WO2003102538A2 - Combinatorial chemical library ii - Google Patents

Combinatorial chemical library ii Download PDF

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Publication number
WO2003102538A2
WO2003102538A2 PCT/US2003/016898 US0316898W WO03102538A2 WO 2003102538 A2 WO2003102538 A2 WO 2003102538A2 US 0316898 W US0316898 W US 0316898W WO 03102538 A2 WO03102538 A2 WO 03102538A2
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WO
WIPO (PCT)
Prior art keywords
amino acid
side chain
alkylene
naturally occurring
independently
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PCT/US2003/016898
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French (fr)
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WO2003102538A3 (en
Inventor
Luis Serrano
Cayetano Gonzales
Luis Bejarano
Original Assignee
European Molecular Biology Laboratory
Williams, Kathleen, M.
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Application filed by European Molecular Biology Laboratory, Williams, Kathleen, M. filed Critical European Molecular Biology Laboratory
Priority to AU2003247435A priority Critical patent/AU2003247435A1/en
Publication of WO2003102538A2 publication Critical patent/WO2003102538A2/en
Publication of WO2003102538A3 publication Critical patent/WO2003102538A3/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/64—Cyclic peptides containing only normal peptide links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
    • C07K1/047—Simultaneous synthesis of different peptide species; Peptide libraries
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • J are the same or different and each J is independently CR I T R.2 ;. and
  • n is an integer from 0 to 10;
  • n is an integer from 0 to 10;
  • T 1 and T 2 are the same or different and each T 1 and each T 2 is independently one of the groups having the structures represented by the formulae:
  • p is an integer from 0 to 20;
  • J are the same or different and each J is independently CR l ⁇ R.2. ;
  • X are the same or different and each X is independently O, S, NR 4 or CR 4 R 4 ;
  • A are the same or different and each A is independently O, S or NR '
  • R 4 are the same or different and each R 4 is independently hydrogen, alkyl or a protecting group
  • protecting group refers to a group which reacts selectively with the desired functionality in good yield to give a derivative that is stable to the reactions for which protection is desired and can be selectively removed from the derivative to yield the desired functionality.
  • pharmaceutically acceptable salt refers to a salt prepared from pharmaceutically acceptable non-toxic acids or bases including inorganic or organic acids and bases.
  • inorganic bases include metallic salts made from aluminium, calcium, lithium, magnesium, potassium, sodium, and zinc.
  • Appropriate organic bases may be selected, for example, from N,N- dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumaine (N-methylglucamine), and procaine.
  • G 1 are the same for each compound of the library of the invention and G 2 are the same for each compound of the library of the invention.
  • At least one group D preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups D, except for any group D which is GR ⁇ T 1 or CT ! T 2 , are independently CR*R 2 wherein each R 1 and each R 2 is independently H, R 4 2 NCR 10 2 (CO)NR 4 -, R 4 O(CO)CR 10 2 NR 4 (CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • at least one of R 1 and R 2 is not H for each group D which is CR !
  • Each T 1 and each T 2 is independently one of the groups having the structures represented by the formulae:
  • each R is H.
  • p is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20.
  • p is not 0. More preferably, p is 4, 5, 6, 7 or 8. Even more preferably, p is 5, 6 or 7, particularly 6.
  • R 3 is NR 4 or O, preferably NR 4 , particularly NH, and when T 1 or T 2 is the group having the structure represented by the formula
  • R is preferably -CX-, more preferably -CO-. This embodiment is particularly preferred when at least one Y is not an amino acid residue mimetic.
  • R 3 is a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • R 3 is a divalent radical of a side chain of a naturally occurring amino acid. This embodiment is particularly preferred when all Y are amino acid residue mimetics, for example -(CX)DA- or -AD(CX)-.
  • each T and each T is independently one of the groups having the structures represented by the formulae:
  • each R is H.
  • L 2 are the same or different and each L 2 is independently CR ⁇ 2 .
  • T 1 and T 2 are each independently one of the groups having the structures represented by the formula:
  • u is an integer from 0 to 20;
  • v is an integer from 0 to 20;
  • w is 0 or 1 ;
  • L are the same or different and each L is independently CR l ⁇ R>2
  • at least two groups D are independently CHT 1 .
  • one group T 1 is one of the groups having the structures represented by the formula:
  • one group T 1 is the group having the structure represented by the structure:
  • each R is H.
  • At least one group D is CHT 1 wherein T 1 is the group having the structure represented by the formula:
  • At least two groups D are independently CHT 1 , wherein in one group D which is CHT 1 , T 1 is the group having the structure represented by the formula:
  • T 1 is -NHCOCH 2 NH 2 .
  • u isO, 1,2,3,4,5,6,7,8,9,10,11,12,13, 14,15, 16,17, 18, 19 or 20.
  • u is not 0. More preferably, u is 1, 2, 3, 4 or 5, still more preferably 2, 3 or 4, particularly 3.
  • v is 0, 1, 2, 3, 4, 5, 6, 1, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20.
  • u is not 0. More preferably, v is 0, 1, 2, 3 or 4, still more preferably 1, 2 or 3, particularly 2.
  • w is 0 or 1, preferably 1.
  • u + v + w is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, more preferably 2, 3, 4, 5, 6, 7, 8 or 9, still more preferably 3, 4, 5, 6, 1, 8 or 9, still more preferably, 5, 6, 7, 8 or 9, still more preferably, 6, 7 or 8, particularly 7.
  • each group D is independently adjacent to a
  • O S NR 4 group which is O, S or NR and adjacent to another group which is ⁇ 1 ⁇ , ⁇ 1 ⁇ , or ⁇ 1 ⁇ .
  • each group D is separated from the nearest other group D by at least one group which is not D.
  • each group D is separated from the nearest other group D by two groups which are not D.
  • each group which is not D is independently O, S,
  • At least one group J preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups J and particularly all groups J are independently CR J R 2 wherein each R 1 and each R 2 is independently H, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • at least one of R 1 and R 2 is not H for each group J.
  • each R 2 is H and each R 1 is independently a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • each R 2 is H and each R 1 is independently a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
  • each R 2 is H and each R 1 is independently a side chain of a naturally occurring positively charged amino acid or a side chain of a naturally occurring aromatic amino acid.
  • at least one group J preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups J which are C*R !
  • R 2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D-configuration.
  • at least one group J preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups J and particularly all groups D which are C*R !
  • C* is a stereogenic centie are at least 60%, more preferably at least 70%), even more preferably at least 80%>, still more preferably at least 90%>, still more preferably at least 95%>, still more preferably at least 96%o, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99%o C* of L-configuration.
  • C* is substantially homochiral.
  • C* is substantially homochiral in the D-configuration.
  • C* is substantially homochiral in the L-configuration.
  • the group K is CR 1 wherein R 1 is H, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • R 1 is H or a side chain of a naturally occurring amino acid.
  • the group K is C*R J wherein C* is a stereogenic centie.
  • C* is at least 60%), more preferably at least 70%>, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D- configuration.
  • C* is at least 60%), more preferably at least 70%>, even more preferably at least 80%), still more preferably at least 90%>, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%>, still more preferably at least 98%), particularly at least 99%) C* of L-configuration.
  • C* is substantially homochiral.
  • C* is substantially homochiral in the D-configuration.
  • C* is substantially homochiral in the L-configuration.
  • At least one group L 1 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L and particularly all groups L 1 are independently CR ⁇ 2 wherein each R 1 and each R 2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups
  • each R is H and each R 1 is independently a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
  • each R 2 is H and each R 1 is independently a side chain of a naturally occurring positively charged amino acid.
  • at least one group L 1 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L 1 and particularly all groups L 1 which are C*R !
  • R 2 wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%), still more preferably at least 90%>, still more preferably at least 95%>, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%o, particularly at least 99%) C* of D-configuration.
  • at least one group L 1 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L 1 and particularly all groups L 1 which are C*R !
  • At least one group L 2 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L 2 and particularly all groups L 2 are independently CR ! R 2 wherein each R 1 and each R 2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups. Even more preferably, each R 2 is H and each R 1 is independently a side chain of a naturally occurring aromatic amino acid.
  • At least one group L 2 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L 2 and particularly all groups L 2 which are C*R L R 2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99%) C* of L-configuration.
  • C* is substantially homochiral.
  • C* is substantially homochiral in the D- configuration.
  • C* is substantially homochiral in the L-configuration.
  • each compound of the library of the invention or the compound of the invention has independently a combination of groups G and G selected from the combinations shown in table 1 below.
  • G 1 and G 2 are the same for each compound of the library of the invention wherein the combination of G 1 and G 2 is selected from the combinations shown in table 1 above.
  • G 1 is CR ! R 2 wherein R 1 is -CH 2 CH(CH 3 ) 2 and R 2 is H and G 2 is CR ! R 2 wherein R 1 is -CH 2 (C 6 H 5 ) and R 2 is H is particularly preferred when no groups D are CR ⁇ or CT ! T 2 .
  • at least one of groups G 1 and G 2 more preferably both groups G 1 and G 2 which are C*R !
  • R 2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%>, still more preferably at least 90%, still more preferably at least 95%>, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%o, particularly at least 99% C* of D-configuration.
  • groups G 1 and G 2 preferably both groups G 1 and G 2 which are C*R !
  • R 2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99% C* of L-configuration.
  • C* is substantially homochiral.
  • C* is substantially homochiral in the D-configuration.
  • C* is substantially homochiral in the L-configuration.
  • each X is independently O or NR 4 , more preferably O or NH. Particularly, X is O.
  • each A is independently O or NR 4 , more preferably O or NH.
  • A is H.
  • each Y is independently -(CX)DA-, -AD(CX)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups.
  • each Y is independently -(CX)DA-, preferably -(CO)D(NR 4 )-, more preferably -(CO)D(NR 4 )-.
  • each Y is independently a divalent radical of a side chain of a naturally occurring amino acid.
  • each R 1 and each R 2 independently is R 4 2 NCR 10 2 (CO)NR 4 -, R 4 O(CO)CR 10 2 NR 4 (CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R 1 is independently a side chain of a naturally occurring amino acid and each R 2 is H.
  • each R 3 not comprising part of T 1 or T 2 is independently a single bond, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R 3 is independently a single bond or a divalent radical of a side chain of a naturally occurring amino acid.
  • each R 3 comprising part of T 1 or T is independently a single bond, -O-, -S-,
  • a divalent radical of a side chain of a naturally occurring amino acid a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more
  • O protecting groups more preferably a single bond, -O-, -NH-, ⁇ X ⁇ or a divalent radical of a side chain of a naturally occurring amino acid.
  • each R 4 is independently hydrogen or a protecting group. More preferably, each R 4 is hydrogen.
  • R 4 2 NCR 10 2 (CO)NR 4 - is R 4 2 NCH 2 (CO)NR 4 -, more preferably
  • R 4 O(CO)CR 10 2 NR 4 (CO> is R 4 O(CO)CH 2 NR 4 (CO)-, more preferably CH 3 (CO)O(CO)CH 2 NR 4 (CO)-
  • alkyl groups means a branched or unbranched, cyclic or acyclic, saturated or unsaturated (e.g. alkenyl or alkynyl) hydrocarbyl radical.
  • the alkyl group is preferably C 3 to C ⁇ 2 , more preferably C 5 to C 10 , more preferably C 5 to C 7 .
  • the alkyl group is preferably Q to C 16 , more preferably d to C 4 , more preferably methyl.
  • Reference in the present specification to an alkoxy group means an alkyl-O- group.
  • alkoyl group means an alkyl-CO- group.
  • the alkyl groups may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent.
  • Substituents may include halogen atoms and halomethyl groups such as CF 3 and CC1 3 ; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitiogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl,
  • amino includes substituted or unsubstituted amino.
  • halogen means a fluorine, chlorine, bromine or iodine radical, preferably fluorine or chlorine radical.
  • a side chain of a naturally occurring amino acid means one of the side chains of the naturally occurring amino acids shown in table 2 below:
  • a side chain of a naturally occurring amino acid may not be the side chain of Cysteine.
  • a side chain of a naturally occurring amino acid may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent.
  • Substituents may include the side chains of the naturally occurring amino acids, T 1 groups as defined above, halogen atoms and halomethyl groups such as CF 3 and CC1 3 ; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitiogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or
  • Preferred substituents are the T 1 groups, particularly -NHCOCH 2 NH 2 .
  • a particularly preferred substituted side chain of a naturally occurring amino acid is the group having the formula represented by the structure:
  • the alkylene groups may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent.
  • Substituents may include halogen atoms and halomethyl groups such as CF 3 and CC1 3 ; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitrogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitrato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl, pyrrolyl,
  • a divalent radical of a side chain of a naturally occurring amino acid means a divalent radical (or trivalent radical in the case of the side chain of Proline) formed from a monovalent side chain (or divalent radical in the case of the side chain of Proline) of the naturally occurring amino acids set out above by the notional removal of one or more atoms.
  • divalent radicals of a side chain of a naturally occurring amino acid are notionally formed by the removal of H, OH or NH 2 .
  • divalent radicals of the side chains of the naturally occurring amino acids include, but are not limited to those shown in table 3 below:
  • the divalent radicals of the side chain are also attached to the N adjacent to the ⁇ carbon.
  • a divalent radical of a side chain of a naturally occurring amino acid may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent.
  • Substituents may include the side chains of the naturally occurring amino acids, T 1 groups as defined above, halogen atoms and halomethyl groups such as CF and CC1 3 ; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitrogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one
  • Preferred substituents are the T 1 groups, particularly -NHCOCH 2 NH2.
  • a particularly preferred substituted side chain of a naturally occurring amino acid is the group having the formula represented by the structure:
  • the positively charged amino acids are Lysine and Arginine.
  • the aromatic amino acids are Phenylalanine, Tyrosine, Tryptophan and Histidine.
  • L-configuration means the absolute configuration of the ⁇ carbon in the partial-structure of the compounds of the present invention as represented by the following formula:
  • D-configuration means the absolute configuration of the ⁇ carbon in the partial-structure of the compounds of the present invention as represented by the following formula:
  • n is selected from the integers 0 to 10;
  • n is selected from the integers 0 to 10;
  • Q 1 are the same or different and each Q 1 is independently CR ⁇ 2 ;
  • Q 2 are the same or different and each Q 2 is independently CR 1 ;
  • T 1 and T 2 are the same or different and each T 1 and each T 2 is independently one of the groups having the structures represented by the formulae:
  • u is an integer from 0 to 20;
  • w is 0 or 1;
  • L 1 are the same or different and each L 1 is independently CR ! R 2 ;
  • R 3 are the same or different and each R 3 is independently a single bond, alkylene,
  • a , A a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
  • R 4 are the same or different and each R 4 is independently hydrogen, alkyl or a protecting group
  • R 10 are the same or different and each R 10 is independently hydrogen or a side chain of a naturally occurring amino acid.
  • a preferred compound of the present invention, or pharmaceutically acceptable derivative thereof, has the structure represented by the formula II:
  • n is selected from the integers 0 to 10;
  • n is selected from the integers 0 to 10;
  • r is selected from the integers 0 to 10; Q 1 are the same or different and each Q 1 is independently CR J -R>2.
  • Q 2 are the same or different and each Q 2 is independently CR 1 ;
  • S' is R' or T 1
  • T 1 and T 2 are the same or different and each T 1 and each T 2 is independently one of the groups having the structures represented by the formulae:
  • u is an integer from 0 to 20;
  • w is 0 or 1;
  • L 1 are the same or different and each L 1 is independently CR R 2 ;
  • L are the same or different and each L is independently CR l ⁇ R ⁇ 2' .
  • Y are the same or different and each Y is independently -(CO)Q 1 (NR 4 )-, -(NR ⁇ Q ⁇ CO)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
  • R 3 are the same or different and each R 3 is independently a single bond, alkylene,
  • a divalent radical of a side chain of a naturally occurring amino acid a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
  • R 4 are the same or different and each R 4 is independently hydrogen, alkyl or a protecting group
  • R , R , R and R are the same or different and are independently H or a side chain of a naturally occurring amino acid, wherein the combination of R 5 , R 6 , R 7 and R 8 is selected from the combinations shown in table 4 below:
  • R 5 are the same for each compound of the library of the invention
  • R 6 are the same for each compound of the library of the invention
  • R 7 are the same for each compound of the library of the invention
  • R 8 are the same for each compound of the library of the invention wherein the combination of R 5 , R 6 , R 7 and R s is selected from the combinations shown in table 4 above.
  • S 1 is T 1 .
  • S 2 i • s ⁇ TT>2.
  • S 1 and S 2 are the same for each compound of the library of the invention.
  • T 1 or T 2 is the group having the formula represented by the structure:
  • each R is H.
  • T 1 is the group having the structure represented by the formula:
  • each R 4 is H.
  • one of T 1 or T 2 is the group having the structure represented by the formula:
  • T is the group having the structure represented by the formula:
  • each compound of the library of the invention or the compound of the invention at least one group Q 1 , preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q 1 and particularly all groups Q 1 are independently C ⁇ R 2 wherein each R 1 and each R 2 is independently H, R 4 2 NCR 10 2 (CO)NR 4 -, R 4 O(CO)CR 10 2 NR 4 (CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid
  • each compound of the library of the invention or the compound of the invention at least one group Q 2 , preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q 2 and particularly all groups Q 2 are independently CR ! R 2 wherein each R 1 and each R 2 is independently H, R 4 2 NCR 10 2 (CO)NR 4 -, R 4 O(CO)CR 10 2 NR 4 (CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R 2 is H and each R 1 is independently a side chain of a naturally occurring amino acid.
  • At least one group Q optionally at least two, optionally at least three, optionally at least four, optionally at least five and optionally all groups Q 2 are C*R* wherein C* is a stereogenic centre.
  • at least one group Q 2 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q 2 and particularly all groups Q 2 which are independently C ⁇ R 1 wherein C* is a stereogenic centre are at least 60%>, more preferably at least 70%>, even more preferably at least 80%>, still more preferably at least 90% still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99% C* of D-configuration.
