Fast liquid chromatography-mass spectrometry reveals side chain oxysterol heterogeneity in breast cancer tumour samples. Issue 192 (September 2019)

• Record Type: Journal Article
• Authors: Solheim, Stian; Hutchinson, Samantha A.; Lundanes, Elsa; Wilson, Steven R.; Thorne, James L.; Roberg-Larsen, Hanne
• Is Part Of: Journal of steroid biochemistry and molecular biology. Issue 192(2019)
• Publication Date: 2019-09
• Subjects: AFFL automatic filtration and filter back-flush -- BCa breast cancer -- ScOHC sidechain hydroxycholesterol; Oxysterols -- Breast cancer -- On-line sample clean-up -- 27-hydroxycholesterol -- Liquid chromatography-mass spectrometry; Steroid hormones -- Periodicals; Biochemistry -- Periodicals; Hormones -- Periodicals; Molecular Biology -- Periodicals; Hormones stéroïdes -- Périodiques; Steroid hormones; Periodicals; 572.579
• DOI: 10.1016/j.jsbmb.2019.02.004
• Languages: English
• ISSNs: 0960-0760

Abstract

Graphical abstract: Highlights: Fast LC–MS method for sensitive side chain oxysterol determination in breast cancer tumour samples. Multiple slices from same tumours reveals oxysterol heterogeneity. No significant differences in oxysterol content between ER-positive and ER-negative breast cancer. Correlation between esterified and free 27-hydroxycholesterol in breast cancer tumours. Abstract: Oxysterols can contribute to proliferation of breast cancer through activation of the Estrogen Receptors, and to metastasis through activation of the Liver X Receptors. Endogenous levels of both esterified and free sidechain-hydroxylated oxysterols were examined in breast cancer tumours from Estrogen Receptor positive and negative breast tumours, using a novel fast liquid chromatography tandem mass spectrometry method. Multiple aliquots of five milligram samples of 22 tumours were analysed for oxysterol content to assess intra- and inter-tumour variation. Derivatization was performed with Girard T reagent (with and without alkaline hydrolysis) and sample clean-up was performed using a robust automatic on-line column switching system ("AFFL"). Oxysterols were separated isocratically on a 2.1 mm inner diameter column packed with ACE SuperPhenylHexyl core shell particles using a mobile phase consisting of 0.1% formic acid in H2 O/methanol/acetonitrile (57/10/33, v/v/v) followed by a wash out step (0.1% formic acid in methanol/acetonitrile, 50/50, v/v). The total analysis time, including sample clean-up and column reconditioning, was 8 min (80% time reduction compared to other on-line systems). Analysis revealed large intra-tumour variations of sidechain oxysterols, resulting in no significant differences in endogenous oxysterols levels between Estrogen Receptor positive and Estrogen Receptor negative breast cancers. However, a correlation between esterified and free 27-hydroxycholesterol was observed. The same correlation was not observed for 24S-hydroxycholesterol or 25-hydroxycholesterol. The oxysterol heterogeneity of tumour tissue is a critical factor when assessing the role of these lipids in cancer.