Abstract 4881: Anti-cancer efficacy of a novel phenothiazine derivative is independent of dopamine and serotonin receptor inhibition
Authors: Vanneste Marion , Venzke Anita , Guin Soumitra , Krysan Damian , Meyers Marvin , Henry Michael 
Published: April 2023

Abstract
Introduction: An attractive, yet unrealized, goal in cancer therapy is repurposing psychiatric drugs that can readily penetrate the blood-brain barrier for the treatment of deadly primary brain tumors and brain metastasis. The phenothiazine (PTZ) class of anti-psychotic agents were originally developed to inhibit dopamine- and serotonin-receptors (DR/5-HTRs). PTZs have demonstrated anti-cancer properties in numerous studies through a variety of proposed mechanisms, including on-target (DR/5-HTR) and off-target (e.g. calmodulin (CaM) inhibition) effects. PTZs inhibit DR/5-HTRs in the low- to mid-nanomolar range whereas anti-cancer effects tend to be observed only at a higher (low micromolar) dose range. Thus, clinical efficacy of existing PTZs may be limited by toxicity of on-target effects (sedation) of these drugs as DR/5-HTR antagonists. However, it has been unclear whether anti-cancer effects of PTZs are entirely separate from their activity as DR/5-HTR antagonists. Here we evaluate the in vitro anti-cancer efficacy of a novel PTZ analog (CWHM-974, a N-benzyl-linked fluphenazine (FLU) derivative), originally developed to treat brain-resident fungal infections, that is a 100-1000-fold less potent DR/5-HTR antagonist than FLU.
Experimental procedures: To evaluate the anti-cancer efficacy of FLU and CWHM-974, we assessed cytotoxicity following 72h exposure against a broad panel of cancer cell lines. To investigate CaM as a target for CWHM-974, we evaluated the correlation between the sensitivity to CMW-974 and sensitivity to the CaM inhibitor W-7 as well as CaM gene expression levels in a panel of cancer cell lines. We also tested the anti-cancer efficacy of a sulfoxide analog of CWHM-974 (SLU-0010894) which lacks CaM binding activity.
Results: We confirmed the anti-cancer efficacy of FLU and demonstrated that CWHM-974 is consistently 2-10-fold more potent than FLU in a broad panel of cancer cell lines. Among the most sensitve cancer cell lines are melanoma, which is a cancer that frequently metastasizes to the brain. Consistent with the hypothesis that CaM is a relevant target for CWHM-974, we found that: 1) CaM mRNA (CALM1) levels are correlated with sensitivity to CWHM-974; 2) the activities of CWHM-974 and W7 are correlated, and 3) most importantly, SLU-0010894 does not exhibit anti-cancer activity.
Conclusions: The anti-cancer efficacy of the PTZ derivative CWHM-974 is seperable from DR/5-HTR antagonism. Our data suggests that further optimization of this compound may preserve the desirable pharmacologic properties for targeting brain cancers while reducing toxicity due to DR/5-HTR antagonism. Moreover, we provide a variety of evidence suggesting that anti-cancer efficacy of CWHM-974 is associated with CaM binding. Ongoing efforts will characterize the activity of CWHM-974 in mouse models of brain metastasis.
Citation Format: Marion Vanneste, Anita Venzke, Soumitra Guin, Damian Krysan, Marvin Meyers, Michael Henry. Anti-cancer efficacy of a novel phenothiazine derivative is independent of dopamine and serotonin receptor inhibition. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4881.