  • At least one group Q 2 preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q 2 and particularly all groups Q 2 which are C*R* wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%), still more preferably at least 95%, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99%> C* of L-configuration.
  • C* is substantially homochiral.
  • C* is substantially homochiral in the D- configuration.
  • C* is substantially homochiral in the L-configuration.
  • any stereogenic carbon atoms between a carbonyl moiety and a nitrogen atom are in the D-configuration.
  • X 1 , X 2 , X 3 and X are the same or different and each X , X , X 3 and X 4 is independently a residue of a naturally occurring amino acid;
  • a "naturally occurring amino acid” means an amino acid selected from those shown in table 5 below:
  • a naturally occurring amino acid may not be Cysteine.
  • the amino acid residues of the compounds of the library are at least 60%, more preferably at least 70%, even more preferably at least 80%>, still more preferably at least 90%, still more preferably at least 95%>, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99%> C* in the D- configuration.
  • a preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
  • R 9 are the same of different and each R 9 is independently a side chain of a naturally occurring amino acid
  • R 5 is a side chain of a naturally occurring amino acid
  • R 7 is a side chain of a naturally occurring amino acid.
  • a particularly preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
  • R 9 are the same of different and each R 9 is independently a side chain of a naturally occurring amino acid
  • R 5 is a side chain of a naturally occurring amino acid
  • R 7 is a side chain of a naturally occurring amino acid.
  • R 5 and R 7 are selected from the combinations shown in table 4 above.
  • R are the same for each compound of the library of the invention and R are the same for each compound of the library of the invention.
  • Preferred compounds of the present invention are the compounds having structures represented by the following formulae:
  • R 9 are the same of different and each R 9 is independently a side chain of a naturally occurring amino acid
  • R 5 is a side chain of a naturally occurring amino acid
  • R 7 is a side chain of a naturally occurring amino acid.
  • Particularly preferred compounds of the present invention are the compounds having structures represented by the following formulae:
  • R 5 is a side chain of a naturally occurring amino acid
  • R 7 is a side chain of a naturally occurring amino acid.
  • Libraries and compounds of the invention maybe prepared by standard solution phase or solid phase methods in which subunits are added stepwise to growing oligomers until a desired oligomer size is reached.
  • Cyclisation may be effected either by end to end coupling of the peptide backbone, side chain to side chain coupling of side chains of the residues or residue side chain to end of peptide backbone coupling (P. Rovero, Homodetic Cyclic Peptide in Solid-Phase Synthesis: A Practical Guide (S.A. Kates and F. Albericio Eds.), Marcel Dekker, Inc., New York, USA (2000), pages 331 - 364).
  • the libraries and of the invention comprise at least one bioactive compound.
  • the compounds of the invention are bioactive.
  • each library having mutually exclusive compounds contained therein, wherein each library possesses a constant portion which is the same for all the compounds contained in the two or more libraries, and a variable portion, the variable portion of each library being constant for the compounds of that library, but variable between the two or more libraries;
  • the steps set out above can be repeated iteratively using different variable or constant portions at each iteration.
  • Such a repetition allows the progressive determination of the most preferred groups for each portion and therefore the most active compound or compounds of the library.
  • the constant and variable portions are selected from the groups D, T , T , J, X, A, Y, E, F, G 1 , G 2 , R 1 , R 2 , R 3 and R 4 , as defined herein.
  • the constant and variable portions are selected from the amino acid residue mimetics -(CX)G 1 A- (e.g. -(CO)CR 5 R 6 NR 4 -) , -(CX)G 2 A- (e.g. -(CO)CR 7 R 8 NR 4 )-, - (CX)DA- (e.g. -(CO)Q 1 NR 4 -), -EDY- (e.g. -NR 4 QT 1 Y-), -YDF- (e.g. -YQT 2 (CO)-), and -(CX)JA- (e.g. -(CO)JNR 4 -).
  • bioactive compounds of the invention have utility in the treatment of one or more of the following: neoplasms, including cancer, brain tumour, glioma, bone tumour, lung tumour, breast tumour, prostate tumour, colon tumour, hemangioma, melanoma, lung, colorectal, breast, pancreas, head and neck and other solid tumours, myeloproliferative disorders, such as leukemia, non-Hodgkin lymphoma, leukopenia, thrombocytopenia, Kaposis' sarcoma, angiogenesis disorders, inflammatory diseases (e.g.
  • allergies and inflammatory bowel disease hematological disease, neutropenia, autoimmune disease, thrombocytopenia, angiogenesis disorders, dermatological disease, ageing, wounds, burns, fibrosis, arthritis, psoriasis, respiratory tract inflammation, asthma, disorders associated with organ transplantation, cardiovascular disease, hypertension, oedema, angina, atherosclerosis, thrombosis, sepsis, disorders relating to aberrant angiogenesis, shock, reperfusion injury, ischemia, central nervous system disease,
  • Alzheimer's disease brain injury and amyotrophic lateral sclerosis, pain, renal disease, diabetes mellitus, osteoporosis, restensosis, heart disease, myeloid cell disorders, lymphoid cell disorders, peripheral vascular disease, coronary artery disease, oedema, thromboembolism, dysmenorrhea, endometiiosis, pre-eclampsia, lung disease, COPD, asthma, bone disease, renal disease, glomerulonephritis, liver disease, Crohn's disease, gastritis, ulcerative colitis, ulcer, immune disorder, autoimmune disease, arthritis, rheumatoid arthritis, psoriasis, epidermolysis bullosa, systemic lupus erythematosus, ankylosing spondylitis, Lyme disease, multiple sclerosis, neurodegeneration, stroke, brain/spinal cord injury, Parkinson's disease, motor neurone disease, neuromuscular disease, ocular
  • a bioactive compound of the invention may alternatively or additionally exhibit one or more of the following activities or effects; promotion of wound healing; effecting biorhythms or circadian cycles or rhythms; effecting the fertility of male or female subjects; effecting the metabolism, catabolism, anabolism, processing, utilization, storage or elimination, of dietary fat, lipid, protein, carbohydrate, vitamins, minerals, co-factors or other nutritional factors or component(s); effecting behavioural characteristics, including, without limitation, appetite, libido, stress, cognition (including cognitive disorders), hormonal or endocrine activity; and, in the case of enzymes, correcting deficiencies of the enzyme and treating deficiency-related diseases.
  • the bioactive compounds of the invention may be useful in treating an infectious disease.
  • Niruses are one example of an infectious agent.
  • viruses include, but are not limited to the following D ⁇ A and R ⁇ A viruses and viral families: Arbovirus, Adenoviridae, Arenaviridae, Arterivirus, Birnaviridae, Bunyaviridae, Caliciviridae, Circoviridae, Coronaviridae, Dengue, EBN, HIV, Flaviviridae, Hepadnaviridae (Hepatitis), Herpesviridae (such as, Cytomegalovirus, Herpes Simplex, He ⁇ es Zoster), Mononegavims (e.g., Paramyxoviridae, Morbillivirus, Rhabdoviridae), Orthomyxoviridae (e.g., Influenza A, Influenza B, and parainfluenza), Papiloma virus, Papovaviridae, Parvoviridae,
  • Viruses falling within these families can cause a variety of diseases or symptoms, including, but not limited to: arthritis, bronchiollitis, respiratory syncytial virus, encephalitis, eye infections (e.g., conjunctivitis, keratitis), chronic fatigue syndrome, hepatitis (A, B, C, E, Chronic Active, Delta), Japanese B encephalitis, Junin, Chikungunya, Rift Valley fever, yellow fever, meningitis, opportunistic infections (e.g., AIDS), pneumonia, Burkitt's Lymphoma, chickenpox, hemorrhagic fever, Measles, Mumps,
  • a bioactive compound of the invention may be used to treat one (or more) of these diseases.
  • Bacteria and fungi are further examples of infectious agents.
  • a bioactive compound having utility in the tieatment of an infection caused by, the following Gram-Negative and Gram-positive bacteria and bacterial families and fungi: Actinomycetales (e.g., Corynebacterium, Mycobacterium, Norcardia), Cryptococcus neoformans, Aspergillosis, Bacillaceae (e.g., Anthrax, Clostiidium), Bacteroidaceae, Blastomycosis, Bordetella, Borrelia (e.g., Borrelia burgdorferi, Brucellosis, Candidiasis, Campylobacter, Coccidioidomycosis, Cryptococcosis, Dermatocycoses, E.
  • Actinomycetales e.g., Corynebacterium, Mycobacterium, Norcardia
  • Cryptococcus neoformans Aspergillosis
  • Enterobacteriaceae Klebsiella, Salmonella (e.g., Salmonella typhi, and Salmonella paratyphi), Serratia, Yersinia), Erysipelothrix, Helicobacter, Legionellosis, Leptospirosis, Listeria, Mycoplasmatales, Mycobacterium leprae, Vibrio cholerae, Neisseriaceae (e.g., Acinetobacter, Gonorrhea, Menigococcal), Meisseria meningitidis, Pasteurellacea Infections (e.g., Actinobacillus, Heamophilus (e.g., Heamophilus influenza type B), Pasteurella), Pseudomonas, Rickettsiaceae, Chlamydiaceae, Syphilis, Shigella spp.,
  • bacterial or fungal families can cause the following diseases or symptoms, including, but not limited to: bacteremia, endocarditis, eye infections (conjunctivitis, tuberculosis, uveitis), gingivitis, opportunistic infections (e.g., AIDS related infections), paronychia, prosthesis-related infections, Reiter's Disease, respiratory tract infections, such as Whooping Cough or Empyema, sepsis, Lyme Disease, Cat-Scratch Disease, Dysentery, Paratyphoid Fever, food poisoning, Typhoid, pneumonia, Gonorrhea, meningitis (e.g., mengitis types A and B), Chlamydia, Syphilis, Diphtheria, Leprosy, Paratuberculosis, Tuberculosis, Lupus, Botulism, gangrene, tetanus, impetigo, Rheumatic Fever, Scarlet Fever, sexually
  • parasitic agents causing diseases include, but not limited to, the following families or class: Amebiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Dourine, Ectoparasitic, Giardiasis, Helminthiasis, Leishmaniasis, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Trichomonas and Sporozoans (e.g., Plasmodium virax, Plasmodium falciparium, Plasmodium malariae and Plasmodium ovale).
  • families or class Amebiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Dourine, Ectoparasitic, Giardiasis, Helminthiasis, Leishmaniasis, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Trichomonas and Sporozoans
  • a bioactive compound of the invention may be used to treat one (or more) of these diseases.
  • a bioactive compound of the invention may be effective in adult and pediatiic oncology including in solid phase tumours/malignancies, locally advanced tumours, human soft tissue sarcomas, metastatic cancer, including lymphatic metastases, blood cell malignancies including multiple myeloma, acute and chronic leukemias, and lymphomas, head and neck cancers including mouth cancer, larynx cancer and thyroid cancer, lung cancers including small cell carcinoma and non-small cell cancers, breast cancers including small cell carcinoma and ductal carcinoma, gastrointestinal cancers including esophageal cancer, stomach cancer, colon cancer, colorectal cancer and polyps associated with colorectal neoplasia, pancreatic cancers, liver cancer, urologic cancers including bladder cancer and prostate cancer, malignancies of the female genital tract including ovarian carcinoma, uterine (including endometiial) cancers, and solid tumour in the ovarian follicle, kidney cancers including renal cell carcinoma, brain cancers including intrinsic brain tumours
  • compositions comprising a bioactive compound of the invention can be administered in therapeutically effective dosages alone or in combination with adjuvant cancer therapy such as surgery, chemotherapy, radiotherapy, thermotherapy, and laser therapy, and may provide a beneficial effect, e. g. reducing tumour size, slowing rate of tumour growth, inhibiting metastasis, or otherwise improving overall clinical condition, without necessarily eradicating the cancer.
  • adjuvant cancer therapy such as surgery, chemotherapy, radiotherapy, thermotherapy, and laser therapy
  • a bioactive compound of the invention could also be administered in therapeutically effective amounts as a portion of an anti-cancer cocktail.
  • An anti-cancer cocktail is a mixture of a bioactive compound of the invention with one or more anti-cancer drugs in addition to a pharmaceutically acceptable carrier for delivery.
  • a bioactive compound of the invention may be used for prophylactic tieatment of cancer.
  • hereditary conditions and/or environmental situations e.g. exposure to carcinogens
  • In vitro models can be used to determine the efficacy of, and/or effective dose, of a bioactive compound of the invention as a potential cancer treatment.
  • tumour cells in Boyden Chamber assays as described in Pilkington et al., Anticancer Res., 17: 4107-9 (1997)
  • angiogenesis assays such as induction of vascularization of the chick chorioallantoic membrane or induction of vascular endothelial cell migration as described in Ribatta et al., Intl. J. Dev. Biol., 40 : 1189-97 (1999) and Li et al., Clin. Exp. Metastasis, 17 : 423-9 (1999), respectively.
  • Suitable tumour cells lines are available, e. g. from American Type Tissue Culture Collection catalogs.
  • a bioactive compound of the invention may have utility in the tieatment of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia
  • Assays for determining the efficacy of a library of the invention, a compound of the library of the invention or a compound of the invention in tieating one of the above-mentioned conditions will be known, or can be readily devised by, persons skilled in the art. Such assays may be in vivo, ex vivo or in vitro assays.
  • a library of the invention, a compound of the library of the invention or a compound of the present invention can be used in assays to test for one or more biological activities. If the compound exhibits activity in a particular assay, it is likely that the compound may useful in treating a disease associated with the biological activity.
  • kits comprising two or more separate libraries of the present invention.
  • a compound of the present invention for use in a method of tieatment, particularly in a method of tieatment of one or more of the conditions mentioned above.
  • the term "tieatment” includes prophylactic treatment.
  • the term “tieating” should be construed accordingly.
  • the therapy is the treatment of bacterial infection.
  • the bacteria may be gram-t ⁇ or gram-.
  • a method of treating a condition comprising administering to a patient in need of such treatment an effective dose of a compound of the present invention.
  • the condition is a bacterial infection.
  • the compounds of the invention will generally be provided in the form of tablets or capsules or as an aqueous solution or suspension.
  • dosage levels used may vary over quite a wide range depending upon the compound used, the severity of the symptoms exhibited by the patient and the patient's body weight.
  • Figure 1 shows the results of a mortality assay on 3T3 cells using libraries of the invention.
  • Figure 2 shows the results of a mortality assay on oncogenic OCM-1 cells using libraries of the invention.
  • Figure 3 shows the results of a mortality assay on E. coli cells using libraries of the invention.
  • Figure 4 shows a comparison of the mortality assays on 3T3 cells, OCM-1 cells and E. coli cells using libraries of the invention.
  • Figures 5 A and 5B show the concentration effect of the positive and negative control libraries on E. coli growth.
  • Figure 8 shows the percentage of living and dead NIH/3T3 cells after 24 hour culture in the presence of libraries of the invention.
  • Figure 9 shows the results of a mortality assay on Listeria cells using libraries of the invention.
  • Figure 10 shows a comparison of the mortality assays on Listeria cells and E. coli cells using libraries of the invention.
  • Figures 11A - 1 IN show the growth curve of Listeria monocytogenes, Sigella flexneri and E. coli with libraries of the present invention.
  • Figure 12 shows the antibacterial activity against E. coli of a library containing compounds without a tail moiety
  • a library kit consisting of 361 libraries in separate vials, was compiled with each library being synthesised on solid support, with conventional solid phase methods, using orthogonal protecting groups that allow side chain to side chain cyclisation on the resin.
  • each R 01 and R 02 were independently selected from the side chains of the natural llyy ooccccuurrririnngg aammiinnoo aacciiddss eexxcceejpt Cysteine such that R 01 and R 02 were together uniquely defined and constant for each library.
  • R x were variable between the compounds of the library and were randomly selected from the side chains of the naturally occurring amino acids except Cysteine. Accordingly, for each R x , each side chain of the naturally occurring amino acids was present in a substantially equimolar amount.
  • each library vial consists a total of 19 4 different peptides, that is 130231 compounds.
  • NIH/3T3 cells were cultured in MEM supplemented with 10%o foetal bovine serum, ImM L-glutamine, 10 U/ml Penicillin and 10 ⁇ g/ml Streptomycin, at 37 °C in a 5%> CO 2 atmosphere.
  • Example 2 Mortality assay on onco genie OCM-1 cells
  • OCM-l(l,2) Malignant coroid human cell medium was cultured in RPMI supplemented with 5% foetal bovine serum, 10 U/ml Penicillin and 10 ⁇ g/ml Streptomycin, at 37 °C in a 5%> CO2 atmosphere.
  • Figures 1 and 2 summarise the effect that the different libraries had on eukaryotic cell viability.
  • all libraries having peptides with a positively charged residue, Lys or Arg, at any of the two defined positions induce cell death, while having negative residues, Asp and Glu, do not compromise cell viability.
  • aromatic residues, Tyr, Phe and T ⁇ tend to have on average higher mortality rate than hydrophobic or neutral polar residues. It is interesting to note that a much higher mortality rate is observed in the cancer cell line (OCM-1), than in NIH/3T3 cells.
  • Example 3 Mortality assay on E. coli cells
  • the numbers are assigned in the same way as in examples 1 - 3. For eukaryotic cells the numbering goes from 1 (no effect) to 5 (all dead). In the case of bacteria the numbers indicate cell culture OD (0 for no growth and 10 (denoted as #) for maximum growth).
  • the density of the cell culture was monitored every hour for a total period of 12 hours using a Tecan Spectia (TECAN Deutschland GmbH) and Delta Soft III software Ver 2.24 (Dr Bechtold and Biometallics Inc.) by following the OD of the cell culture at 600 nm.
  • FIGS 5 A and 5B The results of the analysis are shown in figures 5 A and 5B.
  • the Ref+ lines show the growth curve of the contiol without peptide and the Ref- lines show the control without bacteria.
