17/06-2019 - Alexandre How-Kit : Improved detection and identification of microsatellite instability by new molecular diagnostic methods: application to colorectal cancer patients

15 - Avril - 2019

LES LUNDIS DE SAINT-ANTOINE

Bâtiment Kourilsky - 13h–14h

Salle des Conférences (Rez de Chaussée),

184 rue du Faubourg Saint-Antoine, Paris

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LUNDI 17 JUIN 2019

Improved detection and identification of microsatellite instability by new molecular diagnostic methods: application to colorectal cancer patients

Alexandre How-Kit

CEPH

Equipe Duval invité par Ada Collura

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Microsatellite instability (MSI) is a genomic alteration in which microsatellites, usually of one to four nucleotide repeats, accumulate mutations corresponding to deletions/insertions of a few nucleotides. The MSI phenotype has been extensively characterized in colorectal cancer and is due to a deficiency of the DNA mismatch repair system. MSI has recently been shown to be present in most types of cancer with variable frequencies (from <1% to 30%). It correlates positively to survival outcome and predicts the response to immune checkpoint blockade therapy in solid cancers. The evaluation of MSI status is based on screening tumor DNA for the presence of somatic deletions in DNA repeats using PCR followed by fragment analysis. However this method can lack sensitivity due to the presence of a high level of germline DNA frequently contaminating the core of primary colon tumors. We have developed two new approaches for the sensitive detection of MSI in CRC known as NaMSIE (Nuclease-assisted MicroSatellite Insability Enrichment) and E-ice-COLD-PCR (Enhanced-improved and complete enrichment-CO-amplification at Lower Denaturation temperature-PCR) that are based on the enrichment of mutant alleles prior to and during PCR amplification respectively. We used HSP110 T17, a mononucleotide DNA repeat that was previously proposed as an optimal marker to detect MSI in tumor DNA. Compared to standard PCR, NaMSIE and E-ice-COLD-PCR allowed a strong decrease of the limit of detection of MSI (0.5% and 0.05% respectively), thus greatly improving MSI detection in CRC samples highly contaminated with normal DNA. Both molecular approaches are a rapid, cost-effective, easy-to-implement, and highly sensitive. They may deeply improve the detection of MSI in routine clinics.

Centre de Recherche
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