Targeting O6-methylguanine-DNA Methyltransferase Deficiency in Preclinical Models of Extracranial Human Cancers with a Tumor-Selective DNA-Modifying Agent

Abstract

Temozolomide (TMZ) is an established therapy for gliomas with silencing of the DNA repair gene O6-methylguanine-DNA methyltransferase (MGMT) but is hindered by the frequent development of resistance via loss of mismatch repair (MMR) proteins. To overcome this resistance, a novel 2-fluoroethylating agent, KL-50, was designed to generate toxic DNA interstrand cross-links specifically in MGMT-deficient cells, bypassing MMR. KL-50 has shown efficacy in preclinical MGMT-silenced, TMZ-resistant glioma models. MGMT loss also occurs in a wide range of nonglioma cancers, but the ability of KL-50 to effectively treat these tumors with minimal off-target toxicity remains untested. In this study, we utilize a large-scale cell-line screen, focused panels of cancer cell lines, and tumor xenograft mouse models, including patient-derived xenografts, to interrogate the therapeutic potential of KL-50 in treating intracranial and extracranial tumors. We find that KL-50 is highly efficacious in a wide range of MGMT-deficient human tumor models, including colon cancer, melanoma, and lung cancer in vivo models. In addition, we demonstrate that KL-50 is impervious to loss of MMR and remains effective in tumors with induced TMZ resistance. Finally, we determine that KL-50 possesses higher MGMT selectivity in vitro compared with chloroethylating agents and is associated with less in vivo systemic toxicity in preclinical models. These results establish KL-50 as the first agent to maintain strong selectivity for MGMT loss while inducing tumor toxicity by an MMR-independent mechanism and support further development of 2-fluoroethylating agents for use in a tumor type–agnostic, biomarker-based strategy targeting MGMT-deficient tumors.