Background: Tolfenamic acid (TA) is Non-Steroidal Anti-Inflammatory Drug (NSAID) used to treat migraines. We previously reported that TA exhibits both chemopreventive (esophageal cancer) and chemotherapeutic activities in pancreatic, esophageal and colon cancer models that acts through down-regulating the expression of Sp proteins. Sp proteins regulate the expression of PSA, c-Met, survivin and other critical proteins that are important for cancer cell growth and metastasis. In this preclinical study, we have screened several new TA derivatives to evaluate their anti-cancer activity using prostate cancer model in order to identify robust candidates for the treatment of this and other cancers.

Method: TA derivatives were selected based on their structural relationship with TA. Human prostate cancer cells (PC-3, DU-145 and LNcaP) were grown in the presence of control or various concentrations (25, 50 and 100 µM) of TA or selected analogs and cell proliferation was measured at 24, 48, and 72 h post-treatment using Cell Titer-Glo kit. Further studies were conducted with the two most active TA analogs. PC-3 cells were treated with vehicle or 50 µM TA analogs for 48 h and cell lystaes were prepared and used to evaluate the expression of Sp proteins (Sp1, Sp3 and Sp4), c-Met, Akt and survivin through Western blot analysis. Scratch (wound healing) assay was performed to determine the effect of these drugs on PC-3 cell migration. For in vivo studies, nude mice were injected with PC-3 cells into the right prostate and treated with vehicle or 50 mg/kg of one of the active TA analog/3 times a week and tumor growth was continuously monitored for 4 weeks.

Results: Data showed a significant decrease in cell proliferation that was dose and time dependent. Screening the anti-proliferative activity, we identified at least two TA analogs that inhibited the cell viability more than original TA compound. TA and the two most active analogs significantly diminished the expression of Sp proteins, c-Met, phospho-Akt and survivin and inhibited the migration of cells. Interestingly, the effect of two TA analogs tested was greater than TA as reflected with their higher anti-proliferative activity. In vivo data showed a significant decrease (>50%) in tumor weight and volume after 4 weeks of treatment with selected TA analog confirming a strong correlation between in vitro and in vivo results. These results suggest that TA analogs inhibit prostate cancer cell proliferation, down-regulate the expression of c-Met and survivin, and disrupt phosphorylation of Akt through suppressing Sp proteins.

Conclusion: Targeting key transcription factors such as Sp proteins using small molecules is a new strategy for the treatment of cancer including that of the prostate. In this interesting screening test, we have identified two promising active TA analogs for the potential implications in cancer therapy.Phase I clinical trial is in preparation for the lead compound (TA).

Citation Format: Maen Abdelrahim, Mehmet Asim Bilen, Stephen Safe, Ala Abudayyeh. New tolfenamic acid's derivatives inhibit cancer cells growth and tumor progression: Pre-clinical studies update. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4532. doi:10.1158/1538-7445.AM2014-4532