In vitro profiling of IDRX-42 against secondary and tertiary mutations (AP/AL) found in TKI-resistant GIST.

Abstract

11524 Background: The majority of Gastrointestinal Stromal Tumors (GIST) harbor a KIT mutation and respond to imatinib (IM), but over time clinical resistance emerges due to secondary mutations. These secondary mutations are restricted to two regions of the KIT protein: the drug/ATP binding pocket (AP, exons 13 and 14) and the activation loop (AL, exons 17 and 18). Although additional TKI drugs has been approved for later lines of therapy, these agents lack activity against the entire spectrum of known secondary resistance mutations. For example, sunitinib is potent against AP mutations (V654A, T670I), but has minimal activity against AL mutations. Recently, IDRX-42 has demonstrated promising activity in a phase 1/1b study of patients with TKI-resistant GIST. However, the cellular potency of IDRX-42 against various primary, secondary, and compound AP/AL KIT mutations has not been comprehensively described. Methods: We used a panel of GIST cell lines with various mutations that were derived from the parental GIST T1 cell line (primary KIT exon 11 (K11) deletion mutation). We also transiently expressed additional mutations in CHO cells. Densitometry and curve fitting was used to determine the biochemical IC50 for inhibition of KIT autophosphorylation, an accepted surrogate for KIT kinase activity. We also profiled IM and ripretinib (RIP) in the same cell line models. Results: IM was active against primary K11 mutations but was much less active against K11 + AP, K11 + AL, KIT exon 9 (K9), K9 + AP, and K9 + AL mutations. In addition, IM was inactive against K9 or K11 + AP/AL mutations (with all three mutations in cis). RIP potently inhibited K11, K11 + AL mutations, and K9 + AL mutations. However, RIP was less active against K11 + AP, K9 +AP, and K9 or K11 + AP/AL mutations. IDRX-42 was active against K11, K11 + V654A, K11 + AL, K9, K9 + V654A, and K9 +AL mutations. However, IDRX-42 lacked potency against the gatekeeper T670I (AP) mutation and K9 or K11 + AP/AL mutations. We developed resistant cell lines using GIST T1 cell line subjected to chemical mutagenesis and long-term drug selection with IDRX-42. All IDRX-42 resistant clones had an acquired T670I mutation, but no other secondary mutations were identified. Conclusions: IDRX-42 has superior biochemical potency against a panel of primary and secondary KIT mutations compared with IM or RIP. Additionally, IDRX-42 has activity against K9 or K11 + V654A, and K9 or K11 + AL; However, K9 or K11 with T670I mutant kinases were resistant. Notably, none of these three profiled drugs had activity against K9 or K11 + AP/AL mutations. The optimal clinical use of IDRX-42 may be in earlier lines of therapy, prior to the emergence of AP/AL mutations. Biochemical IC50 for inhibition of KIT autophosphorylation. Cells Mutation IDRX-42 (nM) IM (nM) RIP (nM) GIST T1 K11 S S S GIST T1 K11 + V654A S R R GIST T1 K11 + T670I R R R GIST T1 K11 + AL S R S CHO K9 S R S CHO K9 + V654A S R R CHO K9 + T670I R R R CHO K9 + AL S R S CHO (K9 or K11) + AP/AL R R R