  • Figures 5 A and 5B show that the positive control library Arg-Leu prevents bacteria growth at all concentiations tested, while the negative control library Asp-Asp does not affect growth at all concentiations tested.
  • Example 6 Titration effect of the library Leu-Phe on E. coli growth
  • the results of the analysis are shown in figure 6.
  • the Ref+ line shows the growth curve of the contiol without peptide and the Ref- line shows the contiol without bacteria.
  • Figure 6 demonstrates the concentration effect on the inhibition effect of the Leu-Phe library. It can be seen that the library is still active at very low active concentiations, for example 2.5 nM, and completely abolishes bacterial growth at 7 nM.
  • FIGS 7 A - 7N The results of the analysis are shown in figures 7 A - 7N.
  • the Ref+ line shows the growth curve of the control without peptide and the Ref- line shows the control without bacteria, hi each figure, the left graph shows the initial bacterial growth, while the right one shows the bacterial growth after washing and dilution. If the cells have acquired resistance, a normal growth curve should be observed on the right graph. However, if the problem is peptide degradation, we should see the same growth pattern in both the left and right graphs.
  • the behaviour of the cell culture with a peptide dose allowing partial cell growth is a repetition of the first incubation. Accordingly, bacterial growth after long incubation periods can be attributed to peptide degradation.
  • a cell culture of 3T3 cells were infected with 10 ⁇ l of an overnight culture of E. coli diluted 1000 times. At intervals of 3, 6, 9 and 12 hours, an aliquot of the cell medium was taken, diluted 1000 times and plated on Agar plated. The plates were incubated for 12 hours at 37 °C and the number of colonies counted. Cell mortality was also examined through visual inspection of the cells. In all cases, except with the Arg-Leu library that killed both bacteria and cells, and in the other contiol, library Asp- Asp, that did not prevent bacterial growth, the cells looked healthy and viable and could be cultured longer. The results of this analysis are shown in table 10 below:
  • Table 10 Relative bactericidal activity of the active selected libraries. For some of the libraries, for example, Ile-Val, Leu-Gly, Tyr-Gly, it was found that the bacteria start growing after a certain period of time, suggesting cell division arrest. In other libraries, some growing, followed by a decline in the number of viable cells, was observed, suggesting cell death. It therefore appears that different peptides prevent bacterial death by different mechanisms.
  • Example 9 3T3 cell survival in the presence of the active antibiotic peptides.
  • NIH/3T3 cells were cultured in MEM supplemented with 10% ⁇ foetal bovine serum, 1 mM L-glutamine, 10 U/ml Penicillin and 10 ⁇ g/ml Streptomycin, at 37 °C in a 5 % CO2 atmosphere. The day before the experiment, the cells were spread on a 96 glass bottom well plate at 30%) confluency and 150 ⁇ l of media per well. The media was removed and fresh media was added (150 ⁇ l/well). Then, 15 ⁇ l of a selected vial was added to reach a final active peptide concentration of 14 nM.
  • the Live/Dead assay was made using the LIVE/DEAD Viability/Cytotoxicity Kit (L3224) from Molecular Probes, following the protocol supplied by the manufacturer.
  • the cells were incubated in a 2 ⁇ M Calcein AM and 4 ⁇ M Ethidium homodimer-1 in D-PBS for 20 min at room temperature. Then the cells were observed using a DMIRBE Leica photomicroscope and photographed with a COHU high performance CCD camera.
  • As a negative control (dead culture) one of the wells was treated with 3.7%> formaldehyde for 20 min at room temperature, prior to the staining with the Live/Dead assay.
  • the antibiotic properties of the selected libraries were tested on the pathogenic strains Sigella flexneri SC301, Listeria monocytogenes 10403s and E. coli.
  • Example 12 Comparison of mortality assays on Listeria and E. coli
  • Figure 10 shows a comparison of the mortality effects produced using a total of 5 ⁇ l of each selected peptide library at a concentration of 14 nM on E. coli (right columns) after 12 hours and on Listeria (left columns) after 18 hours.
  • the cell density was measured by recording the absorbance at 600 nm after 18 hours.
  • 6 groups were made, ranging from 10 to 0. "10" corresponds to the OD of the control without peptide and "1" to that after incubating the cells with Kanamycin (30 ⁇ g/ml), Ampicillin (100 ⁇ g/ml), Tetiacyclin (12.5 ⁇ g/ml) or Streptomycin (25 ⁇ g/ml).
  • Example 13 Titration effect of selected libraries on prokaryote growth
  • R 01 is the side chain of the amino acid Leucine and R 02 is the side chain of the amino acid Phenylalanine.

Abstract

A chemical library comprising two or more different cyclic peptide mimetic compounds each independently having a structure represented by the following formula is disclosed. Individual compounds from which the library is comprised are also disclosed. The chemical library is useful in screening of pharmaceutically active compounds, in particular antibacterial compounds.

Description

COMBINATORIAL CHEMICAL LIBRARY II
Field of the Invention
The present invention relates to a combinatorial cyclic peptide library and a method of using a combinatorial cyclic peptide library for screening for pharmaceutically active compounds. The present invention further relates to compounds obtainable from such a library.
Background of the Invention
The use of combinatorial libraries of random-sequence oligonucleotides, polypeptides or synthetic oligomers to generate and search for pharmaceutically active compounds and the advantages thereof are widely known in the art (A. Kramer, Pept Res, 6(6): 314 (1993); R.A. Houghten, NIDA Res Monograph, 134: 66 (1993); R.A. Houghten, Gene, 137(1):7 (1993); R.A. Houghten, BioorgMed Chem Lett, 3:405 - 412 (1993); R.A. Houghten, Biotechniques, 13(3):412 (1992); R.A. Houghten, Nature, 354: 84 - 86 (1991); M.H. Ohlmayer, Proc Nat Acad Sci, 90(23): 10922 (1993); CT. Dooley, Proc Nat Acad Sci, 90(22):10811 (1993); CT. Dooley, Life Sci, 52(18): 1509 (1993); J. Eichler, Biochemistry, 32(41): 11035 (1993); C Pinilla, Gene, 128(1):71 (1993), C Pinilla, Biotechniques, 13(6):901 (1992).
However, there is a present and ongoing need for new therapies against diseases and infections.
Summary of the Invention
According to an aspect of the present invention, there is provided a library comprising two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula I:
Figure imgf000003_0001
formula I
wherein:
n is an integer from 0 to 10;
m is an integer from 0 to 10;
r is an integer from 0 to 10;
D are the same or different and each D is independently CRXR2, CR T1 or CT!T2;
T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000003_0002
Figure imgf000003_0003
Figure imgf000004_0001
wherein
p is an integer from 0 to 20;
q is an integer from 0 to 20;
J are the same or different and each J is independently CR IT R.2 ;. and
K is CR1;
X are the same or different and each X is independently O, S, NR or CR τ R>4.
A are the same or different and each A is independently O, S or NR4;
Y are the same or different and each Y is independently R
E is R3
F is R3;
G1 and G2 are the same or different and are independently CR^2, wherein the combination of G1 and G2 is selected from the combinations shown in table 1 below:
Figure imgf000004_0002
Figure imgf000005_0001
Table 1 R1 and R2 are the same or different and each R1 and each R2 is independently hydrogen, alkyl, alkoyl, alkoxy, carboxy, hydroxy, amino, amido, cyano, nitro, thio, sulphonyl, sulphoxide, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R and R attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R are the same or different and each R is independently a single bond, alkylene, O S NR4
-0-, -S-, -N(R4)-, X Λ ,XA , XA , -(CX)DA-, -AD(CX)-, -C(O)O-, -OC(O)-, -S(O)-, -S(O)2- , -C(O)N(R4)-, -N(R4)C(O)-, -OC(O)N(R4)-, -N(R4)C(O)O-, -C(S)N(R4)-, -N(R4)C(S)-, -S(O)2N(R4)-5 -N(R4)S(O)2-5 -N(R4)CON(R4)-, -N(R4)CSN(R4)-,
-O-alkylene-, -S-alkylene-, ~N(R4)-alkylene-,-C(O)-alkylene-, -C(S)-alkylene-, -C(NR4)- alkylene-,-(CX)DA-alkylene-, -AD(CX)-alkylene-, -C(O)O-alkylene-, -OC(O)-alkylene-, -S(O)-alkylene-, -S(O)2-alkylene-, -C(O)N(R4)-alkylene-, -N(R4)C(O)-alkylene-, -OC(O)N(R4)-alkylene-, -N(R4)C(O)O-alkylene-, -C(S)N(R4)-alkylene-, -N(R4)C(S)-alkylene-, -S(O)2N(R4)-alkylene-, -N(R4)S(O)2-alkylene-,
-N(R4)CON(R4)-alkylene-5 -N(R4)CSN(R4)-alkylene-, -alkylene-O-, -alkylene-S-, -alkylene-N(R4)-, -alkylene-C(O)-, -alkylene-C(S)-, -alkylene-C(NR4)-, -alkylene-(CX)DA-, -alkylene-AD(CX)-, -alkylene-C(O)O-, -alkylene-OC(O)-, -alkylene-S(O)-, -alkylene-S(O) -, -alkylene-C(O)N(R4)-, -alkylene-N(R4)C(O)-, -alkylene-OC(O)N(R4)-, -alkylene-N(R4)C(O)O-, -alkylene-C(S)N(R4)-, -alkylene-N(R4)C(S)-, -alkylene-S(O)2N(R4)-, -alkylene-N(R4)S(O)2-, -alkylene-N(R4)CON(R4)-, -alkylene-N(R4)CSN(R4)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group; and R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
Preferably, the library contains at least 100, more preferably at least 1000, even more preferably at least 10,000 and particularly at least 100,000 different compounds, or zwitterions or salts thereof. Particularly, the library contains all the possible different compounds represented by formula I or zwitterions or salts thereof.
It will be appreciated that each of the compounds of the library may independently be in salt, zwitterion or non-ionic form. In aqueous solution, the form of each compound of the library will depend on the pH of the solution and the acidity or basicity of the functional groups present in the compounds of the library.
According to a further aspect of the present invention, there is provided a compound, or pharmaceutically acceptable derivative thereof, having the structure represented by the formula I:
Figure imgf000007_0001
formula I
wherein:
n is an integer from 0 to 10;
m is an integer from 0 to 10;
r is an integer from 0 to 10;
D are the same or different and each D is independently CR l'τR>22, C.τR. l'τT-11 o „„r / C-nT-l 2;. T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000008_0001
Figure imgf000008_0002
wherein
p is an integer from 0 to 20;
q is an integer from 0 to 20;
J are the same or different and each J is independently CR lτ R.2. ; and
K is CR
X are the same or different and each X is independently O, S, NR4 or CR4R4;
A are the same or different and each A is independently O, S or NR'
Y are the same or different and each Y is independently R ;
E is R3 F is R3;
G1 and G2 are the same or different and are independently CR'R2, wherein the combination of G1 and G2 is selected from the combinations shown in table 1 below:
Figure imgf000009_0001
Figure imgf000010_0001
Table 1
R1 and R2 are the same or different and each R1 and each R2 is independently hydrogen, alkyl, alkoyl, alkoxy, carboxy, hydroxy, amino, amido, cyano, nitro, thio, sulphonyl, sulphoxide, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO>, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R and R attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R are the same or different and each R is independently a single bond, alkylene, O S NR4
-O-, -S-, -N(R 44)-, X AX , X AX , X AX t -(CX)DA-, -AD(CX)-, -C(O)O-, -OC(O)-, -S(O)-, -S(O)2- , -C(O)N(R4)-, -N(R4)C(O)-, -OC(O)N(R4)-, -N(R4)C(O)O-, -C(S)N(R4)-, -N(R4)C(S)-, -S(O)2N(R4)-, -N(R4)S(O)2-, -N(R4)CON(R4)-, -N(R4)CSN(R4)-, -O-alkylene-, -S-alkylene-, -N(R4)-alkylene-,-C(O)-alkylene-, -C(S)-alkylene-, -C(NR4)- alkylene-,-(CX)DA-alkylene-, -AD(CX)-alkylene-, -C(O)O-alkylene-, -OC(O)-alkylene-, -S(O)-alkylene-, -S(O)2-alkylene-, -C(O)N(R4)-alkylene-, -N(R4)C(O)-alkylene-, -OC(O)N(R4)-alkylene-, -N(R4)C(O)O-alkylene-, -C(S)N(R4)-alkylene-, -N(R4)C(S)-alkylene-, -S(O)2N(R4)-alkylene-, -N(R4)S(O)2-alkylene-, -N(R4)CON(R4)-alkylene-, -N(R4)CSN(R4)-alkylene-, -alkylene-O-, -alkylene-S-, -alkylene-N(R4)-, -alkylene-C(O)-, -alkylene-C(S)-, -alkylene-C(NR4)-, -alkylene-(CX)DA-, -alkylene-AD(CX)-, -alkylene-C(O)O-, -alkylene-OC(O)-, -alkylene-S(O)-, -alkylene-S(O)2-, -alkylene-C(O)N(R4)-, -alkylene-N(R4)C(O)-, -alkylene-OC(O)N(R4)-, -alkylene-N(R4)C(O)O-, -alkylene-C(S)N(R4)-, -alkylene-N(R4)C(S)-, -alkylene-S(O)2N(R4)-, -alkylene-N(R4)S(O)2-, -alkylene-N(R4)CON(R4)-, -alkylene-N(R4)CSN(R4)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group; and
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
It will be appreciated that the compounds of the present invention may independently be in salt, zwitterion or non-ionic form. In aqueous solution, the form of each compound will depend on the pH of the solution and the acidity or basicity of the functional groups present in the compounds.
As used herein, the term "protecting group" refers to a group which reacts selectively with the desired functionality in good yield to give a derivative that is stable to the reactions for which protection is desired and can be selectively removed from the derivative to yield the desired functionality.
Suitable protecting groups will be readily apparent to the skilled person and may be found in, for example, Kocienski, Protecting Groups, Thieme, New York, 1994. For example, in the case of hydroxyl groups, suitable protecting groups may include esters, ethers (e.g. silyl ethers or alkyl ethers) or acetals. Some specific examples of typical hydroxyl protecting groups are allyl, Aloe, benzyl, BOM, t-butyl, trityl, TBS, TBDPS, TES, TMS, TIPS, PMB, MEM, MOM, MTM, and THP. In the case of nitrogen atoms, suitable protecting groups may include Boc, Aloe, Troc, benzyl, allyl, Fmoc or silyl. In the case of carboxylic acids, suitable protecting groups may include esters (e.g. benzyl, allyl, methyl or ethyl esters).
The term "pharmaceutically acceptable derivative" as used herein, means any pharmaceutically acceptable salt, zwitterion addition compound, or any other compound which upon administration to a recipient is capable of providing, whether directly or indirectly, a compound of the invention or a pharmaceutically acceptable metabolite. The term "pharmaceutically acceptable metabolite" as used herein, means a metabolite or residue of a compound of the invention which gives rise to a biological activity exhibited by the compounds of the invention.
The term "pharmaceutically acceptable salt", as used herein, refers to a salt prepared from pharmaceutically acceptable non-toxic acids or bases including inorganic or organic acids and bases.
Examples of such inorganic acids are hydrochloric, hydrobromic, hydroiodic, sul uric, and phosphoric acids. Appropriate organic acids may be selected, for example, from aliphatic, aromatic, carboxylic and sulfonic classes of organic acids, examples of which are formic, acetic, propionic, succinic, glycolic, glucuronic, maleic, furoic, glutamic, benzoic, anthranilic, salicylic, phenylacetic, mandelic, embonic (pamoic), methanesulfonic, ethanesulfonic, pantothenic, benzenesulfonic, stearic, sulfanilic, algenic, and galacturonic. Examples of such inorganic bases include metallic salts made from aluminium, calcium, lithium, magnesium, potassium, sodium, and zinc. Appropriate organic bases may be selected, for example, from N,N- dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumaine (N-methylglucamine), and procaine.
As used herein, the term "zwitterion" means an ion carrying both a positive and negative charge, for example Hs ^C^COO".
Preferred Libraries
Optionally, at least one of n, m, r, D, T1, T2, J, K, X, A, Y, E, F, G1 or G2 may be the same for each compound of the library of the invention.
In a preferred embodiment, n are the same for each compound of the library of the invention, m are the same for each compound of the library of the invention and r are the same for each compound of the library of the invention.
In another preferred embodiment, T1 are the same for each compound of the library of the invention and T2 are the same for each compound of the library of the invention. In another preferred embodiment, A are the same for each compound of the library of the invention and X are the same for each compound of the library of the invention.
In another preferred embodiment, E are the same for each compound of the library of the invention, F are the same for each compound of the library of the invention and Y are the same for each compound of the library of the invention.
In a more preferred embodiment, G1 are the same for each compound of the library of the invention and G2 are the same for each compound of the library of the invention.
hi a particular preferred embodiment, n are the same for each compound of the library of the invention, m are the same for each compound of the library of the invention, r are the same for each compound of the library of the invention, T1 are the same for each compound of the library of the invention, T2 are the same for each compound of the library of the invention, J are the same for each compound of the library of the invention, K are the same for each compound of the library of the invention, X are the same for each compound of the library of the invention, A are the same for each compound of the library of the invention, Y are the same for each compound of the library of the invention, E are the same for each compound of the library of the invention, F are the same for each compound of the library of the invention, G1 are the same for each compound of the library of the invention and G are the same for each compound of the library of the invention are the same for each compound of the library of the invention.
Preferred Functionalities
The following represent the preferred functionalities of each compound of the library of the invention or the compound of the invention.
n is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. More preferably n is 0, 1, 2, 3 or 4. Even more preferably n is 1, 2 or 3, particularly 2.
m is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. More preferably m is 0, 1, 2, 3 or 4. Even more preferably m is 1, 2 or 3, particularly 2.
r is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. More preferably r is 0, 1, 2, 3 or 4. Even more preferably r is 1, 2 or 3, particularly 2. Preferably, n + m + r is 1, 2, 3, 4, 5, 6, 7, 8 or 9, more preferably 2, 3, 4, 5, 6, 7 or 8, still more preferably, 4, 5, 6, 7 or 8, still more preferably 5, 6 or 7, most preferably 6.
