Détection des variants de fusion du gène ALK par séquençage massif parallèle ciblé à partir d'ARN et réponse au crizotinib dans les cancers bronchiques non à petites cellules

Abstract : The aim of the study was to test and improve the efficacy of an amplicon-based parallel RNA sequencing (RNA-seq) assay for ALK fusion transcript variants detection in comparison with immunochemistry (IHC) and fluorescent in-situ hybridization (FISH) in a cohort of non-small cell lung cancer (NSCLC) samples. 53 formalin-fixed paraffin-embedded ALK-positive samples by FISH and/or IHC and 23 ALK-negative cases were tested by RNA-seq. Subsequently, a protocol combining RNA-seq and an anchored multiplex PCR was evaluated in 10 samples unsuccessfully analyzed with RNA-seq alone. 75% of the tested samples were successfully analyzed by RNA-seq. RNA-seq sensitivity and specificity were of 80% and 100% respectively compared to IHC and FISH. In IHC- and FISH-positive samples, RNA-seq detected an ALK rearrangement in all cases. RNA-seq showed to be a promising rescue technique, especially for the equivocal and/or borderline-positive IHC/FISH cases (an ALK status could be assigned in 10/10 cases), less for IHC/FISH discordant cases (2/7 cases). Among 10 cases unsuccessfully tested by RNA-seq alone, anchored multiplex PCR allowed the detection of 3 fusion transcripts, one of which was never described (CLIP1(12)-ALK(20)). Median progression free survival was longer in crizotinib-treated patients harboring EML4-ALK variants 1 or 2 compared to variants 3a/b (314 vs 192 days, p = 0.17). RNA-seq appears to be effective for ALK rearrangement diagnostic in NSCLC and to be a promising rescue technique for equivocal and/or borderline cases. It also allows transcript variants identification which could be a pronostic biomarker in these patients.
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Master Thesis
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https://dumas.ccsd.cnrs.fr/dumas-01609830
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Submitted on : Wednesday, October 4, 2017 - 9:38:23 AM
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Julian Pinsolle. Détection des variants de fusion du gène ALK par séquençage massif parallèle ciblé à partir d'ARN et réponse au crizotinib dans les cancers bronchiques non à petites cellules. Médecine humaine et pathologie. 2017. ⟨dumas-01609830⟩

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