ABSTRACT
Experimental data indicate that low-calcemic vitamin D derivatives (VDDs) exhibit anticancer properties, both in vitro and in vivo. In our search for a vitamin D analog as potential anticancer agent, we investigated the influence of chirality in the side chain of the derivatives of 1,25-dihydroxyergocalciferol (1,25D2) on their activities. In this study, we synthesized modified analogs at the side chain and the A-ring, which differed from one another in their absolute configuration at C-24, namely (24S)- and (24R)-1,25-dihydroxy-19-nor-20a-homo-ergocalciferols (PRI-5105 and PRI-5106, respectively), and evaluated their activity. Unexpectedly, despite introducing double-point modifications, both analogs served as very good substrates for the vitamin D-hydroxylating enzyme. Irrespective of their absolute C-24 configuration, PRI-5105 and PRI-5106 showed relatively low resistance to CYP24A1-dependent metabolic deactivation. Additionally, both VDDs revealed a similar antiproliferative activity against HT-29 colorectal cancer cells which was higher than that of 1,25D3, the major biologically active metabolite of vitamin D. Furthermore, PRI-5105 and PRI-5106 significantly enhanced the cell growth-inhibitory activity of 5-fluorouracil on HT-29 cell line. In conclusion, although the two derivatives showed a relatively high anticancer potential, they exhibited undesired high metabolic conversion.
Subject(s)
Antineoplastic Agents/chemical synthesis , Colorectal Neoplasms/metabolism , Ergocalciferols/chemical synthesis , Vitamin D3 24-Hydroxylase/metabolism , Vitamin D/analogs & derivatives , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Proliferation/drug effects , Cell Survival/drug effects , Colorectal Neoplasms/drug therapy , Drug Synergism , Epithelial-Mesenchymal Transition/drug effects , Ergocalciferols/chemistry , Ergocalciferols/pharmacology , Fluorouracil/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , HCT116 Cells , HT29 Cells , Humans , Molecular Structure , Signal Transduction/drug effects , Vitamin D/chemistryABSTRACT
Two unknown impurities were observed during the process development for multigram-scale synthesis of eplerenone (Inspra®). The new process-related impurities were identified and fully characterized as the corresponding (7ß,11α,17α)-11-hydroxy- and (7α,11ß,17α)-9,11-dichloroeplerenone derivatives 12a and 13. Seven other known but poorly described in the literature eplerenone impurities, including four impurities A, B, C and E listed in the European Pharmacopoeia 8.4 were also detected, identified and fully characterized. All these contaminants result from side reactions taking place on the steroid ring C of the starting 11α-hydroxy-7α-(methoxycarbonyl)-3-oxo-17α-pregn-4-ene-21,17-carbolactone (12) and the key intermediate (7α,17α)-9(11)-enester 7, including epimerization of the C-7 asymmetric center, oxidation, dehydration, chlorination and lactonization. The impurities were isolated and/or synthesized and fully characterized by infrared spectroscopy (IR), nuclear magnetic resonance spectroscopy (NMR) and high-resolution mass spectrometry/electrospray ionization (HRMS/ESI). Their ¹H- and 13C-NMR signals were fully assigned. The molecular structures of the eight impurities, including the new (7ß,11α,17α)-11-hydroxy- and (7α,11ß,17α)-9,11-dichloroeplerenone related substances 12a and 13, were solved and refined using single-crystal X-ray diffraction (SCXRD). The full identification and characterization of these impurities should be useful for the quality control and the validation of the analytical methods in the manufacture of eplerenone.
Subject(s)
Antihypertensive Agents/chemistry , Drug Contamination , Spironolactone/analogs & derivatives , Chromatography, High Pressure Liquid , Eplerenone , Magnetic Resonance Spectroscopy , Molecular Structure , Spectrometry, Mass, Electrospray Ionization , Spectrophotometry, Infrared , Spironolactone/chemistryABSTRACT
1,25-dihydroxyvitamin D3 (1,25D), a steroid hormone which regulates calcium/phosphate homeostasis, has a broad spectrum of anti-cancer activities, including differentiation of acute myeloid leukemia (AML) cells. In order to avoid undesirable side effects such as hypercalcemia, low-calcemic analogues should be produced for therapeutic purposes. In this paper, we describe biological activities of double-point modified analogues of vitamin D2 and we compare them to 1,25D and to paricalcitol, the drug used to treat secondary hyperparathyroidism. In vivo, our new analogues have lower calcemic effects, and lower toxicity in comparison to 1,25D. They have enhanced pro-differentiating and transcription-inducing activities in AML cells. Interestingly, differentiation effects do not correlate with the affinities of the analogues to the vitamin D receptor (VDR).