Preferably, at least one group D is CR1!"1 or CI^T2, preferably CR1!1. More preferably, two groups D are each independently CR1!1 or CT1 2, preferably CR1^. Without prejudice to the scope of the invention, it is believed that eukaryotic membrane penetration is facilitated by the presence of tail moiety T1 or T2.
Preferably, at least one group D, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups D, except for any group D which is GR^T1 or CT!T2, are independently CR*R2 wherein each R1 and each R2 is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. Preferably, at least one of R1 and R2 is not H for each group D which is CR!R2. Preferably, each R2 is H and each R1 is independently R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid
1 0 protected by one or more protecting groups for each group D which is CR R . Even more preferably, each R is H and each R is independently a side chain of a naturally occurring amino acid for each group D which is C ^R2.
Preferably, at least one group D, preferably at least two, more preferably at least three, even more preferably at least four and still more preferably at least five and particularly all groups D which are independently CR R , except for any group D which is CH2 or any group D in which R1 = R2, are C*R1R2 wherein C* is a stereogenic centre. Preferably, at least one group D, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups D which are C*R1R2 wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D-configuration. Alternatively, at least one group D, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups D which are C*R1R2 wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, any group D which is CR!T! or CTΨ is CRV. Preferably, each R1 is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R1 is independently H or a side chain of a naturally occurring amino acid. Particularly, each R1 is H.
Preferably, any group D which is CR'T1 or CT1T2, except for any group in which R1 = T1 or T1 = T2, is C^R'T1 or C*T!T2 wherein C* is a stereogenic centre. Preferably, the at least one group D which is C*RLT or CTlT2 wherein C* is a stereogenic centre is at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D-configuration. Alternatively, the at least one group D which is C*R1T1 or CT1"!2 wherein C* is a stereogenic centre is at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%), still more preferably at least 97%>, still more preferably at least 98%, particularly at least 99%o C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000016_0001
Figure imgf000016_0002
Preferably each R is H.
p is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. Preferably, p is not 0. More preferably, p is 4, 5, 6, 7 or 8. Even more preferably, p is 5, 6 or 7, particularly 6.
q is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. Preferably, p is not 0. More preferably, p is 4, 5, 6, 7 or 8. Even more preferably, p is 5, 6 or 7, particularly 6.
In one embodiment of the present invention, when T or T is the group having the structure represented by the formula
Figure imgf000016_0003
R3 is NR4 or O, preferably NR4, particularly NH, and when T1 or T2 is the group having the structure represented by the formula
Figure imgf000017_0001
R is preferably -CX-, more preferably -CO-. This embodiment is particularly preferred when at least one Y is not an amino acid residue mimetic.
In another embodiment of the present invention, R3 is a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups. Preferably, R3 is a divalent radical of a side chain of a naturally occurring amino acid. This embodiment is particularly preferred when all Y are amino acid residue mimetics, for example -(CX)DA- or -AD(CX)-.
1 9
Preferably, each T and each T is independently one of the groups having the structures represented by the formulae:
Figure imgf000017_0002
Figure imgf000017_0003
Figure imgf000018_0001
Preferably, each R is H.
Even more preferably, at least one of groups T 1 and T 9 is one of the groups having the structures represented by the formulae:
Figure imgf000018_0002
Figure imgf000018_0003
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1 ;
provided that the sum u + v + w is not greater than 20; L1 are the same or different and each L1 is independently CRJR2;
L2 are the same or different and each L2 is independently CR^2.
Still more preferably all groups T1 and T2 are each independently one of the groups having the structures represented by the formula:
Figure imgf000019_0001
Figure imgf000019_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1 ;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CR*R2;
L are the same or different and each L is independently CR lτ R>2 Particularly, at least two groups D are independently CHT1. Preferably, one group T1 is one of the groups having the structures represented by the formula:
Figure imgf000020_0001
Figure imgf000020_0002
and another group T1 is the group having the structure represented by the structure:
Figure imgf000020_0003
Preferably, in the case above where at least two groups D are CHT1, one group T1 is the group having the structure represented by the structure:
Figure imgf000021_0001
and another is the group represented by the structure:
Figure imgf000021_0002
Preferably, each R is H.
Still more preferably, at least one group D is CHT1 wherein T1 is the group having the structure represented by the formula:
Figure imgf000022_0001
More preferably, at least another group D is CHT1 wherein T1 is -NHCOCH2NH2.
Particularly, at least two groups D are independently CHT1, wherein in one group D which is CHT1, T1 is the group having the structure represented by the formula:
Figure imgf000023_0001
and in another group D which is CHT1, T1 is -NHCOCH2NH2.
uisO, 1,2,3,4,5,6,7,8,9,10,11,12,13, 14,15, 16,17, 18, 19 or 20. Preferably, u is not 0. More preferably, u is 1, 2, 3, 4 or 5, still more preferably 2, 3 or 4, particularly 3.
v is 0, 1, 2, 3, 4, 5, 6, 1, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. Preferably, u is not 0. More preferably, v is 0, 1, 2, 3 or 4, still more preferably 1, 2 or 3, particularly 2.
w is 0 or 1, preferably 1.
Preferably, u + v + w is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, more preferably 2, 3, 4, 5, 6, 7, 8 or 9, still more preferably 3, 4, 5, 6, 1, 8 or 9, still more preferably, 5, 6, 7, 8 or 9, still more preferably, 6, 7 or 8, particularly 7.
X X
Preferably, each F is independently a single bond or <^X, more preferably -^\, particularly -CO-. Preferably, each E is independently a single bond or A, more preferably A, even more preferably NR4, particularly NH.
X
Preferably, no group A is adjacent to another group A and no group -^\ is adjacent to X another group ^\.
1 0
Preferably, each group D, without considering R or R , is independently adjacent to a
O S NR4 group which is O, S or NR and adjacent to another group which is ^ 1 ^ , ^1 ^ , or ^1 ^ .
1
Particularly, each group D, without considering R or R , is independently adjacent a group
which is NR4 and adjacent to another group which is
Figure imgf000024_0001
Preferably, each group D is separated from the nearest other group D by at least one group which is not D. Particularly, each group D is separated from the nearest other group D by two groups which are not D. Preferably, each group which is not D is independently O, S,
Figure imgf000024_0002
other of the two groups which is not D is NR . Preferably, R 4 i •s H.
Preferably, at least one group J, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups J and particularly all groups J are independently CRJR2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. Preferably, at least one of R1 and R2 is not H for each group J. Preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. Still more preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups. Particularly, each R2 is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid or a side chain of a naturally occurring aromatic amino acid.
Preferably, at least one group J, preferably at least two, more preferably at least three, even more preferably at least four and still more preferably at least five and particularly all groups J, except for any group J which is CH2 or any group J in which Ri = R2, are independently C*R!R2 wherein C* is a stereogenic centre. Preferably, at least one group J, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups D and particularly all groups J which are C*R!R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D-configuration. Alternatively, at least one group J, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups J and particularly all groups D which are C*R!R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%), even more preferably at least 80%>, still more preferably at least 90%>, still more preferably at least 95%>, still more preferably at least 96%o, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99%o C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, the group K is CR1 wherein R1 is H, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. Preferably, R1 is H or a side chain of a naturally occurring amino acid. Preferably, the group K is C*RJ wherein C* is a stereogenic centie. Preferably, C* is at least 60%), more preferably at least 70%>, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99% C* of D- configuration. Alternatively, C* is at least 60%), more preferably at least 70%>, even more preferably at least 80%), still more preferably at least 90%>, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%>, still more preferably at least 98%), particularly at least 99%) C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, at least one group L1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L and particularly all groups L1 are independently CR^2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting
1 1 9 groups. Preferably, at least one of R and R is not H for each group L . Preferably, each R is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups. Even more preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid.
Preferably, at least one group L1, preferably at least two, more preferably at least three, even more preferably at least four and still more preferably at least five and particularly all groups L1, except for any group L which is CH2 or any group L1 in which Ri = R2, are independently C*RJR2 wherein C* is a stereogenic centre. Preferably, at least one group L1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L1 and particularly all groups L1 which are C*R!R2 wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%), still more preferably at least 90%>, still more preferably at least 95%>, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%o, particularly at least 99%) C* of D-configuration. Alternatively, at least one group L1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L1 and particularly all groups L1 which are C*R!R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%), still more preferably at least 90%>, still more preferably at least 95%>, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99%o C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, at least one group L2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L2 and particularly all groups L2 are independently CR!R2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups. Even more preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring aromatic amino acid.
Preferably, at least one group L2, optionally at least two, optionally at least three, optionally at least four, optionally at least five and optionally all groups L2, except for any group L2 which is CH2 or any group L2 in which Ri = R2, are independently C*R!R2 wherein C* is a stereogenic centie. Preferably, at least one group L2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L2 and particularly all groups L2 which are C*R1R2 wherein C* is a stereogenic centie are at least 60%>, more preferably at least 70%>, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%>, still more preferably at least 91%, still more preferably at least 98%, particularly at least 99% C* of D-configuration. Alternatively, at least one group L2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups L2 and particularly all groups L2 which are C*RLR2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99%) C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D- configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, each compound of the library of the invention or the compound of the invention has independently a combination of groups G and G selected from the combinations shown in table 1 below.
Particularly, G1 and G2 are the same for each compound of the library of the invention wherein the combination of G1 and G2 is selected from the combinations shown in table 1 above.
The combination in which G1 is CR!R2 wherein R1 is -CH2CH(CH3)2 and R2 is H and G2 is CR!R2 wherein R1 is -CH2(C6H5) and R2 is H is particularly preferred when no groups D are CRΨ or CT!T2.
1 9 1 Preferably, at least one of groups G and G , preferably both groups G and G , except for any group G1 or G2 which is CH2 or any group G1 or G2 in which Ri = R2, are C*R!R2 wherein C* is a stereogenic centre. Preferably, at least one of groups G1 and G2, more preferably both groups G1 and G2 which are C*R!R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%>, still more preferably at least 90%, still more preferably at least 95%>, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%o, particularly at least 99% C* of D-configuration. Alternatively, at least one of groups G1 and G2, preferably both groups G1 and G2 which are C*R!R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99% C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D-configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, each X is independently O or NR4, more preferably O or NH. Particularly, X is O.
Preferably, each A is independently O or NR4, more preferably O or NH. Particularly, A is H.
Preferably, each Y is independently -(CX)DA-, -AD(CX)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups. In one preferred embodiment, each Y is independently -(CX)DA-, preferably -(CO)D(NR4)-, more preferably -(CO)D(NR4)-. In another more preferred embodiment, each Y is independently a divalent radical of a side chain of a naturally occurring amino acid.
Preferably, each R1 and each R2 independently is R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R1 is independently a side chain of a naturally occurring amino acid and each R2 is H.
Preferably, each R3 not comprising part of T1 or T2 is independently a single bond, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R3 is independently a single bond or a divalent radical of a side chain of a naturally occurring amino acid. Preferably, each R3 comprising part of T1 or T is independently a single bond, -O-, -S-,
O NR4
-N(R ,4 ))- t,x """^ 5,x ^^ 1,x ^"^ . , a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more
O protecting groups, more preferably a single bond, -O-, -NH-, ^ X^ or a divalent radical of a side chain of a naturally occurring amino acid.
Preferably, each R4 is independently hydrogen or a protecting group. More preferably, each R4 is hydrogen.
Preferably, each R10 is H.
Preferably, R4 2NCR10 2(CO)NR4- is R4 2NCH2(CO)NR4-, more preferably
H2NCH2(CO)NH-.
Preferably, R4O(CO)CR10 2NR4(CO>, is R4O(CO)CH2NR4(CO)-, more preferably CH3(CO)O(CO)CH2NR4(CO)-
As used herein, reference to alkyl groups means a branched or unbranched, cyclic or acyclic, saturated or unsaturated (e.g. alkenyl or alkynyl) hydrocarbyl radical. Where cyclic, the alkyl group is preferably C3 to Cι2, more preferably C5 to C10, more preferably C5 to C7. Where acyclic, the alkyl group is preferably Q to C16, more preferably d to C4, more preferably methyl. Reference in the present specification to an alkoxy group means an alkyl-O- group. Reference to alkoyl group means an alkyl-CO- group.
The alkyl groups may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent. Substituents may include halogen atoms and halomethyl groups such as CF3 and CC13; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitiogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, pyrrolidinyl, pyrrolinyl, imidazohdinyl, imidazolinyl, pyrazolidinyl, tetiahydrofuranyl, pyranyl, pyronyl, pyridyl, pyrazinyl, pyridazinyl, benzofuranyl, isobenzofuryl, indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl, isoindazolyl, benzopyranyl, coumarinyl, isocoumarinyl, quinolyl, isoquinolyl, naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl, quinoxadinyl, chromenyl, chromanyl, isochromanyl and carbolinyl; alkyl groups, which may themselves be substituted; and aryl groups, which may themselves be substituted, such as phenyl and substituted phenyl. Alkyl includes substituted and unsubstituted benzyl.
As used herein, the term "amino" includes substituted or unsubstituted amino.
As used herein, the term "halogen" means a fluorine, chlorine, bromine or iodine radical, preferably fluorine or chlorine radical.
As used herein the term "a side chain of a naturally occurring amino acid" means one of the side chains of the naturally occurring amino acids shown in table 2 below:
Figure imgf000031_0001
Figure imgf000032_0001
Figure imgf000033_0001
Table 2
In the case of Proline, the side chain is also attached to the N adjacent to the α carbon.
Optionally, a side chain of a naturally occurring amino acid may not be the side chain of Cysteine.
A side chain of a naturally occurring amino acid may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent. Substituents may include the side chains of the naturally occurring amino acids, T1 groups as defined above, halogen atoms and halomethyl groups such as CF3 and CC13; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitiogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, pyrrolidinyl, pyrrolinyl, imidazohdinyl, imidazolinyl, pyrazolidinyl, tetiahydrofuranyl, pyranyl, pyronyl, pyridyl, pyrazinyl, pyridazinyl, benzofuranyl, isobenzofuryl, indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl, isoindazolyl, benzopyranyl, coumarinyl, isocoumarinyl, quinolyl, isoquinolyl, naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl, quinoxadinyl, chromenyl, chromanyl, isochromanyl and carbolinyl; alkyl groups, which may themselves be substituted; and aryl groups, which may themselves be substituted, such as phenyl and substituted phenyl. Alkyl includes substituted and unsubstituted benzyl.
Preferred substituents are the T1 groups, particularly -NHCOCH2NH2. A particularly preferred substituted side chain of a naturally occurring amino acid is the group having the formula represented by the structure:
Figure imgf000034_0001
As used herein, reference to alkylene groups means a branched or unbranched, cyclic or acyclic, saturated or unsaturated (e.g. alkenylene or alkynylene) hydrocarbylene radical. Where cyclic, the alkylene group is preferably C3 to Cu, more preferably C5 to C10, more preferably C5 to C7. Where acyclic, the alkylene group is preferably Q to Cι6, more preferably Q to C4, more preferably methylene.
The alkylene groups may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent. Substituents may include halogen atoms and halomethyl groups such as CF3 and CC13; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitrogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitrato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, pyrrolidinyl, pyrrolinyl, imidazohdinyl, imidazolinyl, pyrazolidinyl, tetiahydrofuranyl, pyranyl, pyronyl, pyridyl, pyrazinyl, pyridazinyl, benzofuranyl, isobenzofuryl, indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl, isoindazolyl, benzopyranyl, coumarinyl, isocoumarinyl, quinolyl, isoquinolyl, naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl, quinoxadinyl, chromenyl, chromanyl, isochromanyl and carbolinyl; alkyl groups, which may themselves be substituted; and aryl groups, which may themselves be substituted, such as phenyl and substituted phenyl.
As used herein the term "a divalent radical of a side chain of a naturally occurring amino acid" means a divalent radical (or trivalent radical in the case of the side chain of Proline) formed from a monovalent side chain (or divalent radical in the case of the side chain of Proline) of the naturally occurring amino acids set out above by the notional removal of one or more atoms. Typically, divalent radicals of a side chain of a naturally occurring amino acid are notionally formed by the removal of H, OH or NH2.
Examples of divalent radicals of the side chains of the naturally occurring amino acids include, but are not limited to those shown in table 3 below:
Figure imgf000035_0001
Figure imgf000036_0001
35
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Table 3
In the case of Proline, the divalent radicals of the side chain are also attached to the N adjacent to the α carbon.
A divalent radical of a side chain of a naturally occurring amino acid may be substituted or unsubstituted, preferably unsubstituted. Where substituted, there will generally be 1 to 3 substituents present, preferably 1 substituent. Substituents may include the side chains of the naturally occurring amino acids, T1 groups as defined above, halogen atoms and halomethyl groups such as CF and CC13; oxygen containing groups such as oxo, hydroxy, carboxy, carboxyalkyl, alkoxy, alkoxy, alkoyl, alkoyloxy, aryloxy, aryloyl and aryloyloxy; nitrogen containing groups such as amino, amido, alkylamino, dialkylamino, cyano, azide, nitiato and nitro; sulphur containing groups such as thiol, alkylthiol, sulphonyl and sulphoxide; heterocyclic groups containing one or more, preferably one, heteroatom, such as thienyl, furanyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, pyrrolidinyl, pyrrolinyl, imidazohdinyl, imidazolinyl, pyrazolidinyl, tetiahydrofuranyl, pyranyl, pyronyl, pyridyl, pyrazinyl, pyridazinyl, benzofuranyl, isobenzofuryl, indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl, isoindazolyl, benzopyranyl, coumarinyl, isocoumarinyl, quinolyl, isoquinolyl, naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl, quinoxadinyl, chromenyl, chromanyl, isochromanyl and carbolinyl; alkyl groups, which may themselves be substituted; and aryl groups, which may themselves be substituted, such as phenyl and substituted phenyl. Alkyl includes substituted and unsubstituted benzyl.
