WO1997000883A1

WO1997000883A1 - Meiosis regulating compounds

Info

Publication number: WO1997000883A1
Application number: PCT/DK1996/000271
Authority: WO
Inventors: Frederik Christian GRØNVALD; Peter Faarup; Erling Guddal
Original assignee: Novo Nordisk A/S
Priority date: 1995-06-23
Filing date: 1996-06-20
Publication date: 1997-01-09
Also published as: AU6122696A; ZA965332B

Abstract

A compound of general formula (I) wherein R?1, R2 and R3¿, independently, are hydrogen or methyl; and R?4 and R5¿ are either hydrogen or together designate an additional bond between the carbon atoms to which they are bound can be used for regulating the meiosis.

Description

MEIOSIS REGULATING COMPOUNDS

FIELD OF THE INVENTION

The present invention relates to pharmacologically active compounds and to their use as medicaments. More particularly it has been found that the compounds of the invention can be used for regulating the meiosis.

BACKGROUND OF THE INVENTION

Meiosis is the unique and ultimate event of germ cells on which sexual reproduction is based. Meiosis comprises two meiotic divisions. During the first division, exchange between maternal and paternal genes take place before the pairs of chromosomes are separated into the two daughter cells. These contain only half the number (1n) of chromosomes and 2c DNA. The second meiotic division proceeds without a DNA synthesis. This division therefore results in the formation of the haploid germ cells with only 1 c DNA.

The meiotic events are similar in the male and female germ cells, but the time schedule and the differentiation processes which lead to ova and to spermatozoa differ profoundly. All female germ cells enter the prophase of the first meiotic division early in life, often before birth, but all are arrested as oocytes later in the prophase (dictyate state) until ovulation after puberty. Thus, from early life the female has a stock of oocytes which is drawn upon until the stock is exhausted. Meiosis in females is not completed until after fertilisation, and results in only one ovum and two abortive polar bodies per germ cell. In contrast, only some of the male germ cells enter meiosis from puberty and leave a stem population of germ cells throughout life. Once initiated, meiosis in the male cell proceeds without significant delay and produces four spermatozoa.

Only little is known about the mechanisms which control the initiation of meiosis in the male and in the female. In the oocyte, new studies indicate that follicular purines, hypoxanthine or adenosine, could be responsible for meiotic arrest (Downs, S.M. et al. Dev. Biol. 82 (1985) 454-458; Eppig, J.J. et al. Dev. Biol. 119 (1986) 313-321; and Downs, S.M. Mol. Reprod. Dev. 35 (1993) 82-94). The presence of a diffusible meiosis regulating substance was first described by Byskov et al. in a culture system of fetal mouse gonads (Byskov, A.G. et al. Dev. Biol. 52 (1976) 193-200). A meiosis activating substance (MAS) was secreted by the fetal mouse ovary in which meiosis was ongoing, and a meiosis preventing substance (MPS) was released from the moφhologically differentiated testis with resting, non-meiotic germ cells. It was suggested that the relative concentrations of MAS and MPS regulated the beginning, arrest and resumption of meiosis in the male and in the female germ cells (Byskov, AG. et al. in The Physiology of Reproduction (eds. Knobil, E. and Neill, J.D., Raven Press, New York (1994)). Clearly, if meiosis can be regulated, reproduction can be controlled. A recent article (Byskov, AG. et al. Nature 374 (1995) 559-562) describes the isolation from bull testes and from human follicular fluid of certain sterols that activate oocyte meiosis. Unfortunately, these sterols are rather labile and utilisation of the interesting finding would thus be greatly facilitated if more stable meiosis activating compounds were available.

SUMMARY OF THE INVENTION

According to the present invention there are provided novel, stable compounds with interesting pharmacological properties. In particular, the compounds of the invention are useful for regulating the meiosis in oocytes and in male germ cells.

In its broadest aspect, the present invention relates to compounds of the general formula (I)

(I) wherein R¹, R² and R³, independently, are hydrogen or methyl; R⁴ and R⁵ are either hydrogen or together they designate an additional bond between the carbon atoms to which they are bound.

