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Cancers 2019, 11(4), 454; https://doi.org/10.3390/cancers11040454

Assessment of a High Sensitivity Method for Identification of IDH1 R132x Mutations in Tumors and Plasma of Intrahepatic Cholangiocarcinoma Patients

1
Cancer Genomics Laboratory, Fondazione Edo ed Elvo Tempia, Via Malta 3, 13900 Biella, Italy
2
Laboratory of Molecular Oncology, Fondazione Edo ed Elvo Tempia, Via dei Ponderanesi 2, 13875 Ponderano, Biella, Italy
3
Department of Oncology, University of Turin, 10100 Torino, Italy
4
Department of Experimental and Clinical Medicine, University of Firenze, 50100 Firenze, Italy
5
Center for Autoimmune Liver Diseases, Humanitas Clinical and Research Center, 20089 Rozzano, Italy
6
Department of Medical Sciences, University of Turin, 10100 Torino, Italy
7
Pathology Unit, Candiolo Cancer Institute-FPO-IRCCS, Candiolo, 10060 Torino, Italy
8
Division of Medical Oncology, Candiolo Cancer Institute, FPO-IRCCS, Candiolo, 10060 Torino, Italy
*
Author to whom correspondence should be addressed.
These authors contribute equally to this work.
Received: 11 March 2019 / Revised: 26 March 2019 / Accepted: 28 March 2019 / Published: 30 March 2019
(This article belongs to the Special Issue Models of Experimental Liver Cancer)
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PDF [2295 KB, uploaded 30 March 2019]
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Abstract

Hotspot codon 132 mutations (R132xIDH1m) are frequent in intrahepatic cholangiocarcinoma (ICC), are druggable by anti-IDH1m agents, and could represent a marker of disease progression. Developing an assay to identify R132xIDH1m would provide a useful tool to select patients benefitting from targeted treatments. We tested a quantitative real-time allele-specific polymerase chain reaction (qPCR)-based method to detect the main R132xIDH1m in an Italian ICC series (n = 61) of formalin-fixed paraffin-embedded (FFPE) samples, and on circulating-free DNA samples. The outcomes were compared with nested PCR/Sanger sequencing. Reconstitution experiments of plasmids harboring the different R132xIDH1m mixed with wild-type (WT) DNA demonstrated that qPCR is able to detect at least 2% of all mutated allele. High efficiency was also observed on patient-derived mutated DNA mixed with WT DNA (up to 10% and 0.3 ng of mutated template); qPCR detected 16.4% of mutated samples (one R132G, three R132C and six R132L) while nested PCR/Sanger sequencing only 8.2% (four R132L and one R132G). In a single patient with an R132C-mutated tumor, qPCR was also performed on plasma samples collected at four time-points, observing an increase correlating with disease progression. In conclusion, we developed a qPCR assay which could represent a fast, inexpensive and sensitive tool both for detection of R132xIDH1m in ICC samples and monitoring disease progression from liquid biopsy. View Full-Text
Keywords: IDH1 mutation; intrahepatic cholangiocarcinoma; qPCR; liquid biopsy; biomarker IDH1 mutation; intrahepatic cholangiocarcinoma; qPCR; liquid biopsy; biomarker
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Peraldo-Neia, C.; Scatolini, M.; Grosso, E.; Lombardi, P.; Filippi, R.; Raggi, C.; Marchiò, C.; Cavalloni, G.; Aglietta, M.; Leone, F. Assessment of a High Sensitivity Method for Identification of IDH1 R132x Mutations in Tumors and Plasma of Intrahepatic Cholangiocarcinoma Patients. Cancers 2019, 11, 454.

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