Abstract C165: Zurletrectinib, a next-generation TRK inhibitor shows strong intracranial activity against NTRK fusion-positive tumors with on-target resistance to first-generation drugs
Authors: Roa Paola , Foglizzo Valentina , Harada Guilherme , Repetto Matteo , Kulick Amanda , de Stanchina Elisa , Auwardt Supipi , de Marchena Michelle , Ahmed Shaza , Bremer Nicole , Yang Soo-Ryum , Drilon Alexander , Cocco Emiliano 
Published: December 2023

Abstract
NTRK fusions are oncogenic drivers in multiple tumor types. While treatment with TRK inhibitors is effective in ~80% of patients with NTRK fusion-positive tumors, resistance eventually occurs. The most common mechanism of resistance is the emergence of mutations in the kinase domain of the fusions that impair drug binding. Next-generation TRK inhibitors active against these mutants are in clinical development (selitrectinib) or have received breakthrough therapy designation (repotrectinib). Here, we describe the activity of zurletrectinib, a novel next-generation TRK inhibitor. Cell viability assays, in silico modeling, and western blot analyses were used to test the efficacy of zurletrectinib against NTRK fusion-positive models and the most common TRKA resistant mutations. Finally, the in vivo activity of zurletrectinib was assessed in glioma orthotopic models. Selitrectinib, zurletrectinib, and repotrectinib inhibited in vitro cell growth and TRK-mediated signaling of multiple NTRK fusion-positive cell lines with IC50s in the low nanomolar range. Next, we used in silico modeling to predict the binding of zurletrectinib to TRK resistant mutations. These include the TRKA G595R solvent front and the TRKA G667C xDFG substitutions, and the TRKA G595R/G667A or the TRKA G595R/G667C double mutations. Selitrectinib, repotrectinib and zurletrectinib were all predicted to bind the TRKA G595R, and the TRKA G595R/G667A mutants, but not the bulkier TRKA G667C or the TRKA G595R/G667C mutants. To confirm our results, we tested the activity of zurletrectinib against multiple primary human NTRK fusion-positive, TRKA-mutant cell lines. All next-generation inhibitors were active against models expressing the TRKA G595R or the TRKA G595R/G667A double mutation, but not against TRKA G595R/G667C mutant models. Similar results were obtained when the same drugs were tested against mouse NTRK fusion-positive glioma clones engineered by CRISPR to express the mouse Trka ortholog mutations. To determine the in vivo activity of zurletrectinib, mouse glioma orthotopic xenografts harboring the Trka G598R or the Trka G598R/G670A mutations were used. While no significant difference in the survival between selitrectinib, repotrectinib and zurletrectinib was observed in the Trka G598R model, zurletrectinib significantly improved survival of mice harboring the Trka G598R/G670A mutation (median survival=41.5, 66.5, and 104 days for selitrectinib, repotrectinib, and zurletrectinib respectively; P(selitrectinib/zurletrectinib=.0022; P(repotrectinib/zurletrectnib=.038). Together, our data identifies zurletrectinib as a novel, highly potent and selective next-generation TRK inhibitor with strong intracranial activity and a potentially more favorable safety profile than other similar inhibitors.
Citation Format: Paola Roa, Valentina Foglizzo, Guilherme Harada, Matteo Repetto, Amanda Kulick, Elisa de Stanchina, Supipi Auwardt, Michelle de Marchena, Shaza Ahmed, Nicole Bremer, Soo-Ryum Yang, Alexander Drilon, Emiliano Cocco. Zurletrectinib, a next-generation TRK inhibitor shows strong intracranial activity against NTRK fusion-positive tumors with on-target resistance to first-generation drugs [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C165.