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Androgenic biomarker profiling in human matrices and cell culture samples using high throughput, electrospray tandem mass spectrometry
Author(s) -
Wilton John H.,
Titus Mark A.,
Efstathiou Eleni,
Fetterly Gerald J.,
Mohler James L.
Publication year - 2014
Publication title -
the prostate
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.295
H-Index - 123
eISSN - 1097-0045
pISSN - 0270-4137
DOI - 10.1002/pros.22792
Subject(s) - chromatography , chemistry , tandem mass spectrometry , mass spectrometry , dehydroepiandrosterone , lncap , electrospray ionization , triple quadrupole mass spectrometer , selected reaction monitoring , liquid chromatography–mass spectrometry , metabolite , dihydrotestosterone , high performance liquid chromatography , intracrine , electrospray , epitestosterone , endocrinology , androgen , medicine , prostate cancer , hormone , biochemistry , cancer , receptor , paracrine signalling
BACKGROUND A high throughput, high pressure liquid chromatographic (HPLC) method with triple quadrupole mass spectral detection (LC/MS/MS) was validated for the measurement of 5 endogenous androgens in human plasma and serum and applied to various in vivo and in vitro study samples to pursue a better understanding of the interrelationship of the androgen axis, intracrine metabolism, and castration‐recurrent prostate cancer (CaP). METHODS A Shimadzu HPLC system interfaced with a Sciex QTRAP 5500 mass spectrometer with electrospray ionization was used with inline column‐switching. Samples were liquid/liquid extracted and chromatographed on a Luna C18(2) column at 60°C with a biphasic gradient using a 15‐min run time. RESULTS The method was validated for five androgens in human plasma and serum, and applied to four sets of samples. Plasma (n = 188) and bone marrow aspirate (n = 129) samples from patients with CaP, who received abiraterone acetate plus prednisone for up to 945 days (135 weeks), had undetectable androgens after 8 weeks of treatment. Plasma dehydroepiandrosterone (DHEA) concentrations were higher in African Americans than Caucasian Americans with newly diagnosed CaP. Analysis of prostate tumor tissue homogenates demonstrated reproducible testosterone (T) and dihydrotestosterone (DHT) concentrations with a minimal sample size of ∼1.0–2.0 mg of tissue. Finally, cell pellet and media samples from the LNCaP C4‐2 cell line showed conversion of T to DHT. CONCLUSION The proposed LC/MS/MS method was validated for quantitation of five endogenous androgens in human plasma and serum, and effectively profiles androgens in clinical specimens and cell culture samples. Prostate 74:722–731, 2014 . © 2014 Wiley Periodicals, Inc.