Repurposing the serotonin agonist Tegaserod as an anticancer agent in melanoma: molecular mechanisms and clinical implications

Abstract

Background: New therapies are urgently needed in melanoma particularly in late-stage patients not responsive to immunotherapies and kinase inhibitors.Methods: Drug screening, IC50 determinations as well as synergy assays were detected by the MTT assay. Apoptosis using Annexin V and 7AAD staining was assessed using flow cytometry. TUNEL staining was performed using immunocytochemistry. Changes in phosphorylation of key molecules in PI3K/Akt/mTOR and other relevant pathways were detected by western blot as well as immunocytochemistry. To assess in vivo anti-tumor activity of Tegaserod, syngeneic intravenous and subcutaneous melanoma xenografts were used. Immunocytochemical staining was performed to detect expression of active Caspase-3, cleaved Caspase 8 and p-S6 in tumors. Evaluation of immune infiltrates was carried out by flow cytometry.Results: Using a screen of 770 pharmacologically active and/or FDA approved drugs, we identified Tegaserod (Zelnorm, Zelmac) as a compound with novel anti-cancer activity which induced apoptosis in murine and human malignant melanoma cell lines. Tegaserod (TM) is a serotonin receptor 4 agonist (HTR4) used in the treatment of irritable bowel syndrome (IBS). TM's anti-melanoma apoptosis-inducing effects were uncoupled from serotonin signaling and attributed to PI3K/Akt/mTOR signaling inhibition. Specifically, TM blunted S6 phosphorylation in both BRAF V600E and BRAF wildtype (WT) melanoma cell lines. TM decreased tumor growth and metastases as well as increased survival in an in vivo syngeneic immune-competent model. In vivo, TM also caused tumor cell apoptosis, blunted PI3K/Akt/mTOR signaling and decreased S6 phosphorylation. Furthermore TM decreased the infiltration of immune suppressive regulatory CD4 + CD25 + T cells and FOXP3 and ROR-γt positive CD4 + T cells. Importantly, TM synergized with Vemurafenib, the standard of care drug used in patients with late stage disease harboring the BRAF V600E mutation and could be additively or synergistically combined with Cobimetinib in both BRAF V600E and BRAF WT melanoma cell lines in inducing anti-cancer effects.

Mechanisms of Acquired BRAF Inhibitor Resistance in Melanoma: A Systematic Review

“…A considerable number of preclinical studies are investigating other novel targets for overcoming BRAF inhibitor resistance. These include combining BRAF and/or MEK inhibitors with inhibitors of pre-mRNA splicing (to counteract resistance caused by BRAF splicing) [ 251 ], BH3-mimetics [ 252 , 253 ], BCL2 inhibitors [ 254 ], mitochondrial-targeted agents [ 255 , 256 ], inhibitors of p90 ribosomal S6 kinases [ 257 , 258 ], pro-caspase activating compounds [ 259 ], Rho kinase 1 (ROCK1) inhibitors [ 260 ], protein kinase Cδ inhibitors [ 261 ], tubulin inhibitors [ 262 ], ErbB2 or ErbB3 inhibitors [ 222 , 263 , 264 ], activators of the liver-X nuclear hormone receptor [ 265 ], an antibody conjugate targeting the endothelin B receptor [ 266 ], monoclonal antibodies against chondroitin sulfate proteoglycan 4 [ 267 ], inhibitors of sterol regulator element binding protein I (SREBP-1) [ 268 ], copper chelators [ 269 ], polo-like 3 kinase inhibitors (including in models of BRAF + MEK inhibitor resistance) [ 270 , 271 ], anti-nodal antibodies [ 272 ], PAK1 inhibitors [ 273 ], GLI1/2 inhibitors [ 274 ], inhibitors of IQ motif-containing GTPase activating protein 1 (IQGAP1) [ 275 ], serotonin agonists [ 276 ], CK2 inhibitors [ 277 ], p53 activators [ 278 ], metformin [ 279 ], statins [ 280 ], non-steroidal anti-inflammatory drugs [ 281 ], mibefradil [ 282 ], hydroxychloroquine (an autophagy inhibitor) [ 83 ], and A100 (a reactive oxygen species-activated prodrug) [ 283 ].…”

Role of serotonin receptor signaling in cancer cells and anti-tumor immunity

“…Tegaserod also reduces the tumor volume in the mouse melanoma model. However, tegaserod mediates this action through a serotonin-independent signaling mechanism, where it blunts the phosphorylation of S6 through inhibition of the PI3/Akt/mTOR pathway 122 .…”

Tegaserod Maleate Inhibits Esophageal Squamous Cell Carcinoma Proliferation by Suppressing the Peroxisome Pathway

“…Studies have shown that 5-hydroxytryptamine plays a mitogenic role in colon cancer cells, and 5-hydroxytryptamine 4-receptor is significantly expressed in both colon cancer tissue and cells ( 26 ). Tegaserod maleate has been shown to exert anticancer activity by inducing apoptosis in melanoma cells ( 27 ), and it has been identified as a JAK/STAT3 signal inhibitor, impeding the growth of a variety of cancer cells, including lung cancer, prostate cancer, colon cancer, and cervical cancer ( 28 ). JAK/STAT3 pathway also plays an important role in cell proliferation.…”