Chlorpromazine activates cGAS-STING signaling and reprograms the immune response in glioblastoma

Glioblastoma (GBM), the most common and aggressive primary brain tumor in adults, poses a formidable therapeutic challenge, due to its intrinsic radio- and chemoresistance and its ability to create a hostile, immunosuppressive tumor microenvironment (TME). Drug repurposing has emerged as a promising strategy to fight GBM. In this context, our efforts focused on chlorpromazine (CPZ), a first-generation antipsychotic agent previously shown by us to exert anti-tumor effects in both preclinical and clinical settings.

We investigated the role of CPZ in remodeling the GBM microenvironment and shaping immune responses using four GBM cell lines, two standard anchorage-dependent models and two patient-derived neurospheres, enriched for tumoral stem cells. We determined cGAS-STING pathway activation and downstream gene expression via flow cytometry and RT-PCR. The cellular secretome following drug treatment was profiled via a luminescence cytokinome assay using a panel of 27 chemokines. Macrophages were phenotyped by flow cytometry using M1 and/or M2 specific markers and, finally, PD-L1 expression was assessed by quantitative flow cytometry and immunoblot analysis.

We demonstrate that CPZ, alone or in combination with temozolomide (TMZ), the current standard of care, activates the cGAS-STING signaling pathway, thus promoting anti-tumor immune responses. Importantly, CPZ counteracts the immunosuppressive effects of TMZ, hindering some TMZ-induced processes as: i) induction of tumorigenic cytokines; ii) macrophage polarization toward a tumor-supportive M2-like phenotype, and iii) increase of PD-L1 expression, a key mechanism of immune evasion.

This study uncovers that CPZ exerts a previously unrecognized anti-cancer immunomodulatory activity, remodeling the immune microenvironment and enhancing the anti-tumor immune response. By overcoming TMZ resistance, CPZ not only exerts a direct anti-neoplastic effect, but also sensitizes GBM cells to standard therapy.