Deep RNA Sequencing Revealed Fusion Junctional Heterogeneity May Predict Crizotinib Treatment Efficacy in ALK-Rearranged NSCLC

Abstract

Introduction: Gene fusion variants in ALK-rearranged NSCLC may predict patient outcomes, but previous results have been inconclusive. Fusion isoforms coexisting in the same tumor may affect the efficacy of targeted therapy, but they have not been investigated.Methods: Patients with ALK-rearranged NSCLC who received crizotinib treatments were recruited. Precrizotinib tumor tissues were analyzed by the anchored multiplex polymerase chain reaction for targeted RNA sequencing. Kaplan-Meier and Cox regression were used to compare overall and progression-free survivals.Results: Of the 51 studied subjects, EML4-ALK variant types v1, v2, v3, and others were detected in 23 (45.1%), five (9.8%), 19 (37.3%), and four patients (7.8%), respectively. Multiple EML4-ALK RNA isoforms were detected in 24 tumors (47.1%), and single isoform in 27 (52.9%). Most of the v3 tumors (16 of 19) harbored both v3a and v3b RNA isoforms. Multiple isoforms were also detected in eight non-v3 tumors (33.3% of all 24 multiple isoforms; five v1, two v5 0 , and one v2). Compared with patients with single isoform, those with multiple isoforms had worse progression-free (hazard ratio and 95% confidence interval:

Molecular characterization of genomic breakpoints of ALK rearrangements in non‐small cell lung cancer

“…Further comparative analysis of the transcripts and the amino acid sequences showed that partial retention of exon 20 could ensure the in‐frame sequence with integrity of the ALK kinase domain [ 43 , 44 ]. Patients harbouring multiple EML4‐ALK variants implied a poor prognosis due to the high heterogeneity in the tumour tissue [ 45 ]. Therefore, RNA‐based NGS showed an advantage in detecting fusion patterns in which multiple variants coexist, and more precise splicing results at the transcription level were illustrated.…”

Clinically relevant fusion oncogenes: detection and practical implications

“… 252 Similarly, RNA sequencing revealed that ALK junctional heterogeneity in NSCLC may predict resistance to crizotinib. 253 Likewise scRNA-seq of chemoresistant cervical cancer revealed induction of the (PI3K)/AKT pathway. 254 Thus, it is possible to infer the mechanisms of acquired resistance and to monitor clonal changes of tumors in response to therapy.…”

Biomarker-Targeted Therapies in Non–Small Cell Lung Cancer: Current Status and Perspectives

“…Approximately 50–60% of patients with ALK kinase-domain mutations are resistant to second-generation ALK TKIs [ 49 ]. It may be important to note that ALK fusion variants may play a role in the development of ALK inhibitor resistance on-target [ 50 ]. Several recurrent acquired mutations in ALK confer resistance by decreasing drug binding, including I1171T/N/S, V1180L, G1269A and G1202R [ 51 ].…”