Preferred substituents are the T1 groups, particularly -NHCOCH2NH2. A particularly preferred substituted side chain of a naturally occurring amino acid is the group having the formula represented by the structure:
Figure imgf000040_0001
As used herein "the positively charged amino acids" are Lysine and Arginine.
As used herein "the aromatic amino acids" are Phenylalanine, Tyrosine, Tryptophan and Histidine. As used herein, the term "L-configuration" means the absolute configuration of the α carbon in the partial-structure of the compounds of the present invention as represented by the following formula:
Figure imgf000041_0001
and in the case when A is NH and X is O:
Figure imgf000041_0002
As used herein, the term "D-configuration" means the absolute configuration of the α carbon in the partial-structure of the compounds of the present invention as represented by the following formula:
Figure imgf000041_0003
and in the case when A is NH and X is O:
Figure imgf000042_0001
Detailed Description of the Invention
A preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula II:
Figure imgf000042_0002
formula II
wherein:
n is selected from the integers 0 to 10;
m is selected from the integers 0 to 10;
r is selected from the integers 0 to 10;
Q1 are the same or different and each Q1 is independently CR^2; Q2 are the same or different and each Q2 is independently CR1 ;
S' is ROr T
S2 is R2 or T2;
T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000043_0001
Figure imgf000043_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CR!R2;
L are the same or different and each L is independently CR R ; N are the same or different and each Y is independently -(CO)Q1(ΝR4)-, -(NR4)Q1(CO)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
1 1
R and R are the same or different and each R and each R is independently hydrogen, alkyl, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R1 and R2 attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R3 are the same or different and each R3 is independently a single bond, alkylene,
O NR4
-O-, -S-, -N(R 4 )~, A , A , A , a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group;
R , R , R and R are the same or different and are independently H or a side chain of a naturally occurring amino acid, wherein the combination of R5, R6, R7 and R8 is selected from the combinations shown in table 4 below:
Figure imgf000044_0001
Figure imgf000045_0002
Table 4
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
A preferred compound of the present invention, or pharmaceutically acceptable derivative thereof, has the structure represented by the formula II:
Figure imgf000045_0001
formula II
wherein:
n is selected from the integers 0 to 10;
m is selected from the integers 0 to 10;
r is selected from the integers 0 to 10; Q1 are the same or different and each Q1 is independently CRJ -R>2.
Q2 are the same or different and each Q2 is independently CR1;
S' is R' or T1
S2 is R2 or T2;
T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000046_0001
Figure imgf000046_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CR R2;
L are the same or different and each L is independently CR lτ Rι2' . Y are the same or different and each Y is independently -(CO)Q1(NR4)-, -(NR^Q^CO)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
1 9 1 9
R and R are the same or different and each R and each R is independently hydrogen, alkyl, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R1 and R2 attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R3 are the same or different and each R3 is independently a single bond, alkylene,
-0-, -S-, -N(R )-,
Figure imgf000047_0001
, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group;
R , R , R and R are the same or different and are independently H or a side chain of a naturally occurring amino acid, wherein the combination of R5, R6, R7 and R8 is selected from the combinations shown in table 4 below:
Figure imgf000047_0002
Figure imgf000048_0002
Table 4
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
Preferably, R5 are the same for each compound of the library of the invention, R6 are the same for each compound of the library of the invention, R7 are the same for each compound of the library of the invention and R8 are the same for each compound of the library of the invention wherein the combination of R5, R6, R7 and Rs is selected from the combinations shown in table 4 above.
Preferably, S1 is T1. Preferably, S 2 i •s < TT>2. Particularly, S .11 ; i-s, T TI1 and Γ SI2 i 's r Tp2
Preferably, S1 and S2 are the same for each compound of the library of the invention.
Preferably, in each compound of the library of the invention or the compound of the invention, one of T1 or T2 is the group having the structure represented by the formula:
Figure imgf000048_0001
and the other of T1 or T2 is the group having the formula represented by the structure:
Figure imgf000049_0001
Preferably, each R is H.
More preferably, in each compound of the library of the invention or the compound of the invention, T1 is the group having the structure represented by the formula:
Figure imgf000049_0002
and T2 is the group having the structure represented by the structure:
Figure imgf000049_0003
Preferably, each R4 is H.
Even more preferably, in each compound of the library of the invention or the compound of the invention, one of T1 or T2 is the group having the structure represented by the formula:
Figure imgf000050_0001
and the other of T1 or T2 is -NHCOCH2NH2. Particularly, T is the group having the structure represented by the formula:
Figure imgf000051_0001
and T2 is -NHCOCH2NH2.
Preferably, in each compound of the library of the invention or the compound of the invention, at least one group Q1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q1 and particularly all groups Q1 are independently C^R2 wherein each R1 and each R2 is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid
9 1 protected by one or more protecting groups. Preferably, each R is H and each R is independently R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO , a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. Even more preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring amino acid. Preferably, in each compound of the library of the invention or the compound of the invention, at least one group Q1, optionally at least two, optionally at least three, optionally at least four, optionally at least five and optionally all groups , except for any group Q1 which is CH2 or any group Q1 in which R1 = R2, are independently C*R!R2 wherein C* is a stereogenic centre. Preferably, at least one group Q1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q1 and particularly all groups Q1 which are C*R1R2 wherein C* is a stereogenic centie are at least 60%>, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90 still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99% C* of D-configuration.
Alternatively, at least one group Q1, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q1 and particularly all groups Q1 which are C*R1R2 wherein C* is a stereogenic centie are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%, still more preferably at least 95%, still more preferably at least 96%), still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99%> C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D- configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, in each compound of the library of the invention or the compound of the invention, at least one group Q2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q2 and particularly all groups Q2 are independently CR!R2 wherein each R1 and each R2 is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups. More preferably, each R2 is H and each R1 is independently a side chain of a naturally occurring amino acid.
Preferably, in each compound of the library of the invention or the compound of the
9 • invention, at least one group Q , optionally at least two, optionally at least three, optionally at least four, optionally at least five and optionally all groups Q2 are C*R* wherein C* is a stereogenic centre. Preferably, at least one group Q2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q2 and particularly all groups Q2 which are independently C^R1 wherein C* is a stereogenic centre are at least 60%>, more preferably at least 70%>, even more preferably at least 80%>, still more preferably at least 90% still more preferably at least 95%, still more preferably at least 96%, still more preferably at least 97%o, still more preferably at least 98%>, particularly at least 99% C* of D-configuration. Alternatively, at least one group Q2, preferably at least two, more preferably at least three, even more preferably at least four, still more preferably at least five groups Q2 and particularly all groups Q2 which are C*R* wherein C* is a stereogenic centre are at least 60%, more preferably at least 70%, even more preferably at least 80%, still more preferably at least 90%), still more preferably at least 95%, still more preferably at least 96%>, still more preferably at least 97%, still more preferably at least 98%>, particularly at least 99%> C* of L-configuration. Preferably C* is substantially homochiral. Preferably, C* is substantially homochiral in the D- configuration. Optionally, C* is substantially homochiral in the L-configuration.
Preferably, in each compound of the library of the invention or the compound of the invention, any stereogenic carbon atoms between a carbonyl moiety and a nitrogen atom are in the D-configuration.
A more preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
Figure imgf000053_0001
wherein: X1, X2, X3 and X are the same or different and each X , X , X3 and X4 is independently a residue of a naturally occurring amino acid; and
O1 and O2 are the same of different and each O1 and each O2 is independently a residue of a naturally occurring amino acid.
As used herein, the term a "naturally occurring amino acid" means an amino acid selected from those shown in table 5 below:
Figure imgf000054_0001
Table 5
As used herein, the symbol "Ac" means CH3CO-.
Optionally, a naturally occurring amino acid may not be Cysteine.
Preferably, the amino acid residues of the compounds of the library are at least 60%, more preferably at least 70%, even more preferably at least 80%>, still more preferably at least 90%, still more preferably at least 95%>, still more preferably at least 96%, still more preferably at least 97%, still more preferably at least 98%, particularly at least 99%> C* in the D- configuration.
A preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
Figure imgf000055_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above. A particularly preferred library of the present invention comprises two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
Figure imgf000056_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above. Preferably, R are the same for each compound of the library of the invention and R are the same for each compound of the library of the invention.
Preferred compounds of the present invention are the compounds having structures represented by the following formulae:
Figure imgf000057_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above.
Particularly preferred compounds of the present invention are the compounds having structures represented by the following formulae:
Figure imgf000058_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above.
Libraries and compounds of the invention maybe prepared by standard solution phase or solid phase methods in which subunits are added stepwise to growing oligomers until a desired oligomer size is reached. Cyclisation may be effected either by end to end coupling of the peptide backbone, side chain to side chain coupling of side chains of the residues or residue side chain to end of peptide backbone coupling (P. Rovero, Homodetic Cyclic Peptide in Solid-Phase Synthesis: A Practical Guide (S.A. Kates and F. Albericio Eds.), Marcel Dekker, Inc., New York, USA (2000), pages 331 - 364).
Preferably, the libraries and of the invention comprise at least one bioactive compound.
Preferably, the compounds of the invention are bioactive.
The present invention further extends to a method of determining one or more bioactive compounds of the present invention comprising the steps of:
(i) providing two or more libraries of the invention, each library having mutually exclusive compounds contained therein, wherein each library possesses a constant portion which is the same for all the compounds contained in the two or more libraries, and a variable portion, the variable portion of each library being constant for the compounds of that library, but variable between the two or more libraries;
(ii) independently assaying each library;
(iii) determining the bioactivity of each library.
Preferably, the steps set out above can be repeated iteratively using different variable or constant portions at each iteration. Such a repetition allows the progressive determination of the most preferred groups for each portion and therefore the most active compound or compounds of the library.
Preferably, the constant and variable portions are selected from the groups D, T , T , J, X, A, Y, E, F, G1, G2, R1, R2, R3 and R4, as defined herein.
More preferably, the constant and variable portions are selected from the amino acid residue mimetics -(CX)G1A- (e.g. -(CO)CR5R6NR4-) , -(CX)G2A- (e.g. -(CO)CR7R8NR4)-, - (CX)DA- (e.g. -(CO)Q1NR4-), -EDY- (e.g. -NR4QT1Y-), -YDF- (e.g. -YQT2(CO)-), and -(CX)JA- (e.g. -(CO)JNR4-).
As used herein, the term "bioactive compounds" means compounds of the invention which have utility in the treatment of a condition, preferably one or more of the following: cell proliferative disorders (e.g. cancer), autoimmune disorders, inflammatory disorders, immune system disorders (including allergies), cardiovascular disorders, hormonal disorders, nervous system disorders, psychiatric disorders, developmental disorders, reproductive disorders, genetic disorders, metabolic disorders, infections and other pathological conditions.
Preferably, bioactive compounds of the invention have utility in the treatment of one or more of the following: neoplasms, including cancer, brain tumour, glioma, bone tumour, lung tumour, breast tumour, prostate tumour, colon tumour, hemangioma, melanoma, lung, colorectal, breast, pancreas, head and neck and other solid tumours, myeloproliferative disorders, such as leukemia, non-Hodgkin lymphoma, leukopenia, thrombocytopenia, Kaposis' sarcoma, angiogenesis disorders, inflammatory diseases (e.g. allergies and inflammatory bowel disease), hematological disease, neutropenia, autoimmune disease, thrombocytopenia, angiogenesis disorders, dermatological disease, ageing, wounds, burns, fibrosis, arthritis, psoriasis, respiratory tract inflammation, asthma, disorders associated with organ transplantation, cardiovascular disease, hypertension, oedema, angina, atherosclerosis, thrombosis, sepsis, disorders relating to aberrant angiogenesis, shock, reperfusion injury, ischemia, central nervous system disease,
Alzheimer's disease, brain injury and amyotrophic lateral sclerosis, pain, renal disease, diabetes mellitus, osteoporosis, restensosis, heart disease, myeloid cell disorders, lymphoid cell disorders, peripheral vascular disease, coronary artery disease, oedema, thromboembolism, dysmenorrhea, endometiiosis, pre-eclampsia, lung disease, COPD, asthma, bone disease, renal disease, glomerulonephritis, liver disease, Crohn's disease, gastritis, ulcerative colitis, ulcer, immune disorder, autoimmune disease, arthritis, rheumatoid arthritis, psoriasis, epidermolysis bullosa, systemic lupus erythematosus, ankylosing spondylitis, Lyme disease, multiple sclerosis, neurodegeneration, stroke, brain/spinal cord injury, Parkinson's disease, motor neurone disease, neuromuscular disease, ocular disorder, macular degeneration, glaucoma, diabetic retinopathy, ocular hypertension, viral infections, HIN, AIDS, cytomegalovirus infection, parasitic infections, bacterial infections, fungal infections and protistan infections.
A bioactive compound of the invention may alternatively or additionally exhibit one or more of the following activities or effects; promotion of wound healing; effecting biorhythms or circadian cycles or rhythms; effecting the fertility of male or female subjects; effecting the metabolism, catabolism, anabolism, processing, utilization, storage or elimination, of dietary fat, lipid, protein, carbohydrate, vitamins, minerals, co-factors or other nutritional factors or component(s); effecting behavioural characteristics, including, without limitation, appetite, libido, stress, cognition (including cognitive disorders), hormonal or endocrine activity; and, in the case of enzymes, correcting deficiencies of the enzyme and treating deficiency-related diseases.
The bioactive compounds of the invention may be useful in treating an infectious disease. Niruses are one example of an infectious agent. Examples of viruses, include, but are not limited to the following DΝA and RΝA viruses and viral families: Arbovirus, Adenoviridae, Arenaviridae, Arterivirus, Birnaviridae, Bunyaviridae, Caliciviridae, Circoviridae, Coronaviridae, Dengue, EBN, HIV, Flaviviridae, Hepadnaviridae (Hepatitis), Herpesviridae (such as, Cytomegalovirus, Herpes Simplex, Heφes Zoster), Mononegavims (e.g., Paramyxoviridae, Morbillivirus, Rhabdoviridae), Orthomyxoviridae (e.g., Influenza A, Influenza B, and parainfluenza), Papiloma virus, Papovaviridae, Parvoviridae, Picornaviridae, Poxviridae (such as Smallpox or Vaccinia), Reoviridae (e.g., Rotavirus), Retroviridae (HTLN-I, HTLN-II, Lentivirus), and Togaviridae (e.g., Rubivirus). Viruses falling within these families can cause a variety of diseases or symptoms, including, but not limited to: arthritis, bronchiollitis, respiratory syncytial virus, encephalitis, eye infections (e.g., conjunctivitis, keratitis), chronic fatigue syndrome, hepatitis (A, B, C, E, Chronic Active, Delta), Japanese B encephalitis, Junin, Chikungunya, Rift Valley fever, yellow fever, meningitis, opportunistic infections (e.g., AIDS), pneumonia, Burkitt's Lymphoma, chickenpox, hemorrhagic fever, Measles, Mumps,
Parainfluenza, Rabies, the common cold, Polio, leukemia, Rubella, sexually transmitted diseases, skin diseases (e.g., Kaposi's, warts), and viremia. Accordingly, a bioactive compound of the invention may be used to treat one (or more) of these diseases.
Bacteria and fungi are further examples of infectious agents. Preferably, there is provided a bioactive compound having utility in the tieatment of an infection caused by, the following Gram-Negative and Gram-positive bacteria and bacterial families and fungi: Actinomycetales (e.g., Corynebacterium, Mycobacterium, Norcardia), Cryptococcus neoformans, Aspergillosis, Bacillaceae (e.g., Anthrax, Clostiidium), Bacteroidaceae, Blastomycosis, Bordetella, Borrelia (e.g., Borrelia burgdorferi, Brucellosis, Candidiasis, Campylobacter, Coccidioidomycosis, Cryptococcosis, Dermatocycoses, E. coli (e.g., Enterotoxigenic E. coli and Enterohemorrhagic E. coli), Enterobacteriaceae (Klebsiella, Salmonella (e.g., Salmonella typhi, and Salmonella paratyphi), Serratia, Yersinia), Erysipelothrix, Helicobacter, Legionellosis, Leptospirosis, Listeria, Mycoplasmatales, Mycobacterium leprae, Vibrio cholerae, Neisseriaceae (e.g., Acinetobacter, Gonorrhea, Menigococcal), Meisseria meningitidis, Pasteurellacea Infections (e.g., Actinobacillus, Heamophilus (e.g., Heamophilus influenza type B), Pasteurella), Pseudomonas, Rickettsiaceae, Chlamydiaceae, Syphilis, Shigella spp., Staphylococcal, Meningiococcal, Pneumococcal and Stieptococcal (e.g., Streptococcus pneumoniae and Group B Streptococcus). These bacterial or fungal families can cause the following diseases or symptoms, including, but not limited to: bacteremia, endocarditis, eye infections (conjunctivitis, tuberculosis, uveitis), gingivitis, opportunistic infections (e.g., AIDS related infections), paronychia, prosthesis-related infections, Reiter's Disease, respiratory tract infections, such as Whooping Cough or Empyema, sepsis, Lyme Disease, Cat-Scratch Disease, Dysentery, Paratyphoid Fever, food poisoning, Typhoid, pneumonia, Gonorrhea, meningitis (e.g., mengitis types A and B), Chlamydia, Syphilis, Diphtheria, Leprosy, Paratuberculosis, Tuberculosis, Lupus, Botulism, gangrene, tetanus, impetigo, Rheumatic Fever, Scarlet Fever, sexually transmitted diseases, skin diseases (e.g., cellulitis, dermatocycoses), toxemia, urinary tract infections, wound infections. Accordingly, a bioactive compound of the invention may be used to treat one (or more) of these diseases.
