Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07J—STEROIDS
  • C07J9/00—Normal steroids containing carbon, hydrogen, halogen or oxygen substituted in position 17 beta by a chain of more than two carbon atoms, e.g. cholane, cholestane, coprostane
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P3/00—Drugs for disorders of the metabolism
  • A61P3/02—Nutrients, e.g. vitamins, minerals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P3/00—Drugs for disorders of the metabolism
  • A61P3/12—Drugs for disorders of the metabolism for electrolyte homeostasis
  • A61P3/14—Drugs for disorders of the metabolism for electrolyte homeostasis for calcium homeostasis
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
  • C07C401/00—Irradiation products of cholesterol or its derivatives; Vitamin D derivatives, 9,10-seco cyclopenta[a]phenanthrene or analogues obtained by chemical preparation without irradiation
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07J—STEROIDS
  • C07J71/00—Steroids in which the cyclopenta(a)hydrophenanthrene skeleton is condensed with a heterocyclic ring
  • C07J71/0036—Nitrogen-containing hetero ring
  • C07J71/0042—Nitrogen only

Definitions

• This invention relates to a biologically active vitamin D compound.
• the invention relates to a novel 1 ⁇ ,25-dihydroxylated vitamin D compound with a 22,23-cis-double bond in the side chain, and to a method for its preparation.
• Patents 3,741,996; 3,907,843; 3,880,894; 4,226,788; 4,358,406) Most of the known active analogs are characterized by the type of sterol side chain as it occurs in vitamin D 3 (i.e. saturated side chain).
• Known analogs with 22,23-unsaturated side chain are represented by compounds of the vitamin D 2 series (i.e. 22,23-trans-unsaturated with a C-24-methyl substituent), and include, in addition to the compounds named above, 25-hydroxyvitamin D 2 (U.S. Patent 3,585,221) and the 24- and 24,25-dihydroxy derivatives (Jones et al. Acch. Biochem. Biophys.
• This novel compound is characterized by a 22,23-double bond in the side chain having the cis (or Z) geometry. Because of the presence of this 22Z-double bond, which results in a quite different side chain geometry from that pertaining to compounds having the normal saturated side chain (e.g. as in 1 ⁇ ,25-dihydroxyvitamin D 3 ) or a 22,23-trans (22E)-unsaturated side chain (e.g. as in 1 ⁇ ,25-dihydraxyvitamin D 2 ) , it was assumed that this cis-unsaturated product would exhibit lew biological activity, if any. Surprisingly, this material, in spite of its altered side chain structure, shows high activity, being as active as 1 ⁇ ,25-dihydroxyvitamin D 3 in its ability to raise serum calcium levels in test animals.
• novel product of this invention was prepared from a 22Z-dehydrovitamin D precursor having the structure II shown below, by in vitro enzymatic hydroxylation at carbon 25 using a liver homogenate preparation from vitamin D-deficient rats.
• Incubation was carried out in 10 ml incubation medium in a 125 ml Erle ⁇ mayer flask containing an aliquot of liver homogenate representing 1 g of tissue, 0.125 M sucrose, 50 mM phosphate buffer (pH 7.4) , 22.4 mM glucose-6-phosphate, 20 m ATP, 160 mM nicotinamide, 25 mM succinate, 0.4 mM NADP, 5 mM MgCl 2 , 0.1 M KCl and 0.5 units glucose-6-phosphatedehydrogenase.
• the reaction was initiated by addition of 400 ⁇ g of the substrate, compound II above, dissolved in 100 ⁇ l 95% ethanol.
• the incubation mixture was incubated at 37°C with shaking at 80 oscillations/min for 3 h.
• the reaction was stopped by addition of 20 ml methanol and 10 ml dichloromethane.
• the organic phase was collected while the aqueous phase was re-extracted with 10 ml dichloromethane.
• the organic phases from total of three extractions were combined and evaporated with a rotary evaporator.
• the residue containing the desired product was dissolved in 1 ml of CHCl 3 :hexane (65:35) mixture and applied to a Sephadex LH-20 column (0.7 cm x 14 cm) packed, equilibrated and eluted with the same solvent.
• the mass spectrum of the substance contains a molecular ion at m/e 414 as required for a 25-hydroxylated product. Elimination of one and two molecules of H 2 O gives fragment ions at m/e 396 and 378. Loss of the entire steroid side chain (cleavage of C 17 /C 20 bond) results in the fragment of m/e 287, which by elimination of one and two molecules of H 2 O, gives rise to the peaks at m/e 269 and 251. The spectrum shews prominent peaks at m/e 152 and 134 (152-H 2 O) which represent ring A fragments and are diagnostic for 1 ⁇ ,3 ⁇ -dihydroxyvitamin D compounds.
• Serum obtained by centrifugation of the blood was diluted with
• the compound of this invention will find application as a therapeutic agent in the therapy or prophylaxis of disorders such as the various types of rickets, hypoparathyroidism, osteodystrophy, osteomalacia or osteoporosis in the human, or for the treatment of related calcium deficiency diseases (e.g. milk fever, leg weakness, egg shell thinness) in animals.
• disorders such as the various types of rickets, hypoparathyroidism, osteodystrophy, osteomalacia or osteoporosis in the human
• related calcium deficiency diseases e.g. milk fever, leg weakness, egg shell thinness
• the compound may be used for the treatment of certain malignancies, such as human leukemia.
• the compound may be administered by any conventional route of administration and in any form suitable for the method of administration selected.
• the compound may be formulated with any acceptable and innocuous pharmaceutical carrier, in the form of pills, tablets, gelatin capsules, or suppositories, or as solutions, emulsions, dispersions or suspensions in innocuous solvents or oils, and such formulations may contain also other therapeutically active and beneficial ingredients as may be appropriate for the specific applications.
• the compound is advantageously administered in amounts from 0.25 to 10 ⁇ g per day, the specific dosage being adjusted in accordance with the disease to be treated and the medical history, condition and response of the subject, as is well understood by those skilled in the art.
• the 22Z-dehydro precursor substrate, compound II above, required for the preparation of the novel product of this invention is itself prepared by the process depicted in Process Scheme I, appended, and described below.
• compound designation by Arabic numerals e.g. (1) r (2) , (3) , etc.
• the desired substrate (compound II) for the above described 25-hydroxylation reaction is identified by Arabic numeral (11) in Process Scheme I and in the following description.
• Vitamin analog (5) is a known compound (Bogoslovskii et al. , supra) . 1-Hydroxylation of compound (5) . Freshly recrystallized p-toluenesulfonyl chloride (50 mg, 0.26 mmol) was added to a solution of vitamin (5) (50 mg, 0.13 mmol) in dry pyridine (300 ⁇ l) . After 30 h at 4°C, the reaction mixture was poured into ice/saturated NaHCO 3 with stirring.
• the mixture was stirred for 15 min and extracted with benzene.
• the organic extract was washed with saturated NaHCO 3 , saturated copper sulfate and water, dried (Na 2 SO 4 ) and concentrated in vacuo to obtain the oily tosylate (6) .
• the crude tosylate (6) was treated with NaECO 3 (150 mg) in anhydrous methanol (10 ml) and the mixture was stirred for 8.5 h at 55°C. After cooling and concentration to 2 ml the mixture was diluted with benzene (80 ml) , washed with water, dried (Na 2 SO 4 ) and evaporated under reduced pressure.
• the oily 3,5-cyclovitamin D compound (7) thus obtained was sufficiently pure to be used for the following oxidation step without any purification.
• tert-butylhydroperoxide (16.5 ⁇ l, 0.118 mmol) was added.
• dry pyridine 50 ⁇ l was added and the mixture was stirred for additional 25 min at room temperature, diluted with CH 2 Cl 2 (3 ml) and cooled to 0°C.
• the crude 3,5-cyclovitamin product (7) in CH 2 Cl 2 (4.5 ml) was then added.
• the reaction proceeded at 0°C for 15 min and then it was allowed to warm slowly (30 min) to room tarperature.
• the mixture was transferred to a separatory funnel and shaken with 30 ml of 10% NaOH.
• Ether 150 ml was added and the separate organic phase was washed with 10% NaOH, water and dried over Na 2 SO 4 .
• HPLC high pressure liquid chromatography
• a Waters Associates Model ALC/GPC 204 using a Zorbax-Sil (DuPont) (6.2mm x 25 cm column, flow rate 4 ml/min, 1500 psi).
• Column chromatography was performed on Silica Gel 60, 70-230 mesh ASTM (Merck) .
• Preparative thin-layer chromatography (TLC) was carried out on Silica 60 PF-254 (20 x 20 cm plates, 1 mm silica gel). Irradiations were carried out using a Hanovia 608A36 mercury arc lamp fitted with a Vycor filter. All reactions are preferably performed under an inert atmosphere (e.g. argon).
• the compound of this invention can, if desired, be readily obtainedin crystalline form by crystallization from suitable solvents such as hexane, ethers and alcohols (absolute or aqueous) , and mixtures thereof as will be evident and well known to those skilled in the art.
• suitable solvents such as hexane, ethers and alcohols (absolute or aqueous) , and mixtures thereof as will be evident and well known to those skilled in the art.

