Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
  • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
  • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
  • A61K31/52—Purines, e.g. adenine
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
  • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
  • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
  • A61K31/52—Purines, e.g. adenine
  • A61K31/522—Purines, e.g. adenine having oxo groups directly attached to the heterocyclic ring, e.g. hypoxanthine, guanine, acyclovir
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
  • A61K31/575—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of three or more carbon atoms, e.g. cholane, cholestane, ergosterol, sitosterol
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
  • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P15/00—Drugs for genital or sexual disorders; Contraceptives
  • A61P15/08—Drugs for genital or sexual disorders; Contraceptives for gonadal disorders or for enhancing fertility, e.g. inducers of ovulation or of spermatogenesis
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Definitions

• the present invention relates to a pharmaceutical composition and its use to treat infertility.
• 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 (1 n) of chromosomes and 2c DNA. The second meiotic division proceeds without a DNA synthesis. This division therefore results in the formation of the hapioid germ cells with only 1c 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
• Cyclic adenosine 5 ' -monophosphate plays a pivotal role as a second messenger in the signal transduction pathway during meiosis in the oocyte
• cAMP is generated by the action of adenylate cyclase (AC).
• AC adenylate cyclase
• cAMP is degraded by the family of phosphodiesterase enzymes (PDE), which produces inactive second messenger products
• Hypoxanthine is an inhibitor of cAMP PDE (Eppig, J J et al (1985) Biol Reprod. 33 1041-1049). As such, it can prevent the hydrolysis of oocyte cAMP and thereby maintain elevated levels of cAMP in the oocyte.
• agents acting upstream or downstream of cAMP are able to increase cAMP levels.
• activation of AC with forskolin inhibition of PDE with the nonselective 3- isobutyl-1-methyixanth ⁇ ne (lBMX) or inhibition of the oocyte-specific isoform PDE3 with a specific PDE3- ⁇ nhib ⁇ tor, e.g mii ⁇ none, leads to meiotic arrest by maintaining elevated levels of c-AMP within the oocytes (Downs SM and Hunzicker-Dunn M (1995) Dev Biol 172. 72-85, Tsafin A et al (1996) Dev Biol 178. 393-402)
• a PDE3 specific inhibitor has been described as a contraceptive agent (WO98/10765).
• the present invention provides a pharmaceutical composition comprising c-AMP-increasing compounds in low dose and at least one meiosis-stimulating compound for the treatment of infertility in mammals, particulary in humans, more particulary in females.
• c-AMP-increasing compounds in low dose and at least one meiosis-stimulating compound for the treatment of infertility in mammals, particulary in humans, more particulary in females.
• the increase of fertility by low dose c-AMP- increasing compounds is surprising since the teaching of the prior art is that c-AMP- increasing compounds are responsible for meiotic arrest and a PDE3 specific inhibitor has even been used as a contraceptive agent.
• the combination of the low dose cAMP-increasing compounds with at least one meiosis-stimulating compound shows a significantly higher stimulation rate than the meiosis-stimulating compound(s) alone.
• the invention relates to the use of low dose c-AMP- increasing compounds alone or in combination with at least one meiosis-stimulating compound as active substances for the production of a pharmaceutical composition for the treatment of infertility in mammals, particular in humans, more particularly in females.
• Low dose c-AMP-increasing compounds alone increase the fertilization rate compared to the control where no compounds were added and the combination of c- AMP-increasing compounds in a low dose with a meiosis-stimulating compound shows an even higher rate of fertilization than the meiosis-stimulating compound alone or the low dose c-AMP-increasing compound alone.
• the invention relates to the use of low dose c-AMP- increasing compounds alone or in combination with at least one meiosis-stimulating compound to increase the rate of fertilization in a mammal, particularly in humans, more particularly in females.
• This use may be to regulate the fertilization rate in a fertilization culture media.
• the invention includes the use of a low dose c-AMP-increasing compounds alone or in combination with at least one meiosis-stimulating compound for the administration to a germ cell.
• the germ ceil may be an oocyte or a spermatozoon.
• the invention in another embodiment relates to a method of stimulating meiosis in a mammalian germ ceil comprising administering ex vivo or in vivo or in vitro to a germ cell in need of such a stimulation an effective amount of low dose c-AMP-increasing compounds alone or in combination with at least one meiosis-stimulating compound.
• the germ cell may be an oocyte or a spermatozoon.
• the invention in a further embodiment relates to a pharmaceutical kit comprising a dosage unit for a c-AMP-increasing compound in low dose and a dosage unit of at least one meiosis-stimulating compound.
• the c-AMP-increasing compound and the meiosis-stimulating compound may be provided in the same application form or in different application forms.
• Application forms means e.g. tablets, liquid compositions for injections, paste and others well known in the art.
