Treatment Efficacy and Resistance Mechanisms Using the Second-Generation ALK Inhibitor AP26113 in Human NPM-ALK–Positive Anaplastic Large Cell Lymphoma

Abstract

ALK is a tyrosine kinase receptor involved in a broad range of solid and hematologic tumors. Among 70% to 80% of ALK þ 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 þ 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 IC 50 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 F1174VþL1198F and L1122VþL1196M 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.Implications: 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. Mol Cancer Res; 13(4); 775-83. Ó2014 AACR.

The Potent ALK Inhibitor Brigatinib (AP26113) Overcomes Mechanisms of Resistance to First- and Second-Generation ALK Inhibitors in Preclinical Models

“…Thus, while we, and others (28,29), have identified mutants that brigatinib inhibits less potently than native ALK, we predict that when levels of exposure achieved in patients are taken into account, brigatinib has the potential to have pan-ALK inhibitory activity such that no single ALK mutant will be associated with a high degree of resistance, especially at the higher 180-mg dose level.…”

NPM/ALK mutants resistant to ASP3026 display variable sensitivity to alternative ALK inhibitors but succumb to the novel compound PF-06463922

“…Real-time quantitative PCR confirmed the data in SUPM2R cells, but not in K299R cells, where N/A mRNA was only slightly increased (2 to 4-fold) in two out of three cell lines (Table 1). For a comparison, in other ALKi-resistant K299 cells carrying wild-type ALK sequence, we observed 16 to 25-fold increases in N/A mRNA expression and this was a clear effect of oncogene amplification [13]. Therefore, in this case, we cannot definitely ascribe resistance to oncogene overexpression.…”

“…However, we can expect resistance to arise under any treatment. In order to predict the possible evolution of ALK+ ALCL under ALKi pressure, and to study cross-resistance among the available drugs, we recently set up an in vitro model of acquired resistance to selective ALK inhibition in ALCL [12, 13]. As part of this effort, we isolated six NPM/ALK+ cell lines that show high resistance to the novel ALKi, ASP3026 (Astellas Pharma, Japan).…”

Brigatinib, an anaplastic lymphoma kinase inhibitor, abrogates activity and growth in ALK positive neuroblastoma cells, Drosophila and mice

“…Brigatinib has been shown to inhibit ALK activity in NSCLC cell lines carrying the EML4-ALK fusion protein [6,30]. In order to investigate the therapeutic efficacy of brigatinib in a neuroblastoma setting we employed several neuroblastoma cell lines, including CLB-BAR (MYCN amplification, ALK (Δ4-11) and amplified, ALK addicted), CLB-GE (MYCN amplification, ALK (F1174V) amplification, ALK addicted), IMR32 (MYCN amplification, WT ALK) and CLB-PE (MYCN amplified, WT ALK) [21,[31][32][33].…”