Therapeutic targeting of the focal adhesion complex prevents oncogenic TGF-β signaling and metastasis

Abstract

IntroductionMammary tumorigenesis is associated with the increased expression of several proteins in the focal adhesion complex, including focal adhesion kinase (FAK) and various integrins. Aberrant expression of these molecules occurs concomitant with the conversion of TGF-β function from a tumor suppressor to a tumor promoter. We previously showed that interaction between β3 integrin and TβR-II facilitates TGF-β-mediated oncogenic signaling, epithelial-mesenchymal transition (EMT), and metastasis. However, the molecular mechanisms by which the focal adhesion complex contributes to β3 integrin:TβR-II signaling and the oncogenic conversion of TGF-β remain poorly understood.MethodsFAK expression and activity were inhibited in normal and malignant mammary epithelial cells (MECs) either genetically by using lentiviral-mediated delivery of shRNAs against FAK, or pharmacologically through in vitro and in vivo use of the FAK inhibitors, PF-562271 and PF-573228. Altered Smad2/3 and p38 MAPK activation, migration, EMT, and invasion in response to TGF-β1 were monitored in FAK-manipulated cells. TβR-II expression was increased in metastatic breast cancer cells by retroviral transduction, and the metastasis of FAK- and TβR-II-manipulated tumors was monitored by using bioluminescent imaging.ResultsTGF-β stimulation of MECs stabilized and activated FAK in a β3 integrin- and Src-dependent manner. Furthermore, by using the human MCF10A breast cancer progression model, we showed that increased FAK expression in metastatic breast cancer cells mirrored the acquisition of enhanced activation of p38 MAPK by TGF-β. Administering FAK inhibitors or rendering metastatic breast cancer cells FAK deficient abrogated the interaction between β3 integrin and TβR-II, thereby preventing TGF-β from (a) activating p38 MAPK; (b) stimulating MEC invasion, migration, and EMT; and (c) inducing early primary tumor dissemination to the lungs. Finally, in contrast to FAK depletion, adjuvant FAK chemotherapy of mammary tumors decreased their growth in part by diminished macrophage tumor infiltration.ConclusionsOur studies identify an essential function for FAK in mediating the interaction between β3 integrin and TβR-II, and thus in facilitating the oncogenic conversion of TGF-β required for mammary tumor metastasis. Furthermore, this study establishes chemotherapeutic targeting of FAK as an effective, two-pronged approach in preventing tumor progression both by decreasing innate immune cell infiltration, and by inhibiting early TGF-β-dependent metastasis.