Determination of 17α-hydroxypregnenolone in human plasma by routine isotope dilution mass spectrometry using benchtop gas chromatography-mass selective detection

Wudy, Stefan A.; Hartmann, Michaela F.; Solleder, Claudia; Homoki, J.

doi:10.1016/s0039-128x(01)00109-x

Cited by 15 publications

(4 citation statements)

References 12 publications

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“…Pentafluorophenyldimethylsilyl-TMS ether (flophemesyl) derivatives give intense molecular ions in MS that are attractive for GC-MS analysis (Choi and Chung, 1999;Choi et al 2001). Heptafluorobutyrates have been used for quantitative analysis of several steroids (Scherer et al, 1998;Wudy et al, 2000Wudy et al, , 2001Wudy et al, , 2002. Bismethylenedioxypentafluoropropionate derivatives have been used in the analysis of cortisol metabolites (Furuta et al, 2000a, b).…”

Section: Derivative Formationmentioning

confidence: 99%

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Makin

Honour

Shackleton³

et al. 2010

Steroid Analysis

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Section: Derivative Formationmentioning

confidence: 99%

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Makin

Honour

Shackleton³

et al. 2010

Steroid Analysis

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“…In addition, 4-androstenedione, cortisol, DHEA, 11-deoxycortisol, 17-hydroxypregnenolone (17OHPreg), and 17-hydroxyprogesterone (17OHP) were determined by isotope dilution gas chromatography–mass spectrometry (GC–MS) as described elsewhere (Wudy et al, 2001; Wudy and Hartmann, 2004). Briefly, cell supernatant was equilibrated with internal standard (deuterium-labeled analogs of analytes).…”

Section: Methodsmentioning

confidence: 99%

Sunitinib inhibits cell proliferation and alters steroidogenesis by down-regulation of HSD3B2 in adrenocortical carcinoma cells

Kroiß¹

2011

Front. Endocrin.

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The multi-tyrosine kinase inhibitor sunitinib is used in the treatment of several solid tumors. Animal experiments pointed to an adrenotoxic effect of sunitinib. Therefore, we evaluated the expression of key targets of sunitinib in human adrenocortical carcinoma (ACC) tumor samples and investigated its in vitro effects in ACC cell lines. We carried out immunohistochemistry for vascular endothelial growth factor (VEGF) and its receptor (VEGF-R2) in 157 ACC samples and nine normal adrenal glands. VEGF and VEGF-R2 protein were expressed in 72 and 99% of ACC samples, respectively. Using NCI-H295 and SW13 ACC cell lines, we investigated the effects of sunitinib on cell proliferation. Sunitinib reduced dose-dependently cell viability of both NCI-H295 and SW13 cells (SW13: 0.1 μM 96 ± 7%, 1 μM 90 ± 9%*, 5 μM 62 ± 6%*, controls 100 ± 9%; *p < 0.05). To determine sunitinib effects on steroidogenesis, we measured steroid hormones in cell culture supernatant by gas chromatography–mass spectrometry. We observed a pronounced decrease of cortisol secretion (1 μM 90.1 ± 1.5%*, 5 μM 57.2 ± 0.3%*, controls 100 ± 2.4%) and a concomitant increase in the DHEA/4-androstenedione and 17-hydroxypregnenolone/17-hydroxyprogesterone ratios, indicating specific inhibition of 3β-hydroxysteroid dehydrogenase (HSD3B2). In yeast microsomes transformed with HSD3B2, no direct inhibition of HSD3B2 by sunitinib was detected. Sunitinib induced down-regulation of HSD3B2 mRNA and protein in ACC cell lines (mRNA: 1 μM 44 ± 16%*; 5 μM 22 ± 2%*; 10 μM 19 ± 4%*; protein: 1 μM 82 ± 8%; 5 μM 63 ± 8%*; 10 μM 55 ± 9%*). CYP11B1 was down-regulated at mRNA but not at protein level and CYP11A1 remained unchanged. In conclusion, target molecules of sunitinib are expressed in the vast majority of ACC samples. Sunitinib exhibits anti-proliferative effects in vitro, and appears to specifically block adrenal steroidogenesis by down-regulation of HSD3B2, rendering it a promising option for treatment of ACC.

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“…This was a difficult detector to keep for routine use and will not be considered here. GC-MS has been found also to be a very reliable quantitative method for steroids in plasma such as DHAS, 17-OHP, AS, testosterone, 17-hydroxpregnenolone, 11-deoxycortisol and DHAS (7, 8, 9, 10, 11, 12, 13). It is superior to other methods, because of the improved specificity of assays through separation of steroids in the sample extract, which is needed particularly for tests on newborn infants.…”

Section: Gas Chromatographymentioning

confidence: 99%

“…For steroids, those reference methods have relied upon stable-isotope dilution analyses with GC coupled with mass spectrometry (MS) (6, 7, 8). The dual selectivity of the separation of a mixture of components and specific ion detection enables exceptionally high accuracy without interferences (9, 10, 11, 12, 13). Some immunoassays have approached that level of accuracy, but only when the method includes extraction and chromatographic purification, again limited to very specialised laboratories (14, 15).…”

Section: Introductionmentioning

confidence: 99%

Steroid Assays in Paediatric Endocrinology

Honour¹

2010

JCRPE

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Most steroid disorders of the adrenal cortex come to clinical attention in childhood and in order to investigate these problems, there are many challenges to the laboratory which need to be appreciated to a certain extent by clinicians. The analysis of sex steroids in biological fluids from neonates, over adrenarche and puberty present challenges of specificities and concentrations often in small sample sizes. Different reference ranges are also needed for interpretations. For around 40 years, quantitative assays for the steroids and their regulatory peptide hormones have been possible using immunoassay techniques. Problems are recognised and this review aims to summarise the benefits and failings of immunoassays and introduce where tandem mass spectrometry is anticipated to meet the clinical needs for steroid analysis in paediatric endocrine investigations. It is important to keep a dialogue between clinicians and the laboratory, especially when any laboratory result does not make sense in the clinical investigation.Conflict of interest:None declared.

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Determination of 17α-hydroxypregnenolone in human plasma by routine isotope dilution mass spectrometry using benchtop gas chromatography-mass selective detection

Cited by 15 publications

References 12 publications

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

General Methods for the Extraction, Purification, and Measurement of Steroids by Chromatography and Mass Spectrometry

Sunitinib inhibits cell proliferation and alters steroidogenesis by down-regulation of HSD3B2 in adrenocortical carcinoma cells

Steroid Assays in Paediatric Endocrinology

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