Abstract 2433: Targeting tGLI1 pharmacologically as a new therapeutic strategy for breast cancer brain metastases

Abstract

Abstract Breast cancer is the second leading cause of brain metastases in women; patients with breast cancer brain metastasis (BCBM) survive only 6-18 months after diagnosis. Novel agents that act on targetable driver mutations (EGFR, PI3K, etc.) and penetrate the blood-brain barrier (BBB) have shown promise as systemic options for patients with distant lung and melanoma metastases. Nevertheless, brain metastases represent an area of unmet clinical need for which new pathways, novel mechanisms, and innovative therapies are needed. Cancer stem cells are thought to be a driving force behind not only distant metastasis, but also late-stage recurrence. The hedgehog pathway is an important mediator of breast cancer stem cells (BCSC); however, FDA-approved therapies targeting this pathway have demonstrated limited efficacy in breast cancer clinical trials. Despite advances made in understanding BCSC, it remains challenging to effectively target BCSC underscoring the need to identify and inhibit novel mediators of BCSC to treat BCBM. Our laboratory recently reported that truncated glioma-associated oncogene homolog 1 (tGLI1) promotes preferential breast cancer metastasis to the brain by activating BCSC and astrocytes in the tumor microenvironment (Oncogene 39:64-78, 2020). tGLI1 is an alternatively spliced GLI1 variant that functions as a tumor-specific gain-of-function transcription factor and terminal effector of the hedgehog pathway. tGLI1 knockdown abrogated BCBM, providing the rationale to therapeutically target tGLI1. In this study, we conducted cell-based chemical screens followed by validations and found that ketoconazole (KCZ), an FDA-approved antifungal, selectively targets tGLI1-expressing cancer cells, leading to suppression of BCSC in vitro and BCBM in vivo. Modification of KCZ moieties produced derivatives that retained tGLI1-selectivity against both BCSC in vitro and BCBM in vivo with increased BBB penetrance. Mechanistic studies suggest that KCZ-dependent cell kill is mediated through reducing tGLI1’s ability to bind and transactivate its target gene promoters, reducing expression of target genes Nanog, OCT4, and VEGFA. Based on these data, we opened an early Phase I clinical trial to determine if KCZ penetrates the BBB and modulates tGLI1 signaling in BCBM (NCT03796273). Preliminary analysis confirmed KCZ accumulation in collected brain biospecimens. We further investigated structure-activity relationship by synthesizing and testing 22 novel compounds, and identified two tGLI1-selective compounds that no longer inhibit CYP3A4. Since KCZ-mediated CYP3A4 antagonism can result in adrenal insufficiency and drug-drug interactions, these two novel tGLI1 inhibitors could be promising agents superior to KCZ. Collectively, these data establish tGLI1 as an actionable target for BCBM and validate KCZ and its novel derivatives as promising treatment options for patients with BCBM. Citation Format: Daniel Doheny, Sara Manore, Grace L. Wong, Dongqin Zhu, Sherona Sirkisoon, Marlyn Anguelov, Noah R. Augayo, Angelina T. Regua, Anderson Cox, Cristina Furdui, Terrence Smalley, Alexandra Thomas, Adrianna Masters, Roy E. Strowd, Hui-Wen Lo. Targeting tGLI1 pharmacologically as a new therapeutic strategy for breast cancer brain metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2433.