Montrasio Cristina, Cheli Stefania, Clementi Emilio
Unit of Clinical Pharmacology, ASST Fatebenefratelli Sacco, L. Sacco University Hospital, Milan, Italy.
Clinical Pharmacology Unit, Department of Biomedical and Clinical Sciences, L. Sacco University Hospital, Università degli Studi di Milano, Milan, Italy.
Pharmgenomics Pers Med. 2023 Jul 27;16:739-746. doi: 10.2147/PGPM.S412430. eCollection 2023.
The application of pharmacogenetics in oncology is part of the routine clinical practice. In particular, genotyping of dihydropyrimidine dehydrogenase (DPYD) and UDP-glucuronosyltransferase (UGT1A1) is crucial to manage the treatment of patients taking fluoropyrimidines and irinotecan. The unique approach of our laboratory to the pharmacogenetic diagnostic service in oncology is to combine two real-time PCR methods, LightSNiP assay (TIB MOLBIOL), and more recently FRET (Fluorescent Resonance Energy Transfer) probes technology (Nuclear Laser Medicine), plus TaqMan assay (Thermo Fisher) for the confirmation of the presence of variant alleles on DNA from a second extraction. We found that both the FRET and LightSNiP assays, where detection occurs by melting curve analysis, offer an advantage over the competing TaqMan technology. Whereas unexpected genetic variants may be missed using a mutation-specific TaqMan assay, the information thus obtained can be useful to adjust the therapy in case of unexpected post-treatment toxicity. The combination of TaqMan and FRET assays helped us to achieve more accurate genotyping and a correct result for the patient. The added value of the DPYD FRET assay is the possibility of detecting, with the same amplification profile of the polymorphisms detailed in the guidelines, also the c.2194G>A (*6 rs1801160), cited in the recommendations as a variant to be investigated in case of severe toxicity. Regarding the UGT1A1 (TA)n promoter polymorphism (rs3064744), the distinctive and positive feature of the FRET assay is to allow clearly identifying all those potential variant alleles, including the (TA)5 and (TA)8 alleles, that are frequent in African Americans. Our clinical practice emphasizes the importance of not only rapid and easy-to-use assays, such as the new FRET ones, but also of accurate and comprehensive genotyping for good pharmacogenetic diagnostic activity.
药物遗传学在肿瘤学中的应用是常规临床实践的一部分。特别是，二氢嘧啶脱氢酶（DPYD）和尿苷二磷酸葡萄糖醛酸转移酶（UGT1A1）的基因分型对于管理接受氟嘧啶和伊立替康治疗的患者至关重要。我们实验室在肿瘤学药物遗传学诊断服务方面的独特方法是将两种实时PCR方法相结合，即LightSNiP检测法（TIB MOLBIOL），以及最近的荧光共振能量转移（FRET）探针技术（Nuclear Laser Medicine），再加上TaqMan检测法（赛默飞世尔科技），用于从第二次提取的DNA中确认变异等位基因的存在。我们发现，通过熔解曲线分析进行检测的FRET和LightSNiP检测法都比竞争的TaqMan技术具有优势。虽然使用突变特异性TaqMan检测法可能会遗漏意外的基因变异，但在出现意外的治疗后毒性时，由此获得的信息有助于调整治疗方案。TaqMan检测法和FRET检测法的结合帮助我们实现了更准确的基因分型，并为患者得出了正确的结果。DPYD FRET检测法的附加价值在于，在与指南中详细描述的多态性具有相同扩增图谱的情况下，还能够检测到c.2194G>A（*6 rs1801160），该变异在建议中被列为在严重毒性情况下需要研究的变异。关于UGT1A1（TA）n启动子多态性（rs3064744），FRET检测法的独特且积极的特点是能够清晰识别所有那些潜在的变异等位基因，包括在非裔美国人中常见的（TA）5和（TA）8等位基因。我们的临床实践强调了不仅像新型FRET检测法这样快速且易于使用的检测方法的重要性，还强调了准确和全面的基因分型对于良好的药物遗传学诊断活动的重要性。
