Abstract A141: Dasatinib augments the antitumor effect of PLX4720 in a novel immunocompetent mouse model of BRAFV600E-positive anaplastic thyroid cancer.

Abstract

Abstract Background: The MAPK and Src signaling pathways have emerged as critical drivers and mediators of papillary and anaplastic (PTC and ATC) thyroid cancer progression and the individual inhibition of either pathway has marked anti-tumor effects in mouse models. Unfortunately, neither drug induces apoptosis and drug resistance may develop when either is used. We postulated that the inhibition of the Src signaling pathway would further sensitize BRAFV600E-positive thyroid cancer cells to PLX4720 treatment. Methods: Proliferation and apoptosis were analyzed in four novel murine PTC and ATC cell lines derived from genetically engineered mice. Cells were treated with the BRAFV600E-inhibitor, PLX4720 (10μM), or the Src kinase inhibitor, dasatinib (50 nM), or a combination of the two drugs. Single and combinatorial dose response was determined and analyzed using Chou-Talalay. To determine the effect of combinatorial treatment in vivo, 105 murine ATC cells (TBP-3743 - BRAFV600E/WT; p53−/−) were orthotopically implanted in syngeneic immunocompetent mice. Mice were randomized into four groups and placed on treatment regimens 3 days following implantation, consisting of either control chow, PLX4720 chow (ab libitium), dasatinib gavage (50mg/kg) or a combination of PLX4720 chow and dasatinib gavage. Results: Basal pERK, pSrc(Y416) and pFAK(Y861) indicated active MAPK and Src signaling pathways in the thyroid cancer cell lines. PLX4720 treatment reduced pERK in the ATC lines, while dasatinib alone reduced both pSrc(Y416) and pFAK(Y861). The combination index (CI) of PLX4720 and dasatinib was calculated to be <1.0 for each line (TBP-3868 = 0.52, TBP-3743 = 0.45; TBPt-3403 = 0.56; TBPt-3610R = 0.07) indicating a synergistic effect on cellular proliferation. While neither PLX4720 nor dasatinib treatment induced significant apoptosis beyond baseline, combinatorial treatment induced apoptosis in the ATC lines TBP-3743 and TBPt-3403 as determined by caspase 3 and PARP cleavage in vitro. Orthotopic implantation of TBP-3743 resulted in the rapid formation of anaplastic tumors with CD3+, CD8+, B220+ and CD163+ immune cell infiltration. Relative to controls, which developed tumors two weeks post-implantation (231 mm3), treatment with PLX4720, dasatinib or a combination significantly reduced tumor volumes to 92 mm3, 123 mm3 and 62 mm3, respectively. Three weeks post-implantation, mean tumor volume significantly increased in mice treated with PLX4720 alone to 294 mm3 but not in mice treated with both PLX4720 and dasatinib which exhibited a modest increase to 118 mm3. Increased cleaved caspase 3 was detected in tumor sections from combination treated mice. Ultimately, the combination of PLX4720 and dasatinib resulted in increased apoptosis and reduced tumor volume relative to mice treated with PLX4720 alone. Conclusion: Combinatorial treatment of PTC and ATC cells with PLX4720 and dasatinib synergistically reduces proliferation and induces apoptosis in vitro and results in apoptosis and reduced tumor size relative to PLX4720 treatment alone in a pre-clinical mouse model of ATC. Together, these findings suggest that Src-inhibition enhances the anti-tumor effect of BRAFV600E-inhibition and the combination of these two FDA-approved drugs may have potential as a treatment paradigm for patients with ATC or refractory PTC. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A141. Citation Format: Pierre Vanden Borre, Viswanath Gunda, David McFadden, Peter Sadow, Sareh Parangi. Dasatinib augments the antitumor effect of PLX4720 in a novel immunocompetent mouse model of BRAFV600E-positive anaplastic thyroid cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A141.