Mechanisms of Sunitinib Resistance in Gastrointestinal Stromal Tumors Harboring KITAY502-3ins Mutation: An In vitro Mutagenesis Screen for Drug Resistance

Abstract

Purpose: Although tyrosine kinase inhibitors have improved survival in advanced gastrointestinal stromal tumor (GIST), complete response is rare and most patients eventually fail the first-line treatment with imatinib. Sunitinib malate is the only approved second-line therapy for patients with imatinib-resistant or imatinib-intolerant GIST. The clinical benefit of sunitinib is genotype-dependent in regards to both primary and secondary mutations, with GIST patients harboring the KIT AY502-3ins exon 9 mutation being the most sensitive. Experimental Design: As sunitinib resistance is now emerging, our goal was to investigate mechanisms of progression and to test the efficacy of novel tyrosine kinase inhibitor on these resistant mutants in vitro. N-ethyl-N-nitrosourea mutagenesis of Ba/F3 cells expressing the KIT AY502-3ins mutant was used to investigate novel patterns of resistant mutations evolving in the presence of sunitinib. Results: Tumors from patients who developed sunitinib resistance after at least 1 year of radiographic response were analyzed, showing similar findings of a primary KIT AY502-3ins mutation and a secondary mutation in the KIT activation loop. Ba/F3 cells expressing these sunitinib-resistant double mutants showed sensitivity to both dasatinib and nilotinib. Conclusions: Sunitinib resistance in GIST shares similar pathogenetic mechanisms identified in imatinib failure, with acquisition of secondary mutations in the activation domain after an extended initial response to the drug. Moreover, in vitro mutagenesis with or without N-ethyl-N-nitrosourea of Ba/F3 cells expressing KIT AY502-3ins showed acquisition of secondary mutations restricted to the second kinase domain of KIT. In contrast, in vitro resistance to imatinib produces a broader spectrum of secondary mutations including mutations in both KIT kinase domains. (Clin Cancer Res 2009;15(22):6862-70) Although imatinib achieves a partial response or stable disease in most gastrointestinal stromal tumor (GIST) patients, with lasting responses over 5-year period in >20% of patients, complete responses are rare (1). Clinical responses to imatinib in GIST depend on the presence and type of KIT or plateletderived growth factor receptor-α (PDGFRA) gain-of-function mutations. Thus, patients with exon 11 mutations show a partial response rate of 84%, whereas patients with tumors harboring a KIT exon 9 or no detectable mutation had a partial response rate of 48% and 0%, respectively (2). It is now clear that a majority of patients who initially benefit from imatinib eventually become resistant. The most common mechanism of acquired resistance is through a secondary KIT mutation, usually located in either the NH 2 -terminal or the COOH-terminal

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“…For instance, imatinib-resistant CML patients who have progressed on second-line dasatinib treatment harbor mutations that are distinct from those patients having relapsed following second-line treatment with nilotinib (64). This phenomenon has also been documented in KIT-driven GISTs, where primary and secondary KIT mutations have been observed upon relapse to either first-or second-line treatments, respectively (65,66). In this regard, in vitro mutagenesis or in vivo resistance screens combined with structural analysis and genomic characterization of resistant cells can delineate the mechanisms that tumors utilize to evade pharmacological interventions.…”

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“…Mutations that alter the conformational state of ABL are predicted to disfavor imatinib binding and, consequently, its efficacy. Secondary mutations in KIT that occur in the kinase activation loop (which likely promote the active conformation) are associated with resistance to the KIT inhibitors imatinib and sunitinib in vitro and a worse clinical outcome in patients treated with sunitinib (37)(38)(39). In contrast, mutations in the drug/ATP binding pocket of KIT, although common in patients with gastrointestinal stromal cell tumors (GIST) who develop imatinib resistance, are less often detected from in vitro studies or from patients with GISTs that develop sunitinib resistance (37)(38)(39).…”

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“…This offers a survival benefit to the cells effected and is propagated throughout the tumour. This mechanism of acquired resistance has been observed in gastrointestinal tumours treated with sunitinib, but only after approximately a year of response (32). It seems unlikely then that 9 days would be sufficient time for tumours to acquire such mutations.…”