Moreover, parasitic agents causing diseases that may be treated with a bioactive compound of the present invention include, but not limited to, the following families or class: Amebiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Dourine, Ectoparasitic, Giardiasis, Helminthiasis, Leishmaniasis, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Trichomonas and Sporozoans (e.g., Plasmodium virax, Plasmodium falciparium, Plasmodium malariae and Plasmodium ovale). These parasites can cause a variety of diseases or symptoms, including, but not limited to: Scabies, Trombiculiasis, eye infections, intestinal disease (e.g., dysentery, giardiasis), liver disease, lung disease, opportunistic infections (e.g., AIDS related), malaria, pregnancy complications, and toxoplasmosis. Accordingly, a bioactive compound of the invention may be used to treat one (or more) of these diseases.
A bioactive compound of the invention may be effective in adult and pediatiic oncology including in solid phase tumours/malignancies, locally advanced tumours, human soft tissue sarcomas, metastatic cancer, including lymphatic metastases, blood cell malignancies including multiple myeloma, acute and chronic leukemias, and lymphomas, head and neck cancers including mouth cancer, larynx cancer and thyroid cancer, lung cancers including small cell carcinoma and non-small cell cancers, breast cancers including small cell carcinoma and ductal carcinoma, gastrointestinal cancers including esophageal cancer, stomach cancer, colon cancer, colorectal cancer and polyps associated with colorectal neoplasia, pancreatic cancers, liver cancer, urologic cancers including bladder cancer and prostate cancer, malignancies of the female genital tract including ovarian carcinoma, uterine (including endometiial) cancers, and solid tumour in the ovarian follicle, kidney cancers including renal cell carcinoma, brain cancers including intrinsic brain tumours, neuroblastoma, astiocytic brain tumours, gliomas, metastatic tumour cell invasion in the central nervous system, bone cancers including osteomas, skin cancers including malignant melanoma, tumour progression of human skin keratinocytes, squamous cell carcinoma, basal cell carcinoma, hemangiopericytoma and Karposi's sarcoma.
Therapeutic compositions comprising a bioactive compound of the invention can be administered in therapeutically effective dosages alone or in combination with adjuvant cancer therapy such as surgery, chemotherapy, radiotherapy, thermotherapy, and laser therapy, and may provide a beneficial effect, e. g. reducing tumour size, slowing rate of tumour growth, inhibiting metastasis, or otherwise improving overall clinical condition, without necessarily eradicating the cancer.
A bioactive compound of the invention could also be administered in therapeutically effective amounts as a portion of an anti-cancer cocktail. An anti-cancer cocktail is a mixture of a bioactive compound of the invention with one or more anti-cancer drugs in addition to a pharmaceutically acceptable carrier for delivery.
In addition, a bioactive compound of the invention may be used for prophylactic tieatment of cancer. There are hereditary conditions and/or environmental situations (e.g. exposure to carcinogens) known in the art that predispose an individual to developing cancers. Under these circumstances, it may be beneficial to treat these individuals with a therapeutically effective doses of a bioactive compound of the invention to reduce the risk of developing cancers. In vitro models can be used to determine the efficacy of, and/or effective dose, of a bioactive compound of the invention as a potential cancer treatment. These in vitro models include proliferation assays of cultured tumour cells, growth of cultured tumour cells in soft agar (see Freshney, (1987) Culture of Animal Cells: A Manual of Basic Technique, Wily-Liss, New York, NY Ch 18 and Ch 21), tumour systems in nude mice as described in Giovanella et al., J. Natl. Can. Inst., 52 : 921-30 (1974), mobility and invasive potential of tumour cells in Boyden Chamber assays as described in Pilkington et al., Anticancer Res., 17: 4107-9 (1997), and angiogenesis assays such as induction of vascularization of the chick chorioallantoic membrane or induction of vascular endothelial cell migration as described in Ribatta et al., Intl. J. Dev. Biol., 40 : 1189-97 (1999) and Li et al., Clin. Exp. Metastasis, 17 : 423-9 (1999), respectively. Suitable tumour cells lines are available, e. g. from American Type Tissue Culture Collection catalogs.
In view of the foregoing it will be appreciated that a bioactive compound of the invention may have utility in the tieatment of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia
(including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemias (e.g., chronic myelocytic (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphomas (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumours including, but not limited to, sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumour, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumour, cervical cancer, testicular tumour, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, melanoma, neuroblastoma, and retinoblastoma. Assays for determining the efficacy of a library of the invention, a compound of the library of the invention or a compound of the invention in tieating one of the above-mentioned conditions will be known, or can be readily devised by, persons skilled in the art. Such assays may be in vivo, ex vivo or in vitro assays.
For instance, those skilled in the art will appreciate that a library of the invention, a compound of the library of the invention or a compound of the present invention can be used in assays to test for one or more biological activities. If the compound exhibits activity in a particular assay, it is likely that the compound may useful in treating a disease associated with the biological activity.
Those skilled in the art will also appreciate that the efficacy of a library of the invention, a compound of the library of the invention or a compound of the invention for treating a particular condition could be determined by administering a compound to an animal suffering from, or having a predisposition to, a condition of interest. The effect of the compound on the condition may then be determined thus allowing the efficacy of the compound in tieating that condition to be determined.
According to a further aspect of the present invention, there is provided a kit comprising two or more separate libraries of the present invention.
According to a further aspect of the present invention, there is provided a compound of the present invention for use in a method of tieatment, particularly in a method of tieatment of one or more of the conditions mentioned above.
As used herein, the term "tieatment" includes prophylactic treatment. The term "tieating" should be construed accordingly.
The compounds of the present invention may, in particular, be employed in the tieatment of a bacterial infection, more preferably a prokaryotic bacterial infection.
According to a further aspect of the present invention, there is provided the use of a compound of the present invention in the manufacture of a medicament for use in diagnosis or therapy, particularly in the diagnosis or therapy of one or more of the conditions mentioned above.
Preferably, the therapy is the treatment of bacterial infection. The bacteria may be gram-t¬ or gram-.
According to a further aspect of the present invention, there is provided a method of treating a condition comprising administering to a patient in need of such treatment an effective dose of a compound of the present invention. Preferably, the condition is a bacterial infection.
According to a further aspect of the present invention, there is provided a pharmaceutical composition comprising a compound of the present invention in combination with a pharmaceutically acceptable excipient.
The compounds of the present invention maybe administered in a form suitable for oral use, for example a tablet, capsule, aqueous or oily solution, suspension or emulsion; for topical use including transmucosal and transdermal use, for example a cream, ointment, gel, aqueous or oil solution or suspension, salve, patch, plaster or as a component of a lubricant for a condom; for nasal use, for an example a snuff, nasal spray or nasal drops; for vaginal or rectal use, for example a suppository; for administration by inhalation, for example a finely divided powder or a liquid aerosol; for sub-lingual or buccal use, for example a tablet or capsule; or for parenteral use (including intravenous, subcutaneous, intramuscular, intiavascular or infusion), for example a sterile aqueous or oil solution or suspension, or incoφorated in a biodegradable polymer. In general, the above compositions may be prepared in a conventional manner using convention excipients, using standard techniques well known to those skilled in the art of pharmacy. Preferably, the compound is administered orally or topically.
For oral administration, the compounds of the invention will generally be provided in the form of tablets or capsules or as an aqueous solution or suspension.
Tablets for oral use may include the active ingredient mixed with pharmaceutically acceptable excipients such as inert diluents, disintegrating agents, binding agents, lubricating agents, sweetening agents, flavouring agents, colouring agents and preservatives. Suitable inert diluents include sodium and calcium carbonate, sodium and calcium phosphate, and lactose, while corn starch and alginic acid are suitable disintegrating agents. Binding agents may include starch and gelatin, while the lubricating agent, if present, will generally be magnesium stearate, stearic acid or talc. If desired, the tablets may be coated with a material such as glyceryl monostearate or glyceryl distearate, to delay absoφtion in the gastrointestinal tract.
Capsules for oral use include hard gelatin capsules in which the active ingredient is mixed with a solid diluent, and soft gelatin capsules wherein the active ingredient is mixed with water or an oil such as peanut oil, liquid paraffin or olive oil.
For intiamuscular, intraperitoneal, subcutaneous and intravenous use, the compounds of the invention will generally be provided in sterile aqueous solutions or suspensions, buffered to an appropriate pH and isotonicity. Suitable aqueous vehicles mclude Ringer's solution and isotonic sodium chloride. Aqueous suspensions according to the invention may include suspending agents such as cellulose derivatives, sodium alginate, polyvinyl-pyrrolidone and gum tragacanth, and a wetting agent such as lecithin. Suitable preservatives for aqueous suspensions include ethyl and n- propyl p-hydroxybenzoate. The compounds of the invention may also be provided in a biodegradable polymer, for example for use in conjunction with stents in angioplasty (e.g. adsorbed on a stent or applied directly to the site of the procedure for slow release of the active agent).
It will be appreciated that the dosage levels used may vary over quite a wide range depending upon the compound used, the severity of the symptoms exhibited by the patient and the patient's body weight.
Brief Description of the Figures
Figure 1 shows the results of a mortality assay on 3T3 cells using libraries of the invention.
Figure 2 shows the results of a mortality assay on oncogenic OCM-1 cells using libraries of the invention.
Figure 3 shows the results of a mortality assay on E. coli cells using libraries of the invention. Figure 4 shows a comparison of the mortality assays on 3T3 cells, OCM-1 cells and E. coli cells using libraries of the invention.
Figures 5 A and 5B show the concentration effect of the positive and negative control libraries on E. coli growth.
Figure 6 shows the concentration effect of library Leu-Phe on E. coli growth.
Figures 7 A — 7N show the growth curve of bacteria with libraries of the present invention in order to assess the acquisition of resistance by the cell culture to the selected libraries.
Figure 8 shows the percentage of living and dead NIH/3T3 cells after 24 hour culture in the presence of libraries of the invention.
Figure 9 shows the results of a mortality assay on Listeria cells using libraries of the invention.
Figure 10 shows a comparison of the mortality assays on Listeria cells and E. coli cells using libraries of the invention.
Figures 11A - 1 IN show the growth curve of Listeria monocytogenes, Sigella flexneri and E. coli with libraries of the present invention.
Figure 12 shows the antibacterial activity against E. coli of a library containing compounds without a tail moiety
Examples
A library kit, consisting of 361 libraries in separate vials, was compiled with each library being synthesised on solid support, with conventional solid phase methods, using orthogonal protecting groups that allow side chain to side chain cyclisation on the resin.
Each library consisted of compounds of the formula III:
Figure imgf000069_0001
wherein each R01 and R02 were independently selected from the side chains of the natural llyy ooccccuurrririnngg aammiinnoo aacciiddss eexxcceejpt Cysteine such that R01 and R02 were together uniquely defined and constant for each library.
For each library, Rx were variable between the compounds of the library and were randomly selected from the side chains of the naturally occurring amino acids except Cysteine. Accordingly, for each Rx, each side chain of the naturally occurring amino acids was present in a substantially equimolar amount.
As used herein "library A-B" refers to the library in which R01 = the side chain of the naturally occurring amino acid A and R02 = the side chain of the naturally occurring amino acid B. Synthesis of the Library
Automated synthesis was carried out in an Advanced ChemTech 396Ω multiple peptide synthesizer (Louisville, USA) using 9-fluorenylmethyloxycarbonyl/tert-butyl (Fmoc/tBu) strategy. All amino acids used were in the D- configuration. Standard side chain protecting groups were employed for all amino acids except for residues involved in the cyclization step, were orthogonal protecting groups were used. Library was constructed in a positional scanning format (Pinilla, C, Appel, J.R., Blanc, P. & Houghten, R.A. Biotechniques 13, 901-905 (1992)) by randomizing six positions located inside the cycle. Mixture positions were prepared from a set of 19 of the 20 natural coded amino acids, omitting Cys, which was also omitted at the fixed positions.
Deprotection of the orthogonal protecting groups was performed prior to cyclization of the linear sequence on solid support. Final cleavage of the cyclic peptidyl resin yielded the desired cyclic peptide library in solution.
Analysis of the library by MALDI-TOF MS (matrix assisted laser desoφtion ionization mass spectrometiy) techniques provided enough information to ensure the lack of major by-products and the presence of the expected structures in the mixture Furthermore, analysis by amino acid analysis showed that individual amino acids in the mixture positions were present in roughly equal amounts.
A reaction scheme for the synthesis is shown below. The following abbreviations are used:
AM: p-[R,SJ- α-[l -(9H-fluoren-9-yl)-methoxyformamido]-2,4-dimethoxybenzyl]- phenoxyacetic acid
Boc: tert-butyloxycarbonyl
Fmoc/tBu: 9-fluorenylmethyloxycarbonyl/tert-butyl
Pmc: 2,2,4,6,7-pentamethyl-chroman-5-sulfonyl 1) deprotection
Fmoc-AM-NH , Frnoc-Gly-GO-NH-AM-NH
2) Fmoc-Gly-OH
1 ) several deprotection & couplings
2) final acetylation
Ac-arg-arg-arg-trp-phe-Gly-asp-xxx-xxx-ooθ3-ooθ4-xxx-xxx-lys-Gly-CO-NH-AM-NH-
Pm I cPm I c Pm I cBo Ic p I. p I2 _. _„ ,, . , ,.
1 i P1, P2: orthogonal protecting groups
1) P1, P2 deprotection
2) cyclization
3) cleavage from solid support
Ac-arg-arg-arg-trp-p.1e-Gly-asp-xxx-xxx-0003-0004-xxx-xxx-lys-Gly-r.H2
Calculation of Concentration of Active Peptide
In order to compare the biological activities of the active compound of the libraries of the invention with known compounds, it is necessary to have an estimate of the concentration of active compounds in each library.
In each library vial consists a total of 194 different peptides, that is 130231 compounds.
Classifying the amino acids by their chemical similarities, for each R »x . there are six different classes of amino acid side chains. These classifications are:
Figure imgf000071_0001
Using this classification, there are 64, that is 1296, different peptides (active group) with similar activities against a defined biological target. Accordingly, on average there are around 100 different active groups in each library. Thus, diluting 1 mg of peptide (average molecular weight is 1872) in 100 μl of buffer solution will result in a concentration of approximately 4.1 nM for each active group.
Description of the Assays
Example 1: Mortality assay on 3T3 cells
NIH/3T3 cells were cultured in MEM supplemented with 10%o foetal bovine serum, ImM L-glutamine, 10 U/ml Penicillin and 10 μg/ml Streptomycin, at 37 °C in a 5%> CO2 atmosphere.
5μL of selected vials containing peptide library dissolved in Phosphate Buffer Solution
(PBS) at a total peptide concentration of 4.1 μm was added to the wells of 96 glass bottom well plates containing 150μL of NIH/3T3 (Mouse embryo fibroblasts) cell medium. The cell mortality was recorded after 6 hours and 36 hours. By visual inspection 5 groups were made and assigned values from 1 to 5. "1" means that the cells were as alive as the control without peptide. "5" means that the cells were fully dead, while "2" - "4" are intermediate of these. The results are tabulated in figure 1. The left column of the two entries for each library shows the cell mortality after 6 hours, while the right column shows the cell mortality after 36 hours.
Example 2: Mortality assay on onco genie OCM-1 cells
OCM-l(l,2) (Malignant coroid human) cell medium was cultured in RPMI supplemented with 5% foetal bovine serum, 10 U/ml Penicillin and 10 μg/ml Streptomycin, at 37 °C in a 5%> CO2 atmosphere.
5μL of selected vials containing peptide library dissolved in Phosphate Buffer Solution (PBS) at a total peptide concentration of 4.1 μm was added to the wells of 96 glass bottom well plates containing 150μL of OCM-l(l,2) (Malignant coroid human) cell medium and the cell mortality recorded after 6 hours and 36 hours. By visual inspection 5 groups were made, ranging from 1 to 5. "1" means that the cells were as alive as the control without peptide. "5" means that the cells were fully dead, while "2" - "4" are intermediate. The results are shown in figure 2. The left column of the two entries for each library shows the cell mortality after 6 hours, while the right column shows the cell mortality after 36 hours.
Figures 1 and 2 summarise the effect that the different libraries had on eukaryotic cell viability. In general, all libraries having peptides with a positively charged residue, Lys or Arg, at any of the two defined positions induce cell death, while having negative residues, Asp and Glu, do not compromise cell viability. In the case of NIH/3T3 cells, aromatic residues, Tyr, Phe and Tφ tend to have on average higher mortality rate than hydrophobic or neutral polar residues. It is interesting to note that a much higher mortality rate is observed in the cancer cell line (OCM-1), than in NIH/3T3 cells.
Example 3 : Mortality assay on E. coli cells
E. coli cells (W3113 strain) were cultured in L-Broth medium.
5μL of selected vials dissolved in Phosphate Buffer Solution (PBS) at a total peptide concentration of 4.1 μm was added to the wells of 96 glass bottom well plates containing 150μL of E. coli (W3113 strain) cell medium and the cell density measured after 6 hours and 12 hours by recording the OD of the cell culture at 600 nm using a Tecan Spectia (TECAN Deutschland GmbH) and Delta Soft III software Ver 2.24 (Dr Bechtold and Biometallics Inc.) by following the OD of the cell culture at 600 nm. By visual inspection 11 groups were made, ranging from 10 to 0. "10" corresponds to the OD of the control without peptide and "1" to that after incubating the cells with Kanamycin (30 μg/ml), Ampicillin (100 μg/ml), Tetracyclin (12.5 μg/ml) or Streptomycin (25 μg/ml). The results are shown in figure 3. The left column shows the cell mortality after 6 hours, while the right column shows the cell mortality after 12 hours.
As in the case of the eukaryotic cells lines, those peptides containing negatively charged residues do not compromise bacterial viability, with those with positively charged residues tend to arrest growth However, when comparing E. coli growth inhibition with the effect on NIH/3T3 cells, it was noted that those peptides containing one or two aliphatic residues (Val, He, Leu and Met) produce dramatic inhibition of bacterial growth, while having none or moderate effects on NIH/3T3 cells. Example 4: Comparison of the mortality effect of selected subsets of the library
Those vials that had a significant effect on 3T3, OCM-1 and bacteria were selected. On the same day microtiter plates containing these three cell types were incubated with each of the selected vials at 14 nM peptide concentration for 24 hours. After the incubation period, the effect on cell mortality was checked by visual inspection for eukaryotic cells and by absorbance at 600 nm for prokaryotic cells.