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• Health & Medical Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Organic Chemistry (AREA)
• General Health & Medical Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Medicinal Chemistry (AREA)
• General Chemical & Material Sciences (AREA)
• Pharmacology & Pharmacy (AREA)
• Life Sciences & Earth Sciences (AREA)
• Animal Behavior & Ethology (AREA)
• Chemical Kinetics & Catalysis (AREA)
• Public Health (AREA)
• Veterinary Medicine (AREA)
• Diabetes (AREA)
• Engineering & Computer Science (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Hematology (AREA)
• Obesity (AREA)
• Rheumatology (AREA)
• Endocrinology (AREA)
• Nutrition Science (AREA)
• Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
• Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)

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Citations (3)

• 1985
  • 1985-01-07 WO PCT/US1985/000017 patent/WO1985003300A1/en active Application Filing
  • 1985-01-07 CH CH4260/85A patent/CH665835A5/de not_active IP Right Cessation
  • 1985-01-07 DE DE19853590021 patent/DE3590021T/de active Pending
  • 1985-01-07 DE DE19853590021 patent/DE3590021C2/de not_active Expired - Lifetime
  • 1985-01-07 AU AU38372/85A patent/AU587174B2/en not_active Ceased
  • 1985-01-07 JP JP60500432A patent/JPS61501147A/ja active Granted
  • 1985-01-07 NL NL8520009A patent/NL8520009A/nl unknown
  • 1985-01-29 FR FR8501230A patent/FR2558828B1/fr not_active Expired
  • 1985-01-29 IE IE213/85A patent/IE57936B1/en not_active IP Right Cessation
  • 1985-01-29 GB GB08502161A patent/GB2153358B/en not_active Expired
  • 1985-01-29 BE BE0/214412A patent/BE901601A/fr not_active IP Right Cessation
  • 1985-09-27 DK DK437685A patent/DK437685D0/da not_active Application Discontinuation

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