J. M. Aye1, S. Mruthyunjayappa2, L. S. Stafman2, E. F. Garner2, J. E. Stewart2, E. Mroczek-Musulman3, K. J. Yoon4, S. L. Cramer1, E. A. Beierle2 1University Of Alabama at Birmingham,Department Of Pediatrics, Division Of Hematology Oncology,Birmingham, Alabama, USA 2University Of Alabama at Birmingham,Department Of Surgery,Birmingham, Alabama, USA 3University Of Alabama at Birmingham,Department Of Pathology,Birmingham, Alabama, USA 4University Of Alabama at Birmingham,Department Of Pharmacology And Toxicology,Birmingham, Alabama, USA
Introduction: Wilms tumor is the most common pediatric renal tumor. The 4-year-relapse-free survival (RFS) rate for patients with non-metastatic, favorable histology Wilms tumor is greater than 90%; however, the 4-year-RFS rate for those with metastatic Wilms tumor is only 70%. A concentrated research effort is needed for these patients. Limited cell lines are available for the study of metastatic Wilms tumor and long-term passaged cell lines do not always accurately represent the human condition. We have developed patient-derived xenograft (PDX) models of metastatic Wilms tumor for in vitro and in vivo biology studies and these models were found to recapitulate human tumors. Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that controls a variety of cellular pathways involved in tumorigenesis. Inhibition of FAK has been found to decrease cell proliferation and survival in a number of renal tumors, including Wilms tumor. To date, the role of FAK in metastatic Wilms tumor has not been investigated.
Methods: Cells from two PDXs of metastatic Wilms, COA 16, a lung metastasis, and COA 42, a liver metastasis, were utilized. Immunohistochemical staining was used to detect FAK protein expression and phosphorylation in the PDXs. Cells from COA 42 were treated with small molecule FAK inhibitors, PF-573228 and 1,2,4,5-benzenetetraamine tetrahydrochloride (Y15), for 24 hours at increasing concentrations. Cell viability and proliferation were assessed with alamarBlue and CellTiter 96 assays, respectively. Results were compared with student’s t-test and p≤0.05 was considered significant.
Results: Immunohistochemical staining confirmed FAK expression and phosphorylation in both COA 16 and COA 42. Inhibition of FAK with PF-573228 (10 μM) decreased cell viability by 56% and proliferation by 79% in the COA 42 cells compared to untreated control cells. Y15-induced FAK inhibition similarly resulted in decreased cell survival and proliferation. Treatment of the COA 42 cells with Y15 (10 μM) diminished cell viability by 58% and proliferation by 33% compared to untreated control cells.
Conclusion: FAK protein is expressed and phosphorylated in human metastatic lung and liver Wilms tumor PDXs. FAK inhibition with two small molecules led to decreased tumor cell viability and proliferation. These findings suggest that further exploration of FAK as a target for metastatic Wilms tumor should be undertaken.