Figure 4 shows the effect of selected vials on cell mortality which had an inhibitory effect on bacterial growth, while having a small effect on 3T3 cells. The libraries Asp-Asp and Arg- Leu are also shown as contiols. For each selected vial, two rows are shown. The first row shows the results of the first experiment which was completed on different days for each cell type. The second row shows the results of the second experiment which was completed simultaneously in the three cell lines with the selected libraries. Regarding the columns, the first column for each vial corresponds to 3T3 cells, the second to OCM-1 and the third to bacteria (E. coli).
The numbers are assigned in the same way as in examples 1 - 3. For eukaryotic cells the numbering goes from 1 (no effect) to 5 (all dead). In the case of bacteria the numbers indicate cell culture OD (0 for no growth and 10 (denoted as #) for maximum growth).
Antibiotic Properties
The results shown in figures 1 - 4 and referred to in Examples 1 - 4, suggest that some of the vials have antibiotic properties, since they arrest bacterial growth while apparently not killing NIH/3T3 cells.
In order to verify this hypothesis, those vials that consistently found they had a moderate, or no, effect on NIH/3T3 cells, while arresting E. coli growth were tested. The selected vials tested were Gly-Phe, His-Leu, Ile-Thr, Ile-Val, Leu-Ala, Leu-Phe, Leu-Gly, Leu-Thr, Val-Gly, Val-Tyr, Tyr-Phe, Tyr-Gly, Asp- Asp and Arg-Leu. The Asp-Asp and Arg-Leu vials were used as contiols. Vial Arg-Leu killed both bacterial and NIH/3T3 cells, while vial Asp-Asp did not affect these cells. Example 5: Titration effect of the positive and negative control vials on E. coli growth
150 μl of bacterial culture/well was placed in a 96 well microtiter plate (OD600 nm = 0.1) and mixed with varying volumes of positive control library Arg-Leu or varying volumes of negative contiol library Asp-Asp. The concentiations of peptide in the final mixtures is shown below in tables 6 and 7.
Figure imgf000075_0001
Table 6
Figure imgf000075_0002
Table 7
The density of the cell culture was monitored every hour for a total period of 12 hours using a Tecan Spectia (TECAN Deutschland GmbH) and Delta Soft III software Ver 2.24 (Dr Bechtold and Biometallics Inc.) by following the OD of the cell culture at 600 nm.
The results of the analysis are shown in figures 5 A and 5B. The Ref+ lines show the growth curve of the contiol without peptide and the Ref- lines show the control without bacteria. Figures 5 A and 5B show that the positive control library Arg-Leu prevents bacteria growth at all concentiations tested, while the negative control library Asp-Asp does not affect growth at all concentiations tested.
Example 6: Titration effect of the library Leu-Phe on E. coli growth
150 μl of bacterial culture/well was placed in a 96 well microtiter plate (OD600 nm = 0.1) and mixed with varying volumes of library Leu-Phe. The concentrations of peptide in the final mixtures is shown below in table 8.
Figure imgf000076_0001
Table 8
The OD of the cell culture was monitored every hour for a total period of 12 hours.
The results of the analysis are shown in figure 6. The Ref+ line shows the growth curve of the contiol without peptide and the Ref- line shows the contiol without bacteria.
Figure 6 demonstrates the concentration effect on the inhibition effect of the Leu-Phe library. It can be seen that the library is still active at very low active concentiations, for example 2.5 nM, and completely abolishes bacterial growth at 7 nM.
When comparing with conventional antibiotics in table 9 below, it can be seen that the active compounds of the selected libraries of example 4 are around 1000 times more active than Ampicillin or Tetiacyclin. Thus, even if it is assumed that all peptides in the selected libraries are equally active, the libraries will have the same activity as conventional antibiotics.
Figure imgf000077_0001
Table 9. All the selected peptides completely block cell growth at 14 nM, the difference lies in the lowest concentration that still inhibits growth to a cell extent. Thus, a range of concentiations, going from the more to the less active compounds, is given.
Example 7: Assessment of antibiotic resistance.
As seen in Figure 6, at low concentrations of peptide the bacteria grow, although it takes longer for them to reach the maximum OD than in the absence of peptide. This observation could be due to a slow degradation of the peptides, or to the acquisition of resistance by the cell culture. In order to distinguish between these possibilities, a dilution experiment was carried out.
150 μl bacteria culture/well was placed in a 96 well microtiter plate (OD600 nm = 0.1) and mixed with a fixed amount of the selected peptide mixtures to a final concentration of 3.4 nM of the active peptide. At this concentration the bacteria can grow although slower than in the absence of peptide. The OD of the cell culture was monitored every hour for a total period of 12 hours. After 12 hours, the culture was centrifuged, the pellet washed with PBS and then diluted in fresh medium containing the same peptide concentration to an OD600 nm = 0.1, and incubated for another 12 hours.
The results of the analysis are shown in figures 7 A - 7N. The Ref+ line shows the growth curve of the control without peptide and the Ref- line shows the control without bacteria, hi each figure, the left graph shows the initial bacterial growth, while the right one shows the bacterial growth after washing and dilution. If the cells have acquired resistance, a normal growth curve should be observed on the right graph. However, if the problem is peptide degradation, we should see the same growth pattern in both the left and right graphs.
Essentially, in all cases the behaviour of the cell culture with a peptide dose allowing partial cell growth is a repetition of the first incubation. Accordingly, bacterial growth after long incubation periods can be attributed to peptide degradation.
Example 8: Mechanism
It is clear from the above examples that the active peptides in the libraries prevent bacterial growth. In order to determine whether the mechanism of prevention of bacterial growth occurs through cell death or the arrest of cell division, the follow analysis was carried out.
A cell culture of 3T3 cells were infected with 10 μl of an overnight culture of E. coli diluted 1000 times. At intervals of 3, 6, 9 and 12 hours, an aliquot of the cell medium was taken, diluted 1000 times and plated on Agar plated. The plates were incubated for 12 hours at 37 °C and the number of colonies counted. Cell mortality was also examined through visual inspection of the cells. In all cases, except with the Arg-Leu library that killed both bacteria and cells, and in the other contiol, library Asp- Asp, that did not prevent bacterial growth, the cells looked healthy and viable and could be cultured longer. The results of this analysis are shown in table 10 below:
Figure imgf000078_0001
Table 10: Relative bactericidal activity of the active selected libraries. For some of the libraries, for example, Ile-Val, Leu-Gly, Tyr-Gly, it was found that the bacteria start growing after a certain period of time, suggesting cell division arrest. In other libraries, some growing, followed by a decline in the number of viable cells, was observed, suggesting cell death. It therefore appears that different peptides prevent bacterial death by different mechanisms.
Example 9: 3T3 cell survival in the presence of the active antibiotic peptides.
NIH/3T3 cells were cultured in MEM supplemented with 10%ι foetal bovine serum, 1 mM L-glutamine, 10 U/ml Penicillin and 10 μg/ml Streptomycin, at 37 °C in a 5 % CO2 atmosphere. The day before the experiment, the cells were spread on a 96 glass bottom well plate at 30%) confluency and 150 μl of media per well. The media was removed and fresh media was added (150 μl/well). Then, 15 μl of a selected vial was added to reach a final active peptide concentration of 14 nM. After 24 hours incubation, the supernatant was removed and the cells were washed once with Dulbecco's phosphated-buffered saline (D-PBS) at 37 °C. The Live/Dead assay was made using the LIVE/DEAD Viability/Cytotoxicity Kit (L3224) from Molecular Probes, following the protocol supplied by the manufacturer. The cells were incubated in a 2 μM Calcein AM and 4 μM Ethidium homodimer-1 in D-PBS for 20 min at room temperature. Then the cells were observed using a DMIRBE Leica photomicroscope and photographed with a COHU high performance CCD camera. As a negative control (dead culture) one of the wells was treated with 3.7%> formaldehyde for 20 min at room temperature, prior to the staining with the Live/Dead assay.
The results of the assay are shown in figure 8. As can be seen the Arg-Leu peptide kills the majority of the cells as expected, while the Asp- Asp peptide has very little effect on cell viability. The selected antibiotic libraries show very little toxicity except for Gly-Phe.
Example 10: Effect of libraries on other cell lines
To check for possible toxicity effects of the libraries with antibiotic activity, different cell lines were incubated for 12 hours with a peptide concentiation that prevented bacteria growth (14 nM). For all the cell lines tested (3T3, HeLa, OCM1, HEK293 and HS68), only in the case of OCM-1 as described above, was significant cell death found for some libraries. Tests on Other Bacterial Strains
The antibiotic properties of the selected libraries were tested on the pathogenic strains Sigella flexneri SC301, Listeria monocytogenes 10403s and E. coli.
Example 11 : Mortality assay on Listeria cells
5μL of selected peptide library dissolved in Phosphate Buffer Solution (PBS) at a total peptide concentration of 3.33 μm was added to the wells of 96 glass bottom well plates containing 150μL of Listeria monocytogenes 10403 s cell medium and the cell mortality recorded after 18 hours. By visual inspection 6 groups were made, ranging from 10 to 0. "10" corresponds to the OD of the control with peptide without peptide and "1" to that after incubating the cells with Kanamycin (30 μg/ml), ampicillin (100 μg/ml), Tetracyclin (12.5 μg/ml) or Streptomycin (25 μg/ml). Figure 9 shows the results.
Example 12: Comparison of mortality assays on Listeria and E. coli
Figure 10 shows a comparison of the mortality effects produced using a total of 5μl of each selected peptide library at a concentration of 14 nM on E. coli (right columns) after 12 hours and on Listeria (left columns) after 18 hours. The cell density was measured by recording the absorbance at 600 nm after 18 hours. By visual inspection 6 groups were made, ranging from 10 to 0. "10" corresponds to the OD of the control without peptide and "1" to that after incubating the cells with Kanamycin (30 μg/ml), Ampicillin (100 μg/ml), Tetiacyclin (12.5 μg/ml) or Streptomycin (25 μg/ml).
Example 13: Titration effect of selected libraries on prokaryote growth
150 μl bacteria culture per well in 96 well microtiter plate (OD(6oonm)= 0.1) were mixed with different volumes of the selected libraries, to result in the final mixture concentiations (each vial has 130321 different peptides) shown in figures 11 A - 1 IN. The OD of the cell culture was monitored every hour for a total period of 12 hours. In magenta the growth curve of the control without peptide and in blue the control without bacteria are shown. In yellow the growth in the presence of the libraries are shown. The figure shows the growth curves for Listeria monocytogenes, Sigella flexneri and E. coli on the left, centie and right, respectively. Example 14: Relative inhibition of cell growth of the active libraries on three prokaryotics.
A total of 5 μl of dissolved peptide library at a concentration of 14 nM was added to each well and the cell density measured by recording the absorbance at 600 nm. Listeria and Sigella were grown in Bacto Hirn-Hernz Glukose-Bouillon media from DIFCO Laboratories under the usual conditions (37 °C, shaking, aerobic culture). Sigella cultures are supplemented with 100 micrograms/ml ampicillin. E. coli was grown as described in example 2. After 12 hours the cell density was measured and was compared to the cells grown in the absence of the peptide libraries (contiol).
The results of the experiment are summarised in table 11, below. When the OD was similar between the control and the cells grown with peptide libraries "+++" was assigned. When the cells did not grow, "-" was assigned. Intermediate values were assigned "++" and "+". The assignment of "+++", "++" or "+" does not mean that there is no growth inhibition at the beginning of the incubation period.
Table 11 Example 15: Role of the tail in the antibacterial activity
150 μl bacteria culture/well in 96 well microtiter plate (OD(6oonm) = 0.1) were mixed with different volumes of the Leu-Phe vial, to result in the final mixture concentiations (each vial has 130321 different peptides) shown in the figure. The OD of the cell culture was monitored after 12 hours. Figure 12 compares the growth curve of the Leu-Phe vial with the tail and a Leu-Phe vial without the tail having compounds with the formula:
Figure imgf000082_0001
wherein R01 is the side chain of the amino acid Leucine and R02 is the side chain of the amino acid Phenylalanine.
It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims. All publications, patents and patent applications cited herein are hereby incoφorated by reference in their entirety for all puiposes to the same extent as if each individual publication, patent or patent application were specifically and individually indicated to be so incoφorated by reference. Concurrently filed copending provisional patent application 60/387,288 ("Combinatorial Chemical Library I") filed June 7, 2002 (Attorney Docket No. 9000/2051) is incoφorated by reference in its entirety).

Claims

1. A chemical library comprising two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula I:
Figure imgf000083_0001
formula I
wherein:
n is an integer from 0 to 10;
m is an integer from 0 to 10;
r is an integer from 0 to 10;
1 9 1 1 1
D are the same or different and each D is independently CR R , CR T or CT ;
1 1 9
T and T are the same or different and each T and each T is independently one of the groups having the structures represented by the formulae:
Figure imgf000083_0002
Figure imgf000084_0001
wherein
p is an integer from 0 to 20;
q is an integer from 0 to 20;
J are the same or different and each J is independently CR lτ R>2. ; and
K is CR1;
X are the same or different and each X is independently O, S, NR or CR R
A are the same or different and each A is independently O, S or NR ;
Y are the same or different and each Y is independently R3
E is R3;
F is R'
G1 and G2 are the same or different and are independently CR*R2, wherein the combination of G and G is selected from the combinations shown in table 1 below:
Figure imgf000085_0001
R1 and R2 are the same or different and each R1 and each R2 is independently hydrogen, alkyl, alkoyl, alkoxy, carboxy, hydroxy, amino, amido, cyano, nitro, thio, sulphonyl, sulphoxide, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally
1 0 occurring amino acid protected by one or more protecting groups, or R and R attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R are the same or different and each R is independently a single bond, alkylene, O S NR4
-O-, -S-, -N(R 4 4)-, X XX , X XX , X XX , -(CX)DA-, -AD(CX)-, -C(O)O-, -OC(O)-, -S(O)-, -S(O)2- , -C(O)N(R4)-, -N(R4)C(O)-, -OC(O)N(R4)-, -N(R4)C(O)O-, -C(S)N(R4)-, -N(R4)C(S)-, -S(O)2N(R4)-, -N(R4)S(O)2-, -N(R4)CON(R4)-, -N(R4)CSN(R4)-, -O-alkylene-, -S-alkylene-, -N(R4)-alkylene-,-C(O)-alkylene-, -C(S)-alkylene-, -C(NR4)- alkylene-,-(CX)DA-alkylene-, -AD(CX)-alkylene-, -C(O)O-alkylene-, -OC(O)-alkylene-, -S(O)-alkylene-, -S(O)2-alkylene-, -C(O)N(R4)-alkylene-, -N(R4)C(O)-alkylene-, -OC(O)N(R4)-alkylene-, -N(R4)C(O)O-alkylene-, -C(S)N(R4)-alkylene-, -N(R4)C(S)-alkylene-, -S(O)2N(R4)-alkylene-, -N(R4)S(O)2-alkylene-, -N(R4)CON(R4)-alkylene-, -N(R4)CSN(R4)-alkylene-, -alkylene-O-, -alkylene-S-, -alkylene-N(R4)-, -alkylene-C(O)-, -alkylene-C(S)-, -alkylene-C(NR4)-, -alkylene-(CX)DA-, -alkylene-AD(CX)-, -alkylene-C(O)O-, -alkylene-OC(O)-, -alkylene-S(O)-, -alkylene-S(O)2-, -alkylene-C(O)N(R4)-, -alkylene-N(R4)C(O>, -alkylene-OC(O)N(R4)-, -alkylene-N(R4)C(O)O-, -alkylene-C(S)N(R4)-, -alkylene-N(R4)C(S)-, -alkylene-S(O)2N(R4)-, -alkylene-N(R4)S(O)2-, -alkylene-N(R4)CON(R4)-, -alkylene-N(R4)CSN(R4)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group; and
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
2. The library of claim 1 wherein the library contains at least 100 compounds.
3. The library of claim 0 wherein the library contains at least 1 ,000 compounds.
4. The library of claim 3 wherein the library contains at least 10,000 compounds.
5. The library of claim 4 wherein the library contains at least 100,000 compounds.
6. The library of any of preceding claim wherein the library contains all the possible different compounds represented by formula I or zwitterions or salts thereof.
7. The library of any preceding claim wherein n is 1 , 2 or 3.
8. The library of any preceding claim wherein m is 1 , 2 or 3.
9. The library of any preceding claim wherein r is 1 , 2 or 3.
10. The library of any preceding claim wherein two groups D are independently CR1!"1 or CT1T2.
11. The library of any preceding claim wherein the groups D, except for any group D which is CRLT! or CTLT2, are independently CR!R2 wherein each R1 and each R2 is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
12. The library of claim 11 wherein each R2 is H and each R1 is independently a side chain of a naturally occurring amino acid for each group D which is CR^2.
1 9 13. The library of any preceding claim wherein the groups D which are CR R ,
1 9 except for any group D which is CH2 and any group D in which R = R , are independently C*R-R2 wherein c* is a stereogenic centie and wherein C* is substantially homochiral in the D- configuration.
14. The library of any preceding claim wherein at least one of groups T1 and T2 is a group having the structure represented by the formula:
Figure imgf000087_0001
Figure imgf000087_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1 ;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CRXR2;
L2 are the same or different and each L2 is independently CR R2.
15. The library of any preceding claims wherein at least one group D is CHT where T1 is the group having the structure represented by the formula:
Figure imgf000088_0001
and at least another group D is CHT1 where T1 is -NHCOCH2NH2.
