Whole-cell fungal-mediated structural transformation of anabolic drug metenolone acetate into potent anti-inflammatory metabolites

Seven new derivatives, 6 alpha-hydroxy-1-methyl-3-oxo-5 alpha-androst-1-en-17-yl acetate (2), 6 alpha,17 beta-dihydroxy-1-methyl-3-oxo-5 alpha-androst-1-en (3), 7 beta-hydroxy-1-methyl-3-oxo-5 alpha-androst-1-en-17-yl acetate (4), 15 beta,20-dihydroxy-1-methyl-3-oxo-5 alpha-androst-1-en-17-yl acetate (5), 15 beta-hydroxy-1-methyl-3-oxo-5 alpha-androst-1-en-17-yl acetate (6), 12 beta,17 beta-dihydroxy-1-methyl-3-oxoandrosta-1,4-dien (11), and 7 beta,15 beta,17 beta-trihydroxy-1-methyl-3-oxo-5 alpha-androst-1-en (14), along with six known metabolites, 17 beta-hydroxy-1-methyl-3-oxoandrosta-1,4-dien (7), 17 beta-hydroxy-1-methyl-3-oxo-5 alpha-androst-1en (8), 17 beta-hydroxy-1-methyl-3-oxo-5 beta-androst-1-en (9), 1-methyl-5 beta-androst-1-en-3,17-dione (10), 1-methyl-3-oxoandrosta-1,4-dien-3,17-dione (12), and 17 beta-hydroxy-1 alpha-methyl-5 alpha-androstan-3-one (13) of metenolone acetate (1), were synthesized through whole-cell biocatalysis with Rhizopus stolonifer, Aspergillus alliaceous, Fusarium lini, and Cunninghamella elegans. Atamestane (12), an aromatase inhibitor, was synthesized for the first time via F. lini-mediated transformation of 1 as the major product. Hydroxylation, dehydrogenation, and reduction were occurred during biocatalysis. Study indicated that F. lini was able to catalyze dehydrogenation reactions selectively. Structures of compounds 1-14 were determined through NMR, HRFAB-MS, and IR spectroscopic data. Compounds 1-14 were identified as non-cytotoxic against BJ human fibroblast cell line (ATCC CRL-2522). Metabolite 5 (81.0 +/- 2.5%) showed a potent activity against TNF-alpha production, as compared to the substrate 1 (62.5 +/- 4.4%). Metabolites 2 (73.4 +/- 0.6%), 8 (69.7 +/- 1.4%), 10 (73.2 +/- 0.3%), 11 (60.1 +/- 3.3%), and 12 (71.0 +/- 7.2%), also showed a good inhibition of TNF-alpha production. Compounds 3 (IC50 = 4.4 +/- 0.01 mu g/mL), and 5 (IC50 = 10.2 +/- 0.01 mu g/mL) showed a significant activity against T-cell proliferation. Identification of selective inhibitors of TNF-alpha production, and T-cell proliferation is a step forward towards the development of anti-inflammatory drugs. (C) 2020 The Authors. Published by Elsevier B.V. on behalf of Cairo University. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).