Papers by Alexander Misharin
The Journal of Steroid Biochemistry and Molecular Biology, Jun 1, 2023
Topics in Current Chemistry
Archiv der Pharmazie
The aim of this study was to explore the mechanisms of action of alsevirone in prostate cancer (P... more The aim of this study was to explore the mechanisms of action of alsevirone in prostate cancer (PC) in vitro and in vivo: CYP17A1 inhibition, cytotoxic, apoptotic, and antitumor effects in comparison with abiraterone. The CYP17A1-inhibitory activity was investigated in rat testicular microsomes using high-performance liquid chromatography. Testosterone levels were evaluated using enzyme-linked immunoassay. IC50 values were calculated for PC3, DU-145, LNCaP, and 22Rv1 cells using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test. The antitumor effect in vivo was studied in DU-145 and 22Rv1 subcutaneous xenografts in Balb/c nude mice. Alsevirone reduced the CYP17A1-inhibitory activity by 98% ± 0.2%. A statistically significant reduction in the testosterone concentration in murine blood was recorded after the 7th administration of 300 mg/kg alsevirone at 0.31 ± 0.03 ng/ml (p < .001) versus 0.98 ± 0.22 ng/ml (p = .392) after abiraterone administration and 1.52 ± 0.49 ng/ml in control animals. Alsevirone was more cytotoxic than abiraterone in DU-145, LNCaP, and 22Rv1 cells, with IC50 values of 23.80 ± 1.18 versus 151.43 ± 23.70 μM, 22.87 ± 0.54 versus 28.80 ± 1.61 μM, and 35.86 ± 5.63 versus 109.87 ± 35.15 μM, respectively. Alsevirone and abiraterone significantly increased annexin V-positive, caspase 3/7-positive, and activated Bcl-2-positive cells. In 22Rv1 xenografts, alsevirone 300 mg/kg × 10/24 h per os inhibited tumor growth: on Day 9 of treatment, tumor growth inhibition = 59% (p = .022). Thus, alsevirone demonstrated significant antitumor activity associated with CYP17A1 inhibition, apoptosis in PC cells, and testosterone reduction.
European Journal of Medicinal Chemistry
The conjugation of biologically active molecules is a powerful tool for drug discovery used to ta... more The conjugation of biologically active molecules is a powerful tool for drug discovery used to target a variety of multifunctional diseases including cancer. Conjugated drugs can provide combination therapies in a single multi-functional agent and, by doing so, be more specific and powerful than conventional classic treatments. Steroids are widely used for conjugation with other biological active molecules. This review refers to investigations of steroid conjugates as potential anticancer agents carried out mostly over the past decade. It consists of five parts in which the data concerning structure and anticancer activity of steroid conjugates with DNA alkylating agents, metallocomplexes, approved drugs, some biological active molecules, some natural compounds and related synthetic analogs are described.
Steroids
Graphical abstract Figure. No Caption available. HighlightsTestosterone and epitestosterone chemi... more Graphical abstract Figure. No Caption available. HighlightsTestosterone and epitestosterone chemically conjugated with pyropheophorbide a.Spectral properties and molecular models of conjugates are presented.Epitestosterone conjugates inhibit LNCaP and PC‐3 cells growth stronger.Conformationally rigid conjugates possess stronger anti‐proliferative activity. ABSTRACT Conjugates of 17&agr;‐substituted testosterone (1 and 2) and 17&bgr;‐substituted epitestosterone (3 and 4) with pyropheophorbide a were synthesized. The scheme consisted of synthesis of 17&agr;‐hydroxy‐3‐oxopregn‐4‐en‐21‐oic and 17&bgr;‐hydroxy‐3‐oxopregn‐4‐en‐21‐oic acids, and their coupling with pyropheophorbide a by means of either ethylene diamine, or 1,5‐diamino pentane linkers. Mutual influence of steroidal and macrocyclic fragments in conjugates molecules was dependent on configuration of C17 and length of linker, that was established by analysis of 1H NMR spectra and molecular models of conjugates. Studies of interaction of conjugates with prostate carcinoma cells revealed that their uptake and internalization were independent on the androgen receptor activity, but dependent on the structure of conjugates, decreasing in the following row: 3 > 4 ≥ 1 > 2. Conjugates significantly decreased the LNCaP and PC‐3 cells growth at 96 h incubation. Epitestosterone derivatives 3 and 4 also showed superior anti‐proliferative activity versus testosterone ones. Conformationally more rigid conjugates 1 and 3, comprising short linkers, were more active than those with long linkers; conjugate 3 was the most potent.
