Therapeutic targeting of the focal adhesion complex prevents oncogenic TGF-beta signaling and metastasis.
BACKGROUND
Mammary 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-beta function from a tumor suppressor to a tumor promoter. We previously showed that interaction between beta3 integrin and TbetaR-II facilitates TGF-beta-mediated oncogenic signaling, epithelial-mesenchymal transition (EMT), and metastasis. However, the molecular mechanisms by which the focal adhesion complex contributes to beta3 integrin:TbetaR-II signaling and the oncogenic conversion of TGF-beta remain poorly understood.


METHODS
FAK 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-beta1 were monitored in FAK-manipulated cells. TbetaR-II expression was increased in metastatic breast cancer cells by retroviral transduction, and the metastasis of FAK- and TbetaR-II-manipulated tumors was monitored by using bioluminescent imaging.


RESULTS
TGF-beta stimulation of MECs stabilized and activated FAK in a beta3 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-beta. Administering FAK inhibitors or rendering metastatic breast cancer cells FAK deficient abrogated the interaction between beta3 integrin and TbetaR-II, thereby preventing TGF-beta 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.


CONCLUSIONS
Our studies identify an essential function for FAK in mediating the interaction between beta3 integrin and TbetaR-II, and thus in facilitating the oncogenic conversion of TGF-beta 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-beta-dependent metastasis.
Wendt
Michael K
MK
Department of Pharmacology, University of Colorado Denver, Anschutz Medical Campus, Aurora, Colorado 80045, USA. michael.wendt@ucdenver.edu
Schiemann
William P
WP
D000818:Animals; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology / Q000628:therapy*; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D016503:Drug Delivery Systems; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D039661:Integrin beta3 / Q000378:metabolism; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism / Q000473:pathology / Q000628:therapy; D051379:Mice; D008807:Mice, Inbred BALB C; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015363:Quinolones / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000235:genetics; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013450:Sulfones / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
C521108:6-(4-(3-(methylsulfonyl)benzylamino)-5-(trifluoromethyl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one; D007211:Indoles; D039661:Integrin beta3; C527797:N-methyl-N-(3-((2-(2-oxo-2,3-dihydro-1H-indol-5-ylamino)-5-trifluoromethyl-pyrimidin-4-ylamino)-methyl)-pyridin-2-yl)-methanesulfonamide; D015363:Quinolones; D034741:RNA, Small Interfering; D018125:Receptors, Transforming Growth Factor beta; D013449:Sulfonamides; D013450:Sulfones; D016212:Transforming Growth Factor beta; D051416:Focal Adhesion Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II