Abstract A51: Cancer stem cell marker ALDH1A3 is associated with increased resistance of Lovo-1 colon cancer cells to autophosphorylation inhibitor of FAK.

Abstract

Abstract Introduction: Focal Adhesion Kinase is an important survival signal in cancer. Recently, we demonstrated that the autophosphorylation inhibitor of FAK, Y15 effectively inhibited colon cancer cell viability in vitro and blocked xenograft tumor growth in vivo. Among several colon cancer cell lines, Lovo-1 was more resistant to Y15 than other cell lines. To reveal signaling pathways responsible for increased resistance of Lovo-1 cells to autophosphorylation inhibitor of FAK, we performed microarray gene profile study in SW620 and Lovo-1 cells treated with Y15 inhibitor. Experimental Procedures: MTT assay and clonogenicity assays were used to detect the effect of Y15 on viability and colony formation. Western blotting was used to detect protein expression. IlluminaRef 8 whole genome gene expression microarray, bioinformatics and statistical analyses were performed to detect significantly affected genes by Y15 in two colon cancer cell lines: SW620 and Lovo-1 cells. PicoProbe ALDH activity assay kit (Biovision) was used to detect ALDH activity. ALDH1A3 siRNA was used to inhibit ALDH1A3 expression. The specific ALDH inhibitor, diethylaminobenzaldehyde was used to inhibit ALDH activity. Results: MTT, clonogenicity assays and Western blotting with Y397-FAK antibody demonstrated that Lovo-1 colon cancer cells were more resistant to Y15 treatment than SW620 colon cancer cells. The microarray analysis detected 12 up- and 4 significantly down-regulated genes that were affected >2-fold (p<0.05) in Lovo-1, but not in SW620 cells. Among unique genes up-regulated by Y15 in Lovo-1 cells, a stem cell marker- ALDH1A3 was detected to be up-regulated >2-fold. ALDH activity kit demonstrated high activity of ALDH in Lovo-1 colonospheres, and the spheres were resistant to Y15 treatment. Treatment with ALDH1A3 siRNAs sensitized Lovo-1 cells to Y15 inhibitor and decreased cell viability more effectively than each agent alone. The same result was obtained with ALDH inhibitor, DEAB in combination with Y15 inhibitor. Conclusion: The microarray study revealed several important genes up- or down-regulated in Lovo-1 cancer cells in response to Y15 inhibitor compared to SW620 cells. The Lovo-1 cells formed spheres with increased ALDH activity. A cancer stem cell marker, ALDH1A3 was associated with increased resistance of Lovo-1 cancer cells to Y15. In addition, down-regulation of ALDH1A3 and FAK effectively decreased viability of Lovo-1 colon cancer cells, resistant to FAK inhibitor that can be used as a novel combination therapy approach in resistant cell lines. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A51. Citation Format: Baotran Ho, Melissa Heffler, Jeffrey Conroy, Liu Song, Dan Wang, William G. Cance, Vita M. Golubovskaya. Cancer stem cell marker ALDH1A3 is associated with increased resistance of Lovo-1 colon cancer cells to autophosphorylation inhibitor of FAK. [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 A51.