Steroids
Seven new oxazoline, benzoxazole and benzimidazole derivatives were synthesized from 3β-acetoxyan... more Seven new oxazoline, benzoxazole and benzimidazole derivatives were synthesized from 3β-acetoxyandrosta-5,16-dien-17-carboxylic, 3β-acetoxyandrost-5-en-17β-carboxylic and 3β-acetoxypregn-5-en-21-oic acids. Docking to active site of human 17α-hydroxylase/17,20-lyase revealed that all oxazolines, as well as benzoxazoles and benzimidazoles comprising Δ16 could form stable complexes with enzyme, in which steroid moiety is positioned similarly to that of abiraterone and galeterone, and nitrogen atom coordinates heme iron, while 16,17-saturated benzoxazoles and benzimidazoles could only bind in a position where heterocycle is located nearly parallel to heme plane. Modeling of the interaction of new benzoxazole and benzimidazole derivatives with androgen receptor revealed the destabilization of helix 12, constituting activation function 2 (AF2) site, by mentioned compounds, similar to one induced by known antagonist galeterone. The synthesized compounds inhibited growth of prostate carcinoma LNCaP and PC-3 cells at 96 h incubation; the potency of 2'-(3β-hydroxyandrosta-5,16-dien-17-yl)-4',5'-dihydro-1',3'-oxazole and 2'-(3β-hydroxyandrosta-5,16-dien-17-yl)-benzimidazole was superior and could inspire further investigations of these compounds as potential anti-cancer agents.
Steroids
&NA; A number of isoxazole, 1,2,3‐triazole, tetrazole, and 1,2,4‐oxadiazole derivatives of [1... more &NA; A number of isoxazole, 1,2,3‐triazole, tetrazole, and 1,2,4‐oxadiazole derivatives of [17(20)E]‐21‐norpregnene comprising 3&bgr;‐hydroxy‐5‐ene and 3‐oxo‐4‐ene fragments were prepared. Among the key steps for the synthesis of isoxazoles, 1,2,3‐triazoles, and tetrazoles were (i) 1,3‐dipolar cycloaddition of nitrile oxides or azides to acetylenes or nitriles and ii) dehydration of 17&bgr;‐hydroxy‐17&agr;‐methylene‐azoles to [17(20)E]‐21‐norpregnene derivatives. 1,2,4‐Oxadiazoles were prepared through the formation of acetimidamides. Potency of the synthesized compounds to inhibit CYP17A1 and to suppress growth of prostate carcinoma cells was investigated. Among the new azole derivatives, four compounds were found possessing high anti‐proliferative activity. Graphical abstract Figure. No caption available.
Macroheterocycles
Two new conjugates of pyropheophorbide a with testosterone and dihydrotestosterone: 17 3 [2-(17β-... more Two new conjugates of pyropheophorbide a with testosterone and dihydrotestosterone: 17 3 [2-(17β-hydroxy-3-oxopregn-4-en-21-oylamido)ethylamido]pyropheophorbide a (10) and 17 3 [2-(17β-hydroxy-3-oxopregnan-21-oylamido)ethylamido]pyropheophorbide a (11) were synthesized. IC 50 for conjugates 10 and 11 at 96 h incubation in LNCaP and PC-3 prostate carcinoma cells were 1.4 μM and 3.3 μM for compound 10, and 4.5 μM and 6.1 μM for compound 11, respectively. Irradiation with light at wavelength of 660 nm increased toxicity of the conjugates.
Macroheterocycles
Two novel complex conjugates (containing three functional units: pyropheophorbide a, 17α-substitu... more Two novel complex conjugates (containing three functional units: pyropheophorbide a, 17α-substituted testosterone, and lipophylic hexadecyl chain, connected with L-lysine joining block) were synthesized. The scheme consisted of condensation of N(α)-Fmoc-N(ε)-Boc-Lys with hexadecyl amine, followed by consecutive removal of N-protective groups and coupling of obtained intermediates either with pyropheophorbide a, or with 17β-hydroxy-3-oxopregn-4-en-21-oic acid. Mutual influence of steroidal and macrocyclic fragments depending on conjugate structure was established by analysis of NMR spectra and molecular models of conjugates. Complex conjugates easily formed mixed micelles with phosphatidyl choline and pluronic F68; these mixed micelles efficiently internalized by human hepatocarcinoma Hep G2 cells, and slightly-by human prostate carcinoma LNCaP cells. The binding of complex conjugates to cells was dependent on the conjugate structure.
