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Validation of a routine gas chromatography mass spectrometry method for 2-hydroxyglutarate quantification in human serum as a screening tool for detection of idh mutations
Journal of Chromatography B ( IF 3 ) Pub Date : 2018-02-28 , DOI: 10.1016/j.jchromb.2018.02.038
Esther Fernández-Galán , Núria Massana , Marina Parra-Robert , Susana Hidalgo , Gregori Casals , Jordi Esteve , Wladimiro Jiménez

High circulating levels of 2-hydroxyglutarate (2HG) have been reported in patients with determinate isocitrate dehydrogenase (IDH) mutated tumors. Recent studies indicate that in malignancies such as acute myeloid leukemia (AML), measurements of 2HG in serum provide useful diagnostic and prognostic information and improve patient selection and monitoring of IDH-targeted treatments. In the current study, we validated a sensitive and specific gas chromatography mass spectrometry (GC–MS) method specifically intended to quantify serum levels of 2HG in routine clinical laboratories. Extraction was liquid-liquid with ethyl acetate, and derivatization was reduced to 3 min of microwave irradiation. The analytical method was linear over a wide dynamic range, presenting acceptable intraday and day-to-day precision and accuracy. The limit of quantification was 10 ng/mL, process efficiency ranged between 38% and 49%, and recovery of added 2HG was 99–105%. 2HG was found to be stable in serum for up to 48 h at both 4 °C and at ambient temperature, and after three freeze-thaw cycles. Microwave derivatizated extracts in the autosampler were found to be stable for up to 120 h. In summary, the present method is useful for quantification of 2HG serum levels in patients with IDH mutated malignancies in clinical laboratories.



中文翻译:

常规气相色谱质谱法用于人血清中2-羟基戊二酸定量作为验证IDH突变的筛选工具的验证

据报道,患有确定的异柠檬酸脱氢酶(IDH)突变的肿瘤患者体内高水平的2-羟基戊二酸(2HG)循环水平。最近的研究表明,在诸如急性髓细胞性白血病(AML)的恶性肿瘤中,血清中2HG的测定可提供有用的诊断和预后信息,并改善患者选择和监测IDH靶向治疗的情况。在当前的研究中,我们验证了灵敏且特异性的气相色谱质谱(GC-MS)方法,该方法专门用于定量常规临床实验室中2HG的血清水平。用乙酸乙酯液-液萃取,并且衍生化减少至微波辐射3分钟。该分析方法在很宽的动态范围内是线性的,呈现出可接受的日内和日常精度和准确性。定量限为10 ng / mL,工艺效率介于38%和49%之间,添加的2HG的回收率为99–105%。发现2HG在4°C和环境温度下以及经过三个冻融循环后,在血清中最多可稳定48小时。发现自动进样器中的微波衍生提取物可稳定长达120小时。总之,本方法可用于在临床实验室中对具有IDH突变的恶性肿瘤患者的2HG血清水平进行定量。

更新日期:2018-02-28
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