Combination of clotam and vincristine enhances anti-proliferative effect in medulloblastoma cells

• Published in: Gene Vol. 705; pp. 67 - 76
• Main Authors: Patil, Shruti, Sankpal, Umesh T., Hurtado, Myrna, Bowman, W. Paul, Murray, Jeffrey, Borgmann, Kathleen, Ghorpade, Anuja, Sutphin, Robert, Eslin, Don, Basha, Riyaz
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 15-07-2019
• Subjects: Antineoplastic Combined Chemotherapy Protocols - pharmacology; Cell Cycle - drug effects; Cell Line, Tumor; Cell Proliferation - drug effects; Cell Survival - drug effects; Cerebellar Neoplasms - drug therapy; Cerebellar Neoplasms - metabolism; Combination Index; Dose-Response Relationship, Drug; Down-Regulation; Drug Synergism; Gene Expression Regulation, Neoplastic - drug effects; Humans; Inhibitory Concentration 50; Medulloblastoma; Medulloblastoma - drug therapy; Medulloblastoma - metabolism; ortho-Aminobenzoates - pharmacology; Survivin - metabolism; Survivin expression; Tolfenamic acid; Vincristine; Vincristine - pharmacology; VCR; Tolfenamic acid; DMSO; NSAID; Survivin expression; CDK; ATCC; DMEM; SHH; ANOVA; MB; c-PARP; FBS; HSD; EA; CI; Dm; PSN; MEE; COX; TA; Vincristine; EMEM; r; DNA; Medulloblastoma; IAP; SEM; DPBS; Combination Index
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/j.gene.2019.04.037

Medulloblastoma (MB) is characterized by highly invasive embryonal neuro-epithelial tumors that metastasize via cerebrospinal fluid. MB is difficult to treat and the chemotherapy is associated with significant toxicities and potential long-term disabilities. Previously, we showed that small molecule, clotam (tolfenamic acid: TA) inhibited MB cell proliferation and tumor growth in mice by targeting, survivin. Overexpression of survivin is associated with aggressiveness and poor prognosis in several cancers, including MB. The aim of this study was to test combination treatment involving Vincristine® (VCR), a standard chemotherapeutic drug for MB and TA against MB cells. DAOY and D283 MB cells were treated with 10 μg/mL TA or VCR (DAOY: 2 ng/mL; D283: 1 ng/mL) or combination (TA + VCR). These optimized doses were lower than individual IC50 values. The effect of single or combination treatment on cell viability (CellTiterGlo kit), Combination Index (Chou-Talalay method based on median-drug effect analysis), activation of apoptosis and cell cycle modulation (by flow cytometry using Annexin V and propidium iodide respectively) and the expression of associated markers including survivin (Western immunoblot) were determined. Combination Index showed moderate synergistic cytotoxic effect in both cells. When compared to individual agents, the combination of TA and VCR increased MB cell growth inhibition, induced apoptosis and caused cell cycle (G2/M phase) arrest. Survivin expression was also decreased by the combination treatment. TA is effective for inducing the anti-proliferative response of VCR in MB cells. MB has four distinct genetic/molecular subgroups. Experiments were conducted with MB cells representing two subgroups (DAOY: SHH group; D283: group 4/3). TA-induced inhibition of survivin expression potentially destabilizes mitotic microtubule assembly, sensitizing MB cells and enhancing the efficacy of VCR.
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•Combination of clotam and vincristine induces anti-cancer activity in medulloblastoma cells.•Clotam and vincristine combination indiscriminately inhibiting SHH and 4/3 sub group cell lines•Targeting survivin with clotam is effective for sensitizing medulloblastoma cells to chemotherapy.•Clotam potentially destabilizing the microtubules for inducing the growth arrest caused by vincristine