Steroids, 2018
Four new 4,5-dihydro-1,3-oxazole, and four new benzo-[d]-oxazole derivatives of [17(20)E]-21-norp... more Four new 4,5-dihydro-1,3-oxazole, and four new benzo-[d]-oxazole derivatives of [17(20)E]-21-norpregnene, differing in the structure of steroid moiety, were synthesized and evaluated for their potency to inhibit 17α-hydroxylase/17,20-lyase (CYP17A1) activity. Among new compounds, the only oxazolinyl derivative comprising 5-oxo-4,5-seco-3-yn- moiety potently inhibited CYP17A1. Binding modes of the oxazolinyl derivatives of [17(20)E]-21-norpregnene were analyzed by molecular dynamics simulations, and model of alternate, water-bridged type II interaction was proposed for these compounds. Eight new compounds, together with two CYP17A1-inhibiting oxazolinyl derivatives synthesized earlier, abiraterone and galeterone were evaluated for their potency to inhibit prostate carcinoma PC-3 and LNCaP cells growth. Oxazolinyl and benzoxazolyl derivatives comprising 3β-hydroxy-5-ene moieties potently inhibited prostate carcinoma cell growth; inhibitory potencies of 3-oxo-4-en- and 5-oxo-4,5-seco-3...
Steroids, 2016
Five 4,5-dihydro-1,3-oxazole derivatives of [17(20)E]-21-norpregnene, comprising 3β-hydroxy-5-ene... more Five 4,5-dihydro-1,3-oxazole derivatives of [17(20)E]-21-norpregnene, comprising 3β-hydroxy-5-ene (1), 3,6-dioxo-4-ene (2), 3-oxo-4-ene (3), 3α,5α-cyclo-6-oxo (4), 3β-hydroxy-6-oxo (5) fragments were synthesized. Synthesis was conducted with improved procedure, based on reaction of suitably protected [17(20)E]-pregnen-21-oic acids with ethanolamine in presence of triphenyl phosphine, carbon tetrachloride, and triethyl amine. Potency of the compounds 1-5 to inhibit 17α-hydroxylase/17,20-lyase (CYP17A1) activity was studied by highly sensitive electrochemical method, using the enzyme immobilization technique. Compounds 1 and 3 were found to be potent CYP17A1 inhibitors, compounds 2 and 5 were not active, compound 4 strongly and irreversibly suppressed the enzyme activity. Molecular docking of compounds 1-5 in the active site of CYP17A1 showed that positions of all compounds in the enzyme active site were similar.
Biochemistry Supplement Series B Biomedical Chemistry, 2007
Novel synthetic oxysterols (22S,23S)-3 β-hydroxy-22,23-oxido-5 α-ergost-8(14)-en-15-one (I) and (... more Novel synthetic oxysterols (22S,23S)-3 β-hydroxy-22,23-oxido-5 α-ergost-8(14)-en-15-one (I) and (22R,23R)-3 β-hydroxy-22,23-oxido-5 α-ergost-8(14)-en-15-one (II) efficiently inhibited cholesterol biosynthesis in human hepatoma Hep G2 cells during short-term incubation in a serum free medium (IC 50 values of 1.9 ± 0.2 and 0.6 ± 0.2 μ M, respectively). Cultivation of Hep G2 cells in the presence of 5 μ M concentration of either (I) or (II) resulted in significant reduction of cholesterol biosynthesis (52% and 57% from control), and also changes in biosynthesis of fatty acids, triglycerides, and cholesteryl esters. Compounds (I) and (II) stimulated transformation of exogenous cholesterol to polar products secreted into the culture medium (156 % and 175% of control) as it that was shown in experiments in Hep G2 cells prelabeled with [ 3 H]cholesterol.
