Effects of 3β-HSD1 on back-conversion and androgen signaling in CRPC.

Abstract

238 Background: Dihydrotestosterone (DHT), the principal androgen driving prostate cancer progression, is synthesized in the prostate from gonadal and adrenal precursors. It is then metabolized to downstream 5α-reduced androgens to be eliminated. 3β-Hydroxysteroid Dehydrogenase type 1 (3β-HSD1) catalyzes C3 oxidation in androgen synthesis. Inheritance of the adrenal-permissive HSD3B1 allele prolongs enzyme stability and increases DHT synthesis from adrenal precursors, contributing to castration resistance and prostate cancer mortality. Abiraterone acetate (AA) is a steroidal drug that may also be subject to 3β-HSD1 metabolism. We investigated whether 3β-HSD1 mediates the back-conversion of 3β-androstanediol (3β-diol) to DHT and 3β-OH-abiraterone (3β-OH-Abi) to 3-keto-abiraterone (3-keto-Abi), a weak androgen receptor agonist, thereby potentially promoting CRPC progression. Methods: Vector control and HSD3B1 -overexpressing 293T cells were treated with 3β-diol or 3β-OH-Abi. Genetic and pharmacologic blockade of 3β-HSD1 in C4-2 CRPC cells were used to assess DHT metabolism and back-conversion of 3β-diol to DHT via mass spectrometry. Expression of androgen-responsive genes (ARGs) was measured in parental and CRISPR-knockdown cells following 3β-diol treatment. In the clinical setting, serum 3β-OH-Abi levels were compared between adrenal-permissive and adrenal-restrictive HSD3B1 genotypes across treatment cycles in the PANTHER trial (mCRPC patients treated with apalutamide and AA + prednisone). Results: Overexpression of HSD3B1 in 293T cells enabled back-conversion of 3β-diol to DHT. These cells also back-converted 3β-OH-Abi to 3-keto-Abi. C4-2 cells metabolize DHT to both C3 stereoisomers of androstanediol. Pharmacologic inhibition with trilostane blocked 3β-HSD1-mediated conversion of 3β-diol to DHT. Similarly, back-conversion of 3β-diol was abolished in HSD3B1 -knockdown cells, as DHT was detected in both media and intracellularly in parental C4-2 cells following treatment with 3β-diol, but was absent in knockdown cells. Also, 3β-diol treatment upregulated ARGs in parental cells. Pharmacokinetic data from the PANTHER trial demonstrated higher serum 3β-OH-Abi levels in men with the adrenal-restrictive genotype at 4- and 8-hour timepoints during treatment cycles 1 and 2. Conclusions: 3β-HSD1 activity sustains intra-tumoral DHT by back-converting its downstream inactive metabolite, 3β-diol, thereby maintaining androgen signaling activity. This pathway also mediates back-conversion of 3β-OH-Abi to 3-keto-Abi, a weak androgen receptor agonist. These findings provide further mechanistic insight into the adverse outcomes associated with the adrenal-permissive HSD3B1 genotype and suggest new avenues for CRPC management.