Abstract
ALK is a tyrosine kinase receptor involved in a broad range of solid and hematologic tumors. Among 70% to 80% of ALK(+) anaplastic large cell lymphomas (ALCL) are caused by the aberrant oncogenic fusion protein NPM-ALK. Crizotinib was the first clinically relevant ALK inhibitor, now approved for the treatment of late-stage and metastatic cases of lung cancer. However, patients frequently develop drug resistance to Crizotinib, mainly due to the appearance of point mutations located in the ALK kinase domain. Fortunately, other inhibitors are available and in clinical trial, suggesting the potential for second-line therapies to overcome Crizotinib resistance. This study focuses on the ongoing phase I/II trial small-molecule tyrosine kinase inhibitor (TKI) AP26113, by Ariad Pharmaceuticals, which targets both ALK and EGFR. Two NPM-ALK(+) human cell lines, KARPAS-299 and SUP-M2, were grown in the presence of increasing concentrations of AP26113, and eight lines were selected that demonstrated resistance. All lines show IC50 values higher (130 to 1,000-fold) than the parental line. Mechanistically, KARPAS-299 populations resistant to AP26113 show NPM-ALK overexpression, whereas SUP-M2-resistant cells harbor several point mutations spanning the entire ALK kinase domain. In particular, amino acid substitutions: L1196M, S1206C, the double F1174V+L1198F and L1122V+L1196M mutations were identified. The knowledge of the possible appearance of new clinically relevant mechanisms of drug resistance is a useful tool for the management of new TKI-resistant cases. This work defines reliable ALCL model systems of AP26113 resistance and provides a valuable tool in the management of all cases of relapse upon NPM-ALK-targeted therapy.
Highlights
The pharmacologic inhibition of the Anaplastic lymphoma kinase (ALK) tyrosine kinase has become in the last years an issue of great interest, given its involvement in different malignancies and the recent development of several ALK inhibitors
For KARPAS-299–derived cell lines, we observed oncogene overexpression as the main resistance mechanism, whereas in SUPM2–derived cell lines, we identified several point mutations located within the NPM-ALK kinase domain, which could explain drug resistance
Because the IC50 value observed for all resistant cell lines is extremely high, we explored the presence of low-frequency point mutations in ALK kinase domain as an additional resistance mechanism by deep sequencing
Summary
The pharmacologic inhibition of the ALK (anaplastic lymphoma kinase) tyrosine kinase has become in the last years an issue of great interest, given its involvement in different malignancies and the recent development of several ALK inhibitors. To find a way to overcome AP26113 resistance, we explored the cross-resistance of our KARPAS-299–derived cell lines and mutated Ba/F3 NPMALK against other clinically relevant ALK inhibitors nowadays available, namely Crizotinib, ceritinib, alectinib, ASP3026. Our data predict in a human cell-based model the appearance of different mechanisms of resistance to AP26113, and we explored different ways to overcome resistance using a set of clinically relevant ALK inhibitors. This kind of knowledge is a powerful tool to manage clinical cases of Crizotinib and AP26113 relapse
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