Biochemistry Supplement Series B Biomedical Chemistry, 2008
This review is devoted to contemporary status of investigation of C-29 and C-28 plant sterols (ph... more This review is devoted to contemporary status of investigation of C-29 and C-28 plant sterols (phytosterols) in relation to their biological activity in mammals and mammalian cells. On the basis of experimental studies published during the last decade the following questions are discussed: phytosterols and nutrition; phytosterols and body cholesterol level; phytosterols and intestinal absorption of lipids, the role of phytosterols in lipid metabolism regulation; phytosterols and cultured mammalian cells; products of phytosterols oxidation; phytoecdysteroids and induced gene expression.
Biochemistry (Moscow) Supplement Series B: Biomedical Chemistry, 2008
25HC-25-hydroxycholesterol; ACAT-acyl-coenzyme A:cholesterol acyl transferase (formal systematic ... more 25HC-25-hydroxycholesterol; ACAT-acyl-coenzyme A:cholesterol acyl transferase (formal systematic name: SOAT sterol-3-O-acyl transferase; http://www.gene.ucl.ac.uk/cgi-bin/nomenclature/searchgenes.pl:); CE-cholesteryl esters; COcholesteryl oleate; FCS-fetal calf serum; PBS-phosphate buffered saline; PC-egg yolk phosphatidylcholine; PMSF-phenylmethylsulfonyl fluoride; VLDLvery low density lipoproteins.
Russian Journal of Bioorganic Chemistry, 2006
3β-Acetoxy-20-oxomethylpregn-5-ene and 3β-acetoxy-20-hydroxymethylpregn-5-ene were synthesized fr... more 3β-Acetoxy-20-oxomethylpregn-5-ene and 3β-acetoxy-20-hydroxymethylpregn-5-ene were synthesized from (22R, 23R)-sitost-5-ene-3β,22,23-triol in 66% overall yields.
Russian Journal of Bioorganic Chemistry, 2004
cal synthesis and characterization of palmitoyl derivatives of isomeric 15-oxygenated ∆ 8(14)-ste... more cal synthesis and characterization of palmitoyl derivatives of isomeric 15-oxygenated ∆ 8(14)-sterols (II), (IV), (VIII), (IX), (XV), and (XVI) (Schemes 1, 2). RESULTS AND DISCUSSION Acylation of 15-ketosterol (I) with palmitic acid in the presence of TPS and DMAP or N-methylimidazole by the method [14] resulted in palmitate (II) (Scheme 1) in yields 75-85%. To obtain 3 α-palmitoyloxy-5 αcholest-8(14)-en-15-one (IV), (I) was treated with MsCl in pyridine and, then, 3 β-mesylate (III) was converted into 3 α-palmitate (IV) by the reaction with cesium palmitate [16, 17]. The reaction in DMF [16] resulted in a better yield (72% after boiling for 40 min) in comparison with the modified procedure [17] [after the 12-h boiling with a cesium palmitate excess in toluene in the presence of 0.5 equiv of DMAP reacted only 15% of (III)]. An alkaline hydrolysis of palmitate (IV) led to the previously described [18] 3 α-hydroxy-5 α-cholest-8(14)-en-15-one, which unequivocally confirmed 3 α-configuration of (IV).
Russian Journal of Bioorganic Chemistry, 2004
Ergosteryl acetate was converted through three stages into 3 β-acetoxy-24-methyl-5 α-cholesta-8(1... more Ergosteryl acetate was converted through three stages into 3 β-acetoxy-24-methyl-5 α-cholesta-8(14),22-diene-15-one in 32% overall yield. The product was transformed to 3 β-hydroxy-24-methyl-5 αcholesta-8(14),22-diene-15-one, 3 α-hydroxy-24-methyl-5 α-cholesta-8(14),22-diene-15-one, and 24-methyl-5 α-cholesta-8(14),22-diene-3,15-dione. The compounds were characterized by 1 H and 13 C NMR spectra.
Bioorganicheskaia khimiia
The reduction of 3 beta-triphenylmethoxy-5 alpha-cholest-8(14)-en-15-one with lithium aluminum hy... more The reduction of 3 beta-triphenylmethoxy-5 alpha-cholest-8(14)-en-15-one with lithium aluminum hydride resulted in a quantitative yield of 3 beta-triphenylmethoxy-5 alpha-cholest-8(14)-en-15 beta-ol.
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Papers by Alexander Misharin