In a preferred aspect, the present invention relates to a compound of the general formula (I) wherein R¹ is hydrogen.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R¹ is methyl.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R² is hydrogen.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R² is methyl.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R³ is hydrogen.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R³ is methyl.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R⁴and R5 are both hydrogen.

In another preferred aspect, the present invention relates to a compound of the general formula (I) wherein R⁴ and R5 together designate an additional double bond between the carbon atoms to which they are bound.

In a further aspect, the present invention relates to the use of a compound of the general formula (I) in the manufacture of a medicament.

In a further aspect, the present invention relates to the use of a compound of the general formula (I) in the manufacture of a medicament for use in the regulation of meiosis.

In the present context, the expression "regulating the meiosis" is understood to indicate that the compounds can be used for stimulating the meiosis in vitro, in vivo, and ex vivo.

In a still further aspect, the present invention relates to the use of a compound of the general formula (I) in the regulation of the meiosis of oocytes or male germ cells.

In a yet still further aspect, the present invention relates to a method of regulating the meiosis in a mammalian germ cell which method comprises administering an effective amount of a compound of the general formula (I) above to a germ cell in need of such a treatment.

DETAILED DESCRIPTION OF THE INVENTION

The compounds of the present invention will induce resumption of meiosis in oocytes as well as in male germ cells. The existence of a meiosis inducing substance in nature has been known for some time. However, until recently the identity of the meiosis inducing substance or substances has been unknown.

The prospects of being able to influence the meiosis are several. According to a preferred embodiment of the present invention, the compounds of the general formula (I) are used to stimulate the meiosis. According to another preferred em¬ bodiment of the present invention, the compounds of the general formula (I) are used to stimulate the meiosis in humans. Thus, the compounds of formula (I) are promising as new fertility regulating agents without the usual side effect on the somatic cells which are known from the hitherto used hormonal contraceptives which are based on estrogens and/or gestagens. For use as a contraceptive agent in females, a meiosis inducing substance can be administered so as to prematurely induce resumption of meiosis in oocytes while they are still in the growing follicle, before the ovulatory peak of gonadotropins occurs. In women, the resumption of the meiosis can, for example, be induced a week after the preceding menstruation has ceased. When ovulated, the resulting overmature oocytes are then most likely not to be fertilised. The normal menstrual cycle is not likely to be affected. In this connection it is important to notice, that the biosynthesis of progesterone in cultured human granulosa cells (somatic cells of the follicle) is not affected by the presence of a meiosis inducing substance whereas the estrogens and gestagens used in the hitherto used hormonal contraceptives do have an adverse effect on the biosynthesis of progesterone.

According to another aspect of this invention, a meiosis inducing substance of the general formula (I) can be used in the treatment of certain cases of infertility in females, including women, by administration thereof to females who, due to an insufficient own production of MIS, are unable to produce mature oocytes. Also, when in vitro fertilisation is performed, better results can be achieved, when a meiosis inducing substance of the general formula (I) is added to the medium in which the oocytes are kept. Also, when infertility in males, including men, is caused by an insufficient own production of the meiosis inducing substance administration of a meiosis inducing substance of the general formula (I) can be used for relieving the problem.

The route of administration of the compositions containing a compound of formula (I) may be any route which effectively transports the active compound to its site of action.

Thus, when the compounds of this invention are to be administered to a mammal, they are conveniently provided in the form of a pharmaceutical composition which comprises at least one compound of formula (I) in connection with a phar¬ maceutically acceptable carrier. For oral use, such compositions are preferably in the form of capsules or tablets.

From the above it will be understood that administrative regimen called for will depend on the condition to be treated. Thus, when used in the treatment of infertility the administration may be once only, or for a limited period, e.g. until pregnancy is achieved. When used as a contraceptive, the meiosis inducing substance of the general formula (I) will either have to be taken continuously or cyclically. When used as a contraceptive by women and not taken continuously, the timing of the administration relative to the menstrual cycle will be important.

The pharmaceutical compositions may comprise carriers, diluents, absoφtion enhancers, preservatives, buffers, agents for adjusting the osmotic pressure, tablet disintegrating agents and other ingredients which are conventionally used in the art. Examples of solid carriers are magnesium carbonate, magnesium stearate, dextrin, lactose, sugar, talc, gelatine, pectin, tragacanth, methyl cellulose, sodium carboxymethyl cellulose, low melting waxes and cocoa butter.

