Development of a novel, targeted small molecule inhibitor of the nucleoside salvage pathway through murine studies to enable further clinical advancement in non-small-cell lung cancer

PROJECT SUMMARY Non-small cell lung cancer (NSCLC) is the most common type of lung cancer, the leading cause of cancer related deaths worldwide. FDA approval of mutant selective KRAS inhibitors (KRASi) has transformed the management of KRAS-G12C mutant NSCLC, which accounts for over 12% of cases. Yet, KRASi have major limitations. Only 30-40% of patients respond to KRASi, and many of those who do respond eventually acquire resistance. New research has identified persistent nucleoside salvage pathway activity in cancer cells as a KRASi escape route that enables their survival by providing the metabolites necessary for DNA replication and repair. Trethera is a clinical stage biopharmaceutical company that has developed a first-in-class small molecule drug, TRE-515, to block the rate limiting nucleoside salvage pathway enzyme deoxycytidine kinase (dCK). Notably, TRE-515 is the only dCK inhibitor to enter clinical development. Trethera is currently evaluating TRE-515 in a Phase 1a open- label, dose escalation study in patients with solid tumors (NCT05055609), which has enrolled 27 patients across 7 separate cohorts (40 to 720 mg). TRE-515 is well tolerated (no dose limiting clinical or laboratory toxicities have been observed in any treated patient) with mild side effects, compelling target engagement, and early signs of clinical benefit. These encouraging results highlight the immediate potential for TRE-515 to improve outcomes when administered as a component of a rationally designed combination therapy. Phase I of the proposed research (Aims 1 and 2) will establish dCK as a targetable KRASi resistance mechanism in NSCLC models. In Aim 1, the improved efficacy of KRASi with TRE-515 will be defined in two human KRAS-G12C mutant NSCLC xenograft models. In Aim 2, xenograft tumors derived from engineered dCK knockout NSCLC cells will be evaluated to confirm that blockade of cancer cell dCK activity enhances KRASi efficacy. Phase II research (Aims 3, 4, and 5) will generate the necessary data to support a clinical trial testing KRASi with TRE-515 for NSCLC. Aim 3 will apply a clinically relevant positron emission tomography (PET) imaging assay to track tumor dCK activity as a noninvasive treatment response biomarker. In NSCLC xenograft models, baseline and treatment induced alterations in tumor dCK specific PET probe uptake will be associated with combination therapy efficacy. Studies in Aim 4 will test the antitumor effects of KRASi with TRE-515 across KRAS-G12C mutant NSCLC models in vitro to determine the mechanism of anticancer activity and if specific co-occurring mutations dictate combination treatment efficacy. Therapy response will be evaluated using mass spectrometry metabolite quantification alongside flow cytometry analysis of cell cycle alterations or cell death. In Aim 5, TRE-515 will be tested with KRASi across an expanded panel of NSCLC xenograft tumor models including a KRAS-G12C mutant patient derived xenograft model and a lung orthotopic tumor model. The data generated through the successful completion of proposed Aims will (i) establish the enhanced efficacy of KRASi with TRE-515, (ii) enable a Phase 1b trial for NSCLC evaluating KRASi with TRE-515, and (iii) attract outside funding for the proposed trial.