• Meiosis-stimulating compounds according to the present invention are all compounds that can activate meiosis. Compounds being known to stimulate meiosis and their synthesis are described, i.e., in WO 96/27658, WO97/00884, WO96/00235, W098/28323 and W098/52965. In preferred embodiments of all modes of the invention the meiosis-stimulating compound is FF-MAS (4,4-dimethyl-5 ⁇ -cholesta- 8,14,24-trien-3 ⁇ -ol).
• the stimulation rate is at least 40 - 50 %, preferred 50 - 75%, and more preferred 75 - 100 %.
• Low dose cAMP-increasing compounds according to the present invention is a dose of cAMP-increasing compounds that lead to meiotic maturation without inducing meiotic arrest.
• the cAMP- increasing compounds are applied in a dose below 3mM, more preferred in a dose of 0.003 - 1 mM, and especially preferred in a dose of 0.1-0.5mM.
• the cAMP- increasing compounds are purines, unspecific PDE-inhibitors, specific PDE 3 - inhibitors or synthetic membrane permeable cAMP.
• a purine is e.g. hypoxanthine or adenosine.
• Unspecific PDE-inhibitors are nonselective inhibitors which inhibit all types of PDEs, e.g. IBMX, theophylline or SQ20,006 (1-ethyl-4-hydrazino-14- pyrazolo-(3,4-b)-pyridine-5-carboxylicacidethyiester).
• PDE3 inhibitors examples include milrinone, cilostamide, amrinone, enoximone, lixazinone, indolidan and other inhibitors as listed in WO98/10765. This reference also gives the methods of preparation of these compounds.
• An example for a synthetic membrane permeable cAMP is dibutyryl-c-AMP (dbcAMP).
• the meiosis-stimulating compound is FF-MAS and the cAMP-increasing compound is a purine, preferably hypoxanthine.
• compositions may comprise pharmaceutically acceptable excipients well known in the art like carriers, diluents, absorption enhancers, preservatives, buffers, agents for adjusting the osmotic pressure, tablet disintegrating agents and other ingredients which are conventionally used in the art.
• solid carriers are magnesium carbonate, magnesium stearate, dextrin, lactose, sugar, talc, gelatin, pectin, tragacanth, methylcellulose, sodium carboxymethyl cellulose, low melting waxes and cacao 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, in vivo ,and in vitro fertilization.
• the liquid compositions may contain other ingredients which are conventionally used in the art, some of which are mentioned in the list above.
• a composition for transdermal administration of a compound of this invention may be provided in the form of a patch
• a composition for nasal administration may be provided in the form of a nasal spray in liquid or powder form
• a composition for intra-vaginal administration may be provided in the form of a tampon or other intra-vaginal devices.
• the dosage to be administered depends to a large extent on the condition and the size of the subject being treated as well as the frequency of treatment and the route of administration.
• compositions of the invention are prepared by intimately bringing into association the active compound or compounds with the liquid or solid auxiliary ingredients and then, if necessary, shaping the product into the desired formulation.
• Example 1 Test of hypoxanthine, FF-MAS and the combination of FF-MAS and hypoxanthine in the oocyte assay
• Naked oocytes (NO) and cumulus enclosed oocytes (CEO) were isolated from follicles from immature (C57B1/6J x DBA/2J)F ⁇ mice (age 21-24 days), that had received 10 I.U. PMSG i.p. 48h prior to collection.
• the oocytes were cultured in 4- well multidishes in a modified ⁇ -MEM medium containing 1mg fetuin/ml culture medium. Each well contained 0.4 ml of the oocyte culture medium and 35-45 oocytes.
• the control and test cultures were made with different concentrations of the compounds to be tested as indicated in the tables.
• the cultures were kept at 37°C and 100% humidity with 5% C0 2 in the air for 18 hours.
• Oocytes arrested in meiosis are characterised by an intact nucleus with a prominent nucleolus, known as germinal vesicle (GV).
• GV germinal vesicle
• Table 1 Activation of meiosis in oocytes using FF-MAS, low dose hypoxanthine and the combination of FF-MAS + hypoxanthine
• GVB germinal vesicle breakdown
• n number of
• hypoxanthine used was obtained from Sigma, Deisenhofen, Germany. Similar to the control, low dose hypoxanthine (0.4mM) is able to lead to meiotic maturation in most oocytes. However, 3mM hypoxanthine nearly completely prevents meiotic maturation. It is evident that FF-MAS not only when given together with a meiosis - inhibiting dose of hypoxanthine (3mM), but also when given together with low dose hypoxanthine (0.4mM) is able to lead to meiotic maturation in nearly all oocytes.
• Example 2 Test of hypoxanthine, FF-MAS and the combination of FF-MAS and hypoxanthine in the in-vitro fertilization (IVF) assay
• GVB germinal vesicle breakdown
• hypoxanthine used was obtained from Sigma, Deisenhofen, Germany. Oocytes cultured in the presence of 3mM hypoxanthine were unable to get fertilized. 0.4mM hypoxanthine and 10 ⁇ M FF-MAS + 3 mM hypoxanthine increased the fertilization rate of the control group by about 50%. However, the combination of 10 ⁇ M FF-MAS + 0.4 mM hypoxanthine could nearly double the control IVF-rate.

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• 1999
  • 1999-05-07 AU AU54089/99A patent/AU5408999A/en not_active Abandoned
  • 1999-05-07 CA CA002333320A patent/CA2333320A1/en not_active Abandoned
  • 1999-05-07 WO PCT/EP1999/003138 patent/WO1999061010A2/en not_active Application Discontinuation
  • 1999-05-07 KR KR1020007013278A patent/KR20010043828A/ko not_active Application Discontinuation
  • 1999-05-07 JP JP2000550470A patent/JP2002516272A/ja active Pending
  • 1999-05-07 EP EP99939972A patent/EP1098652A2/de not_active Withdrawn
• 2000
  • 2000-11-24 NO NO20005954A patent/NO20005954L/no not_active Application Discontinuation

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