Liquid compositions include sterile solutions, suspensions and emulsions. Such liquid compositions may be suitable for injection or for use in connection with ex vivo and in vitro fertilisation. The liquid compositions may contain other ingredients which are conventionally used in the art, some of which are mentioned in the list above.

Further, a composition for transdermal administration of a compound of this invention may be provided in the form of a patch and a composition for nasal administration may be provided in the form of a nasal spray in liquid or powder form.

The dose of a compound of the invention to be used will be determined by a physician and will depend, inter alia, on the particular compound employed, on the route of administration and on the purpose of the use.

The compounds of the general formula (I) can be synthesised by methods known per se. Examples of preferred compounds of the general formula (I) are the following:

18-nor-5α, 17α(H)-cholesta-8, 11 ,13-triene-3β-ol;

12-methyl-18-nor-5α, 17α(H)-cholesta-8, 11 ,13-triene-3β-ol;

4,4-dimethyl-12-methyl-18-nor-5α, 17α(H)-cholesta-8, 11 ,13-triene-3β-ol;

4,4-dimethyl-18-nor-5α, 17α(H)-cholesta-8, 11 ,13-triene-3β-ol;

18-norcholesta-8, 11 ,13,24-tetraene-3β-ol; 12-methyl-18-norcholesta-8, 11 ,13,24-tetraene-3β-ol;

4,4-dimethyl-12-methyl-18-norcholesta-8, 11 ,13,24-tetraene-3β-ol; and

4,4-dimethyl-18-norcholesta-8, 11 ,13,24-tetraene-3β-ol.

The present invention is further illustrated by examples which, however, are not to be construed as limiting the scope of protection. The features disclosed in the foregoing description and in the examples may, either separately or in any combination thereof, be material for realising the invention in diverse forms thereof.

EXAMPLES

EXAMPLE 1 Preparation of 12-methyl-18-nor-5α(H)cholesta-8,11 ,13-triene-3β-ol.

12-methyl-18-nor-5α(H)-cholesta-8,11,13-triene-3β-ol was prepared by a series of steps according to a Anastasia M et al. Lipids 17 (1982) 220. The product showed the characteristics as described.

¹H-NMR: Hδ: 2.34 (3H, s, CH₃ at C₁₂); 6.90 (1H, d.-H).

Claims

1. A compound ofthe general formula (I)

(I) wherein R^ , R² and R³, independently, are hydrogen or methyl; and R⁴ and R^ are either hydrogen or together designate an additional bond between the carbon atoms to which they are bound.

2. A compound according to claim 1 wherein R^ and R² are both hydrogen.

3. A compound according to claim 1 wherein one of R-' and R² is hydrogen while the other is methyl.

4. A compound according to claim 1 wherein R^ and R² are both methyl.

5. A compound according to any one of the claims 1 to 4 wherein R³ is hydrogen.

6. A compound according to any one of the claims 1 to 4 wherein R³ is methyl.

7. A compound according to any one of the claims 1 to 6 wherein R⁴ and R^ are both hydrogen.

8. A compound according to any one of the claims 1 to 6 wherein R⁴ and R⁵ together designate a bond between the carbon atoms to which they are bound.

9. A compound of general formula (I) as described in any of the claims 1 to 8 for use as a medicament.

10. A compound of general formula (I) as described in any of the claims 1 to 8 for use in the regulation of meiosis.

11. A method of regulating the meiosis in a mammalian germ cell which method comprises administering an effective amount of a compound according to any one of the claims 1 to 8 to a germ cell in need of such a treatment.

12. A method according to claim 11 wherein a compound according to any one of the claims 1 to 8 is administered to a germ cell by administering it to a mammal hosting said cell.

13. A method according to claim 11 or 12 wherein the germ cell the meiosis of which is to be regulated is an oocyte.

14. A method according to claim 11 wherein a compound according to any one of the claims 1 to 8 is administered to an oocyte ex vivo.

15. A method according to claim 12 wherein the germ cell the meiosis of which is to be regulated is a male germ cell.

16. Use of a compound of general formula (I) as described in any of the claims 1 to 8 in the manufacture of a medicament.