Subject(s)
Ergocalciferols/chemistry , Ergocalciferols/chemical synthesis , Active Transport, Cell Nucleus , Animals , CCAAT-Enhancer-Binding Protein-beta/metabolism , Cell Differentiation , Drug Evaluation, Preclinical , Ergocalciferols/genetics , HL-60 Cells , Homeostasis , Humans , Hypercalcemia/metabolism , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Lipopolysaccharide Receptors/metabolism , Mice , Point Mutation , Receptors, Calcitriol/metabolism , Transcriptional Activation/drug effects , Up-Regulation , Vitamin D3 24-Hydroxylase/metabolismABSTRACT
Analogs of 1,25-dihydroxyergocalciferol, modified in the side-chain and in the A-ring, were tested for their antiproliferative activity against a series of human cancer cell lines in vitro and in vivo toxicity. The proliferation inhibition caused by the analogs was higher than that of the parent compounds, while the toxicity, measured as the serum calcium level, was lower. All analogs were able to induce, in HL-60 and MV4-11 leukemic cells, G0/G1 cell cycle arrest and differentiation expressed as morphological signs typical for monocytes. The analogs also induced the expression of CD11b and/or CD14 cell-differentiation markers. The most potent analogs, PRI-5105, PRI-5106, PRI-5201 and PRI-5202, were also able to induce vitamin D receptor (VDR) protein expression, mainly in the cytoplasmic fraction of HL-60 or MV4-11 cells. The most active analogs were the 19-nor ones with an extended and rigidified side-chain (PRI-5201 and PRI-5202), as in the former analogs PRI-1906 and PRI-1907. Epimerization at C-24 (PRI-5101) or introduction of an additional hydroxyl at C-23 (PRI-5104) reduced the toxicity of the analog with retained antiproliferative activity.
Subject(s)
Ergocalciferols/pharmacology , Cell Cycle/drug effects , Cell Differentiation/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Ergocalciferols/chemistry , Gene Expression Regulation, Neoplastic/drug effects , HL-60 Cells , Humans , Receptors, Calcitriol/metabolismABSTRACT
As predicted by single crystal X-ray crystallography, and contrary to the reported suggestions, the anhydrous form of calcipotriol, a therapeutically important vitamin D analog, was found stable enough to be used as an active pharmaceutical ingredient. The crystal and molecular structure of calcipotriol anhydrate was solved and refined using single crystal X-ray diffraction. The analog was obtained by a novel convergent synthesis from the vitamin D C-22 sulfone, as an advanced intermediate and a side-chain fragment. The homo-chiral side-chain aldehyde was obtained from cyclopropanecarboxyaldehyde by the chromatographic separation of the intermediate diastereomeric salts with (S)-naproxen. Calcipotriol anhydrate showed a single peak in differential scanning calorimetry and the absence of a peak from a water molecule, typical for the monohydrate. Calcipotriol anhydrate, as the only 1,25-dihydroxylated analog of vitamin D3 , exists as a mixture of both α- and ß-forms of the A-ring, present in the asymmetric part of the unit cell of the crystal lattice. The intermolecular hydrogen bonding between both conformers in the crystal lattice indicated that the stability of calcipotriol anhydrate might be at least the same as for the known monohydrate. The usefulness of calcipotriol anhydrate as an active pharmaceutical ingredient was confirmed by the stability study in the standard conditions used for the storage of vitamin D analogs.
Subject(s)
Calcitriol/analogs & derivatives , Vitamin D/analogs & derivatives , Vitamins/chemistry , Water/chemistry , Calcitriol/chemical synthesis , Calcitriol/chemistry , Calorimetry, Differential Scanning , Crystallography, X-Ray , Drug Stability , Models, Molecular , Vitamin D/chemical synthesis , Vitamin D/chemistry , Vitamins/chemical synthesisABSTRACT
The hybrid analogs of 1,25-dihydroxyergocalciferol (PRI-5201 and PRI-5202) were synthesized as potential anticancer agents using a convergent strategy. The analogs were designed by combining a 19-nor modification of the A-ring with the homologated and rigidified ergocalciferol-like side-chain of the previously obtained analogs PRI-1906 and PRI-1907. The strategy also allowed the novel efficient synthesis of 19-nor-1,25-dihydroxyergocalciferol (paricalcitol, PRI-5100) and its (24R)-diastereomer (PRI-5101). The single crystal X-ray structures of the 19-nor analogs (PRI-5100 and PRI-5101) were solved and refined. The A-ring of both analogs adopts exclusively chair ß-conformation in the solid state. The side-chain of these analogs is coplanar with the CD-ring plane, while it is perpendicular in 1,25-dihydroxycholecalciferol.
Subject(s)
Antineoplastic Agents/chemical synthesis , Ergocalciferols/chemical synthesis , Antineoplastic Agents/chemistry , Crystallography, X-Ray , Ergocalciferols/chemistry , Hydrogen Bonding , Models, Molecular , Molecular Conformation , Oxidation-ReductionABSTRACT
The 17-phenyl PGF(2α) analogue bimatoprost (10a) is the most efficacious ocular hypotensive agent currently available for the treatment of glaucoma or ocular hypertension. A novel convergent synthesis of 13,14-en-15-ol prostamideF(2α) analogues was developed employing Julia-Lythgoe olefination of the structurally advanced phenylsulfone (+)-(5Z)-15 with an enantiomerically pure aldehyde ω-chain synthon (-)-(S)-16a. Subsequent hydrolysis of protecting groups and final amidation of the diol 26a yielded bimatoprost (10a). The main advantage of the current strategy is the preparation of high-purity bimatoprost (10a). The novel convergent strategy allows the synthesis of a whole series of 13,14-en-15-ol prostamideF(2α) analogues with the desired C-15 asymmetric center configuration from a common and structurally advanced prostaglandin intermediate (+)-(5Z)-15. The preparation and identification of two synthetic impurities, 15-epi isomer (10b) of bimatoprost and a new prostaglandin related amide (+)-(5Z)-18, are also described.