Chemoproteomic Discovery of a Ritanserin-Targeted Kinase Network Mediating Apoptotic Cell Death of Lung Tumor Cells

Abstract

Ritanserin was tested in the clinic as a serotonin receptor inverse agonist but recently emerged as a novel kinase inhibitor with potential applications in cancer. Here, we discovered that ritanserin induced apoptotic cell death of non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) cells via a serotonin-independent mechanism. We used quantitative chemical proteomics to reveal a ritanserin-dependent kinase network that includes key mediators of lipid [diacylglycerol kinase , phosphatidylinositol 4-kinase] and protein [feline encephalitis virus-related kinase, rapidly accelerated fibrosarcoma (RAF)] signaling, metabolism [eukaryotic elongation factor 2 kinase, eukaryotic translation initiation factor 2- kinase 4], and DNA damage response [tousled-like kinase 2] to broadly kill lung tumor cell types. Whereas ritanserin exhibited polypharmacology in NSCLC proteomes, this compound showed unexpected specificity for c-RAF in the SCLC subtype, with negligible activity against other kinases mediating mitogen-activated protein kinase signaling. Here we show that ritanserin blocks c-RAF but not B-RAF activation of established oncogenic signaling pathways in live cells, providing evidence in support of c-RAF as a key target mediating its anticancer activity. Given the role of c-RAF activation in RAS-mutated cancers resistant to clinical B-RAF inhibitors, our findings may have implications in overcoming resistance mechanisms associated with c-RAF biology. The unique target landscape combined with acceptable safety profiles in humans provides new opportunities for repositioning ritanserin in cancer.

Chemoproteomic profiling of kinases in live cells using electrophilic sulfonyl triazole probes

“… 32 Further investigations into ritanserin activity revealed secondary activity against DGKα 33 and other kinases. 34 RF001 showed improved specificity for DGKα compared with ritanserin as determined by activity-based profiling studies using ATP acyl phosphate probes. 35 The proteome-wide activity of RF001, however, is currently unknown and is important for evaluating whether RF001 is a suitable fragment for developing kinase probes and inhibitors.…”

“…The lipid kinase tree was generated in-house as previously described. 34 All data shown are representative of 3 experiments (n ¼ 3 biologically independent experiments).…”

Raf‐ERK1/2 signalling pathways mediate steroid hormone synthesis in bovine ovarian granulosa cells

“…Therefore, we focused on the roles of B-Raf or C-Raf in steroid hormones, but not A-Raf. The Raf-ERK1/2 signalling pathways are originally reported to mediate various cellular responses, such as cell proliferation (Zhang, Shu, Li, Li, & Li, 2015), migration (Wu et al, 2013), differentiation (Ji & Andrisani, 2005) and death (Campbell et al, 2018), whereas, emerging evidence also suggests that Raf-ERK1/2 may play essential roles in ovarian physiological functions. FSH (follicle-stimulating hormone) induces phosphorylation of ERK1/2, thereby controlling granulosa cell differentiation and the expression of genes characteristic of the luteal cell phenotype (Wayne, Fan, Cheng, & Richards, 2007).…”

Ritanserin blocks CaV1.2 channels in rat artery smooth muscles: electrophysiological, functional, and computational studies

“…Recently, evidence that ritanserin inhibits several lipids (e.g., diacylglycerol kinase-α, DGKα, a novel, potential therapeutic target in various cancers as well as in immunotherapy) [ 7 – 9 ] and protein kinases (e.g., the feline encephalitis virus-related kinase FER) [ 10 ], as well as the rapidly accelerated fibrosarcoma kinase c-RAF [ 11 ] has been provided. The capacity of perturbing cellular signaling pathways important for cell survival and proliferation, through serotonin-independent mechanisms suggests that ritanserin may be a viable option for in vivo translation and a novel therapeutic tool with potential applications in various tumors.…”