Mohammad Alyamani (Presenter)
Cleveland State University
Bio: I'm a graduate student whose research focuses on prostate cancer treatment. The main objective of my research is to study the metabolism of steroids and some therapeutic agents used in the treatment of Castration-Resistant Prostate Cancer (CRPC) and its effect on the androgen receptor (AR); which are important for the growth of prostate cancer cells. I'm studying the metabolism by LC-MS/MS methods using CRPC patients’ serum. We assume that knowing more about steroids metabolism may solve the puzzle of the resistance for current therapy in treating prostate cancer.
Authorship: Mohammad Alyamani, MSc.(1,2) Zhenfei Li, PhD.(2) Nima Sharifi, MD.(1,2)
(1) Cleveland State University, OH (2) Cleveland Clinic, OH
| Short Abstract Advanced prostate cancer is the second leading cause of cancer-related deaths among American men. The androgen receptor (AR) is vital for prostate cancer progression. Abiraterone acetate (AA) prolongs survival in castration-resistant prostate cancer (CRPC) by blocking CYP17A1, an enzyme required for androgen synthesis, but resistance invariably occurs. We hypothesize that abiraterone (abi) is endogenously converted by the enzyme 3β-hydroxysteroid dehydrogenase/isomerase (3βHSD) to a novel metabolite delta-4-abiraterone (D4A). We developed a liquid chromatography/tandem mass spectrometry (LC-MS/MS) method to analyze abi and its metabolite. Analysis of serum from CRPC patients treated with AA showed that abi is converted to the novel metabolite D4A. |
Long Abstract
Introduction
Advanced prostate cancer (PCa) is the second cause of cancer related deaths among American men. The estimated number of new PCa cases is about 220,000 and about 27,000 deaths due to PCa in the United States in 2015. Tumor progression is driven by androgen receptor (AR). The activation of AR depends on two steroidal hormones; Testosterone (T) and dihydrotestosterone (DHT), which act as fuel for malignant cell growth; DHT is more potent than T as an agonist for AR. The first line for advanced prostate cancer treatment is androgen deprivation therapy (ADT) either by medical or surgical castration. In many cases and despite the low serum T levels, the cancer become resistant to this treatment and develops castration-resistance prostate cancer (CRPC). In CRPC, DHT can be synthesized either de novo from cholesterol or from adrenal steroids. In both cases DHT formation is achieved via several enzymatic reactions.
Abiraterone, (3β)-17-(pyridine-3-yl)androsta-5,16-diene-3-ol (abi), is a steroidal compound that is used in the treatment of CRPC, it is administered as the prodrug abiraterone acetate (AA). Abi inhibits CYP17A1, an enzyme required for androgen biosynthesis. We hypothesized that the similarity in structure between abi and dehydroepiandrosterone (DHEA) makes abi a possible substrate of the enzyme 3 beta hydroxysteroid dehydrogenase/isomerize (3βHSD) and converts it to 3 keto delta-4-abi (D4A) . In vitro studies in our lab on prostate cancer cell lines show that D4A is more potent than abi in blocking CYP17A1; it also inhibits other enzymes essential for DHT synthesis. Furthermore D4A is an AR antagonist(1).
Methods
Serum samples were collected from CRPC patients treated with AA. Liquid chromatography mass spectrometry method was developed to detect abi and D4A using AB Sciex Qtrap 5500 mass analyzer coupled with a Shimadzu Nexera UPLC station. Electrospray ionization in positive mode was applied and the analytes were separated using a Zorbax Eclipse Plus C18 150*2.1mm, 3.5µm column at 30°C, and isocratic mobile phase 35% A (formic acid) , 65% B (methanol:acetonitrile; 60:40) and flow rate 0.2 ml/min, with injection volume 10µl. Data were processed using Analyst 1.6.2 software (AB Sciex). Analytes and the IS (Abirateronel-d4) were extracted from 100µl human serum with 2 ml methyl tert-butyl ether. After evaporation, the residue was reconstituted using 300 µl 50% methanol. For quantitation purpose, a 7-point calibration curve for both analytes was constructed, and six quality control samples (2 from each level, low, mid, and high) were injected with the samples.
Results
The precursor/product ions and retention times for abi and D4A were m/z 350.3/156.1 (6.75 min) and 348.2/156.3 (6.18 min), respectively. Analysis in human serum showed that abi was linear in the range of 2.0-400 ng/ml, and D4A was linear in the range of 0.1-20 ng/ml. In all samples obtained from CRPC patients, abi and D4A were both detected.
In conclusion, abi is endogenously converted by 3βHSD to the more potent analogue D4A.
References & Acknowledgements:
1. Li Z, Bishop AC, Alyamani M et al. Conversion of abiraterone to D4A drives anti-tumour activity in prostate cancer. Nature 2015; doi: 10.1038/nature14406.
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