16. The library of any preceding claim wherein each F is -CO-.
17. The library of any preceding claim wherein each E is independently NR4.
18. The library of claim 17 wherein R4 is H.
19. The library of any one of claims 14 to 18 wherein the groups L1 are independently
1 9 1 0
CR R wherein each R and each R is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups and a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
20. The library of claim 19 wherein each R2 is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid.
21. The library of any one of claims 14 to 20 wherein groups L1, except for any group L1 which is CH2 or any group L1 in which R\ - R2, are independently C*RJR2 wherein C* is a stereogenic centre and wherein C* is substantially homochiral in the D-configuration.
22. The library of any one of claims 14 to 21 wherein the groups L2 are independently CR R2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups and a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
23. The library of claim 22 wherein each R2 is H and each R1 is independently a side chain of a naturally occurring aromatic amino acid.
24. The library of any one of claims 14 to 23 wherein groups L2, except for any groups L2 which is CH2 or any group L2 in which Ri = R2, are independently C*R1R2 wherein C* is a stereogenic centre and wherein C* is substantially homochiral in the D-configuration.
25. The library of any preceding claim wherein the groups G1 and G2 are independently CR!R2 wherein each R1 and each R2 is independently selected from a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid and a side chain of a naturally occurring amino acid protected by one or more protecting groups.
9 1
26. The library of claim 25 wherein each R is H and each R is independently a side chain of a naturally occurring amino acid.
27. The library of any preceding claim wherein groups G1 and G2, except for any group G1 or G2 which is CH2 or any group G1 or G2 in which Ri = R2, are independently C*R!R2 wherein C* is a stereogenic centie and wherein C* is substantially homochiral in the D- configuration.
28. The library of any preceding claim wherein each group A is NR4.
29. The library of claim 28 wherein R 4 i •s H.
30. A chemical library comprising two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula II:
Figure imgf000090_0001
formula II
wherein:
n is selected from the integers 0 to 10;
m is selected from the integers 0 to 10;
r is selected from the integers 0 to 10;
Q1 are the same or different and each Q1 is independently CR*R2;
Q2 are the same or different and each Q2 is independently CR1;
S1 is R2 or T1;
S2 is R2 or T2;
T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000091_0001
Figure imgf000091_0002
wherein
u is an integer from 0 to 20; v is an integer from 0 to 20;
w is 0 or 1;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CR'R2;
9 9 1 L are the same or different and each L is independently CR R ;
Y are the same or different and each Y is independently -(CO)Q1(NR4)-,
-(NR4)Q1(CO)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R1 and R2 are the same or different and each R1 and each R2 is independently hydrogen, alkyl, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R1 and R2 attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R3 are the same or different and each R3 is independently a single bond, alkylene, O S NR4
-O-, -S-, -N(R , )-, 11 , I , I ^ , a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group; R , R , R and R are the same or different and are independently H or a side chain of a naturally occurring amino acid, wherein the combination of R5, R6, R7 and R8 is selected from the combinations shown in table 4 below:
Figure imgf000093_0001
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
31. The library of claim 30 wherein any stereogenic carbon atoms between a carbonyl moiety and a nitrogen atom are in the D-configuration.
32. The library of claim 30 or claim 31 wherein R5 is -H, R6 is -H, R7 is -CH2(C6H5) and R8 is -H.
33. The library of claim 30 or claim 31 wherein R5 is
Figure imgf000094_0001
, R6 is -H,
R7 is -CH2CH(CH3)2 and R8 is -H.
34. The library of claim 30 or claim 31 wherein R5 is -CH(CH3)CH2CH3, R6 is -H, R7 is -CH2(OH)CH3 and R8 is -H.
35. The library of claim 30 or claim 31 wherein R5 is -CH(CH3)CH2CH3, R6 is -H, R7 is -CH(CH3)2 and R8 is -H.
36. The library of claim 30 or claim 31 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is - CH3 and R8 is -H.
37. The library of claim 30 or claim 31 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -CH2(C6H5) and R8 is -H.
38. The library of claim 30 or claim 31 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is - H and R8 is -H.
39. The library of claim 30 or claim 31 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -CH(OH)CH3 and R8 is -H.
40. The library of claim 30 or claim 31 wherein R5 is -CH(CH3)2, R6 is -H, R7 is -CH3 and R8 is -H.
41. The library of claim 30 or claim 31 wherein R5 is -CH(CH3)2, R6 is -H, R7 is - CH2(C6H4)OH and R8 is -H.
42. The library of claim 30 or claim 31 wherein R5 is -CH2(C6H4)OH, R6 is -H, R7 is - CH2(C6H5) and R8 is -H.
43. The library of claim 30 or claim 31 wherein R5 is -CH2(C6H4)OH, R6 is -H, R7 is - H and R8 is -H.
44. The library of claim 30 or claim 31 wherein R5 is -CH2COOH, R6 is -H, R7 is - CH2COOH and R8 is -H.
45. The library of claim 30 or claim 31 wherein R5 is -(CH2)3NH(CNH)NH2, R6 is - H, R7 is -CH2CH(CH3)2 and R8 is -H.
1 ^ 46. The library of any of claims 30 to 45 wherein one of T or T is the group having the structure represented by the formula:
Figure imgf000095_0001
and the other of T1 or T2 is -NHCOCH2NH2.
47. A chemical library comprising two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
1
Figure imgf000096_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above.
48. A chemical library comprising two or more different compounds, or zwitterions or salts thereof, each compound independently having a structure represented by the formula:
Figure imgf000097_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above.
49.' A compound, or pharmaceutically acceptable derivative thereof, having a structure represented by the formula I:
Figure imgf000098_0001
formula I
wherein:
n is an integer from 0 to 10;
m is an integer from 0 to 10;
r is an integer from 0 to 10;
D are the same or different and each D is independently CR l Rr>2 , n CRτ> l T or CT T
T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000098_0002
Figure imgf000098_0003
Figure imgf000099_0001
wherein
p is an integer from 0 to 20;
q is an integer from 0 to 20;
J are the same or different and each J is independently CR lτ R>2. ; and
K is CR1;
X are the same or different and each X is independently O, S, NR4 or CR4R4;
A are the same or different and each A is independently O, S or NR4;
Y are the same or different and each Y is independently R3;
E is R'
F is R';
G1 and G2 are the same or different and are independently CR^2, wherein the combination of G1 and G2 is selected from the combinations shown in table 1 below:
Figure imgf000099_0002
Figure imgf000100_0001
R1 and R2 are the same or different and each R1 and each R2 is independently hydrogen, alkyl, alkoyl, alkoxy, carboxy, hydroxy, amino, amido, cyano, nitro, thio, sulphonyl, sulphoxide, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally
1 9 occurring amino acid protected by one or more protecting groups, or R and R attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R3 are the same or different and each R3 is independently a single bond, alkylene,
O NR4
-O-, -S-, -N(R4)-: X X X -(CX)DA-, -AD(CX)-, -C(O)O-, -OC(O)-, -S(O)-, -S(O)2- , -C(O)N(R4)-, -N(R4)C(O)-, -OC(O)N(R4)-, -N(R4)C(O)O-, -C(S)N(R4)-, -N(R4)C(S)-, -S(O)2N(R4)-, -N(R4)S(O)2-, -N(R4)CON(R4)-, -N(R4)CSN(R4)-, -O-alkylene-, -S-alkylene-, -N(R4)-alkylene-,-C(O)-alkylene-, -C(S)-alkylene-, -C(NR4)- alkylene-,-(CX)DA-alkylene-, -AD(CX)-alkylene-, -C(O)O-alkylene-, -OC(O)-alkylene-, -S(0)-alkylene-, -S(O)2-alkylene-, -C(O)N(R4)-alkylene-, -N(R4)C(O)-alkylene-, -OC(O)N(R4)-alkylene-, -N(R4)C(O)O-alkylene-, -C(S)N(R4)-alkylene-, -N(R4)C(S)-alkylene-, -S(O)2N(R4)-alkylene-, -N(R4)S(O)2-alkylene-, -N(R4)CON(R4)-alkylene-, -N(R4)CSN(R4)-alkylene-, -alkylene-O-, -alkylene-S-, -alkylene-N(R4)-, -alkylene-C(O)-, -alkylene-C(S)-, -alkylene-C(NR4)-, -alkylene-(CX)DA-, -alkylene-AD(CX)-, -alkylene-C(O)O-, -alkylene-OC(O)-, -alkylene-S(O)-, -alkylene-S(O)2-, -alkylene-C(O)N(R4)-, -alkylene-N(R4)C(O)-, -alkylene-OC(O)N(R4)-, -alkylene-N(R4)C(O)O-, -alkylene-C(S)N(R4)-, -alkylene-N(R4)C(S)-, -alkylene-S(O)2N(R4)-, -alkylene-N(R4)S(O)2-, -alkylene-N(R4)CON(R4)-, -alkylene-N(R4)CSN(R4)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group; and
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
50. The compound of claim 49 wherein n is 1, 2 or 3.
51. The compound of claim 49 or claim 50 wherein m is 1 , 2 or 3.
52. The compound of any one of claims 49 to 51 wherein r is 1 , 2 or 3.
53. The compound of any one of claims 49 to 52 wherein two groups D are independently CR.lΥl or CT'T2.
54. The compound of any one of claims 49 to 53 wherein the groups D, except for
1 1 1 9 1 9 1 9 any group D which is CR T or CT T , are independently CR R wherein each R and each R is independently H, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO , a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups.
55. The compound of claim 54 wherein each R2 is H and each R1 is independently a side chain of a naturally occurring amino acid for each group D which is CR*R2.
56. The compound of any one of claims 49 to claim wherein the groups D which are CR^2, except for any group D which is CH2 and any group D in which R1 = R2, are
1 9 independently C*R R wherein C* is a stereogenic centie and wherein C* is substantially homochiral in the D-configuration.
57. The compound of any one of claims 49 to 56 wherein at least one of groups T1 and T2 is a group having the structure represented by the formula:
Figure imgf000102_0001
Figure imgf000102_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1;
provided that the sum u + v + w is not greater than 20;
1 1 1 9
L are the same or different and each L is independently CR R ;
L2 are the same or different and each L2 is independently CRXR2.
58. The compound of claim 57 wherein at least one group D is CHT1 where T1 is the group having the structure represented by the formula:
Figure imgf000103_0001
and at least another group D is CHT1 where T1 is -NHCOCH2NH2.
59. The compound of any one of claims 49 to 58 wherein each F is -CO-.
60. The compound of any one of claims 49 to 59 wherein each E is independently NR4.
61. The compound of claim 60 wherein R 4 i ■s H
62. The compound of any one of claims 49 to 61 wherein the groups L1 are independently CRJR2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups and a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
63. The compound of claim 62 wherein each R2 is H and each R1 is independently a side chain of a naturally occurring positively charged amino acid.
64. The compound of any one of claims 49 to wherein groups L1, except for any group L1 which is CH2 or any group L1 in which Ri = R2, are independently C*R1R2 wherein C* is a stereogenic centie and wherein C* is substantially homochiral in the D-configuration.
65. The compound of any one of claims 49 to 64 wherei •n the groups L 9 are independently CR^2 wherein each R1 and each R2 is independently H, a side chain of a naturally occurring positively charged amino acid, a side chain of a naturally occurring aromatic amino acid, a substituted side chain of a naturally occurring positively charged amino acid, a substituted side chain of a naturally occurring aromatic amino acid, a side chain of a naturally occurring positively charged amino acid protected by one or more protecting groups or a side chain of a naturally occurring aromatic amino acid protected by one or more protecting groups.
9 1
66. The compound of claim 65 wherein each R is H and each R is independently a side chain of a naturally occurring aromatic amino acid.
67. The compound of any one of claims 49 to 66 wherein groups L2, except for any groups L2 which is CH2 or any group L2 in which Ri = R , are independently C*RXR2 wherein C* is a stereogenic centre and wherein C* is substantially homochiral in the D-configuration.
68. The compound of any one of claims 49 to 67 wherein the groups G1 and G2 are independently CR!R2 wherein each R1 and each R2 is independently selected from a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid and a side chain of a naturally occurring amino acid protected by one or more protecting groups.
9 1
69. The compound of claim 68 wherein each R is H and each R is independently a side chain of a naturally occurring amino acid.
70. The compound of any one of claims 49 to 69 wherein groups G1 and G2, except for any group G1 or G2 which is CH2 or any group G1 or G2 in which Ri = R2, are independently
C*RXR2 wherein C* is a stereogenic centie and wherein C* is substantially homochiral in the D- configuration.
71. The compound of any one of claims 49 to 70 wherein each group A is independently NR4.
72. The compound of claim 71 wherein R4 is H.
73. A compound, or pharmaceutically acceptable derivative thereof, having the structure represented by the formula II:
Figure imgf000105_0001
formula II
wherein:
n is selected from the integers 0 to 10;
m is selected from the integers 0 to 10;
r is selected from the integers 0 to 10;
1 1 1 9
Q are the same or different and each Q is independently CR R ;
Q2 are the same or different and each Q2 is independently CR1;
S1 is R2 or T1;
S2 is R2 or T2; T1 and T2 are the same or different and each T1 and each T2 is independently one of the groups having the structures represented by the formulae:
Figure imgf000106_0001
Figure imgf000106_0002
wherein
u is an integer from 0 to 20;
v is an integer from 0 to 20;
w is 0 or 1;
provided that the sum u + v + w is not greater than 20;
L1 are the same or different and each L1 is independently CR!R2;
L2 are the same or different and each L2 is independently CR!R2;
Y are the same or different and each Y is independently -(CO)Q1(NR4)-, -(NR >4Λ)Q~> 11(/CO)-, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups; 1 9 1 9
R and R are the same or different and each R and each R is independently hydrogen, alkyl, R4 2NCR10 2(CO)NR4-, R4O(CO)CR10 2NR4(CO)-, a side chain of a naturally occurring amino acid, a substituted side chain of a naturally occurring amino acid or a side chain of a naturally occurring amino acid protected by one or more protecting groups, or R1 and R2 attached to the same carbon atom together complete an alicyclic or heterocyclic system;
R3 are the same or different and each R3 is independently a single bond, alkylene,
-O-, -S-, -N(R4)-:
Figure imgf000107_0001
, a divalent radical of a side chain of a naturally occurring amino acid, a divalent radical of a substituted side chain of a naturally occurring amino acid or a divalent radical of a side chain of a naturally occurring amino acid protected by one or more protecting groups ;
R4 are the same or different and each R4 is independently hydrogen, alkyl or a protecting group;
R5, R6, R7 and R8 are the same or different and are independently H or a side chain of a naturally occurring amino acid, wherein the combination of R5, R6, R7 and R8 is selected from the combinations shown in table 4 below:
Figure imgf000107_0002
Figure imgf000108_0003
Table 4
R10 are the same or different and each R10 is independently hydrogen or a side chain of a naturally occurring amino acid.
74. The compound of claim 73 wherein any stereogenic carbon atoms between a carbonyl moiety and a nitrogen atom are in the D-configuration.
75. The compound of claim 73 or claim 74 wherein R is -H,
Figure imgf000108_0001
is - CH2(C6H5) and R8 is -H.
76. The compound of claim 73 or claim 74 wherein R5 is
Figure imgf000108_0002
, R6 is
H, R >7' . is -CH2CH(CH3)2 and Rδ is -H.
77. The compound of claim 73 or claim 74 wherein R5 is -CH(CH3)CH2CH3, R6 is -
H, R7 is -CH2(OH)CH3 and R8 is -H.
78. The compound of claim 73 or claim 74 wherein R5 is -CH(CH3)CH2CH3, R6 is - H, R7 is -CH(CH3)2 and R8 is -H.
79. The compound of claim 73 or claim 74 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -CH3 and R8 is -H.
80. The compound of claim 73 or claim 74 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -CH2(C6H5) and R8 is -H.
81. The compound of claim 73 or claim 74 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -H andR8 is -H.
82. The compound of claim 73 or claim 74 wherein R5 is -CH2CH(CH3)2, R6 is -H, R7 is -CH(OH)CH3 and R8 is -H.
83. The compound of claim 73 or claim 74 wherein R5 is -CH(CH3)2, R6 is -H, R7 is - CH3andR8is-H.
84. The compound of claim 73 or claim 74 wherein R5 is -CH(CH3)2, R6 is -H, R7 is - CH2(C6H4)OH and R8 is -H.
85. The compound of claim 73 or claim 74 wherein R5 is -CH2(C6H4)OH, R6 is -H, R7 is-CH2(C6H5)andR8is-H.
86. The compound of claim 73 or claim 74 wherein R5 is -CH2(C6H4)OH, R6 is -H, R7 is-HandR8is-H.
87. The compound of claim 73 or claim 74 wherein R5 is -CH2COOH, R6 is -H, R7 is -CH2COOHandR8is-H.
88. The compound of claim 73 or claim 74 wherein R5 is -(CH2)3NH(CNH)NH2, R6 is -H, R7 is -CH2CH(CH3)2 and R8 is -H.
89. The compound of any one of claims 73 to 88 wherein one of T1 or T2 is the group having the structure represented by the formula:
Figure imgf000110_0001
and the other of T1 or T2 is -NHCOCH2NH2.
90. A compound, or pharmaceutically acceptable derivative thereof, having the structure represented by the formula:
Figure imgf000110_0002
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid,
wherein the combination of R5 and R7 is selected from the combinations shown in table 4 above.
91. A compound, or pharmaceutically acceptable derivative thereof, having the structure represented by the formula:
Figure imgf000111_0001
wherein:
R9 are the same of different and each R9 is independently a side chain of a naturally occurring amino acid;
R5 is a side chain of a naturally occurring amino acid; and
R7 is a side chain of a naturally occurring amino acid.
wherein the combination of R and R is selected from the combinations shown in table 4 above.
92. A kit comprising two or more libraries of claims 1 to 48.
93. A compound of any one of claims 49 to 91 for use in the treatment of a bacterial infection.
94. A method of treating a bacterial infection comprising administering to a patient in need of such treatment an effective dose of a compound of any one of claims 49 to 91.
95. A pharmaceutical composition comprising a compound of any one of claims 49 to 91 in combination with a pharmaceutically acceptable excipient.
Ill
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