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Structural basis for the selective inhibition of human 3beta-hydroxysteroid dehydrogenase 1 in human breast tumor
James L Thomas1, Kevin M Bucholtz, Jingping Sun
1Division of Basic Medical Sciences, Mercer University School of Medicine, Mercer University, 1550 College Street, Macon, GA 31207, USA. thomas.j@Mercer.edu
Abstract:
Human 3beta-hydroxysteroid dehydrogenase/isomerase type 1 (3beta-HSD1) is a critical enzyme in the conversion of DHEA to estradiol in breast tumors and may be a target enzyme for inhibition in the treatment of breast cancer in postmenopausal women. Human 3beta-HSD2 participates in the production of cortisol and aldosterone in the human adrenal gland in this population. In our recombinant human breast tumor MCF-7 Tet-off cells that express either 3beta-HSD1 or 3beta-HSD2, trilostane and epostane inhibit the DHEA-induced proliferation of MCF-7 3beta-HSD1 cells with 12- to 16-fold lower IC(50) values compared to the MCF-7 3beta-HSD2 cells. The compounds also competitively inhibit purified human 3beta-HSD1 with 12- to 16-fold lower K(i) values compared to the noncompetitive K(i) values measured for human 3beta-HSD2. Using our structural model of 3beta-HSD1, trilostane or 17beta-acetoxy-trilostane was docked in the active site of 3beta-HSD1, and Arg195 in 3beta-HSD1 or Pro195 in 3beta-HSD2 was identified as a potentially critical residue (one of 23 non-identical residues in the two isoenzymes). The P195R mutant of 3beta-HSD2 were created, expressed and purified. Kinetic analyses of enzyme inhibition suggest that the high affinity, competitive inhibition of 3beta-HSD1 by trilostane and epostane may be related to the presence of Arg195 in 3beta-HSD1 vs. Pro195 in 3beta-HSD2.
Insights
Trilostane and epostane effectively inhibit human 3beta-hydroxysteroid dehydrogenase/isomerase type 1 (3beta-HSD1), an enzyme crucial for breast tumor growth. This selective inhibition, linked to a key amino acid difference, offers a potential therapeutic strategy for breast cancer treatment.
Area of Science:
- Biochemistry
- Enzymology
- Pharmacology
Background:
- Human 3beta-hydroxysteroid dehydrogenase/isomerase type 1 (3beta-HSD1) is vital for converting DHEA to estradiol in breast tumors.
- Inhibition of 3beta-HSD1 is a potential therapeutic strategy for postmenopausal breast cancer.
- Human 3beta-HSD2 plays a role in cortisol and aldosterone production in the adrenal gland.
Purpose of the Study:
- To investigate the inhibitory effects of trilostane and epostane on human 3beta-HSD1 and 3beta-HSD2.
- To explore the structural basis for the differential inhibition of 3beta-HSD1 and 3beta-HSD2 by these compounds.
- To assess the potential of 3beta-HSD1 as a drug target for breast cancer treatment.
Main Methods:
- Utilized recombinant human breast tumor MCF-7 Tet-off cells expressing 3beta-HSD1 or 3beta-HSD2.
- Performed enzyme inhibition assays to determine IC(50) and K(i) values.
- Employed molecular docking studies using a structural model of 3beta-HSD1.
- Created and kinetically analyzed a Pro195Arg mutant of 3beta-HSD2.
Main Results:
- Trilostane and epostane showed 12- to 16-fold lower IC(50) values in inhibiting 3beta-HSD1 compared to 3beta-HSD2.
- Competitive inhibition was observed for 3beta-HSD1, while noncompetitive inhibition was noted for 3beta-HSD2.
- Molecular modeling identified Arg195 in 3beta-HSD1 and Pro195 in 3beta-HSD2 as critical residues.
- Kinetic analysis of the P195R mutant supported the role of this residue in differential inhibition.
Conclusions:
- Trilostane and epostane selectively and competitively inhibit human 3beta-HSD1.
- The differential inhibition is attributed to the presence of Arg195 in 3beta-HSD1 versus Pro195 in 3beta-HSD2.
- These findings support the targeting of 3beta-HSD1 for breast cancer therapy.
