Microarray Approach Combined with ddPCR: An Useful Pipeline for the Detection and Quantification of Circulating Tumour DNA Mutations

Cells

Luca Gianni¹  Valentina Burgio¹  Monica Ronzoni¹  Silvia Galbiati²  Nadia Soriani²  Maurizio Ferrari²  Marcella Chiari³  Lucia Ferraro³  Francesco Damin³ 

[1] Dipartimento di Oncologia Medica, IRCCS Ospedale San Raffaele, 20132 Milan, Italy;Genomic Unit for the Diagnosis of Human Pathologies, Division of Genetics and Cell Biology, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy;Istituto di Chimica del Riconoscimento Molecolare, Consiglio Nazionale delle Ricerche, 20131 Milan, Italy;

关键词: ctDNA;    liquid biopsy;    metastatic colorectal cancer;    microarray;    ddPCR;   

DOI  :  10.3390/cells8080769

来源: DOAJ

【 摘 要 】

It has now been established that in biological fluids such as blood, it is possible to detect cancer causing genomic alterations by analysing circulating tumour DNA (ctDNA). Information derived from ctDNA offers a unique opportunity to enrich our understanding of cancer biology, tumour evolution and therapeutic efficacy and resistance. Here, we propose a workflow to identify targeted mutations by a customized microarray-based assay for the simultaneous detection of single point mutations in different oncogenes (KRAS, NRAS and BRAF) followed by droplet digital PCR (ddPCR) to determine the fractional abundance of the mutated allele. Genetic variants were determined in the plasma of 20 metastatic colorectal cancer (mCRC) patients previously genotyped on tissue biopsy at the diagnosis for medication planning (T0) and following the tumour genetic evolution during treatment phase (T1 and T2) with the objective of allowing therapy response prediction and monitoring. Our preliminary results show that this combined approach is suitable for routine clinical practice. The microarray platform enables for a rapid, specific and sensitive detection of the most common mutations suitable for high-throughput analysis without costly instrumentation while, the ddPCR, consents an absolute quantification of the mutated allele in a longitudinal observational study on patients undergoing targeted therapy.

【 授权许可】

Unknown