[CANCER RESEARCH 61, 2022–2030, March 1, 2001] Stimulation of  1 Integrin Down-Regulates ICAM-1 Expression and ICAM-1-dependent Adhesion of Lung Cancer Cells through Focal Adhesion Kinase 1 Manabu Yasuda, Yoshiya Tanaka, 2 Masahito Tamura, Koichi Fujii, Masakazu Sugaya, Tomoko So, Mitsuhiro Takenoyama, and Kosei Yasumoto Second Department of Surgery [M. Y., M. S., T. S., M. Tak., K. Y.], First Department of Internal Medicine [Y. T., K. F.], and Kidney Center [M. Tam.], School of Medicine, University of Occupational and Environmental Health, Japan, Kitakyushu 807-8555, Japan ABSTRACT Adhesion molecules are involved in intracellular signaling in various physiological and pathological processes including metastasis and growth of tumor cells. Tumor cells interact with various host cells as well as with extracellular matrices through certain adhesion molecules such as inte- grins. We here propose that stimulation of 1 integrin reduces intercel- lular adhesion molecule (ICAM)-1-mediated interaction of lung cancer cells with CTLs. This concept is based on the following findings: (a) engagement of 1 integrins on certain lung cancer cells by a specific antibody or by ligand matrices such as fibronectin and collagen markedly reduced ICAM-1 expression on the cell surface and induced sICAM-1; (b) down-regulation of ICAM-1 by stimulation of 1 integrins was abrogated by tyrosine kinase inhibitors or by transfection of dominant negative truncations of focal adhesion kinase (FAK); (c) engagement of 1 inte- grins also reduced ICAM-1-dependent adhesion of lung cancer cells to T cells, a process completely inhibited by tyrosine kinase inhibitors and by transfection of dominant negative forms of FAK; and (d) stimulation of 1 integrins prevented killing of lung cancer cells by autologous CTLs. In malignant tumors, cancer cells, including lung cancer cells, are sur- rounded by extracellular matrix proteins such as fibronectin and collagen. This suggests that the engagement of 1 integrins by matrix proteins potentially occurs in cancer cells in vivo and that continuous stimulation via 1 integrins reduces ICAM-1-expression, ICAM-1-mediated adhesion of cancer cells to CTLs and their killing by CTLs. Our results suggest that such processes can lead to the escape of lung cancer cells in vivo from immunological surveillance. INTRODUCTION Metastasis, the spread of cells from the primary neoplasm to distant sites and their growth at that location, is the most fearsome aspect of cancer. Despite significant improvements in early diagnosis, surgical techniques, general patient care, and local and systemic adjuvant therapies, most deaths from cancer are attributable to metastases that are resistant to conventional therapies. During the metastatic cascade, tumor cells interact with various host cells as well as with extracel- lular matrices and basement membrane components including lami- nin, fibronectin, and type I collagen through certain adhesion mole- cules such as integrins (1–3). Such adhesive interaction may lead to the enhancement of survival, arrest, or invasiveness of tumor cells and is one of the most important events in the metastatic process (4 –7). Adhesion molecules are involved in intracellular signaling in a variety of physiological and pathological processes including metas- tasis and tumor growth. Recent studies have indicated that certain adhesion molecules serve not only as adhesive substances but also regulate several cellular functions by influencing signaling, desig- nated “outside-to-in signaling.” The many better-known molecules are integrins 1 and 2 and CD28, which induce costimulatory signals in the binding of T cells to antigen-presenting cells via multiple cellular signaling molecules, including FAKs, 3 resulting in cell activation and cytokine production (8). Cell adhesion to matrices is primarily medi- ated by integrins, cell-surface receptors that comprise an expanding family of transmembrane heterodimers of  and  subunits (9 –12). Several studies have demonstrated that integrins play a key role in the malignant behavior of neoplastic cells (13–18). Interaction of inte- grins with their protein ligands increases tyrosine phoshorylation and triggers the assembly of cytoskeletal proteins, signaling enzymes, and their substrates into membrane-substratum junctions referred to as focal adhesions (19 –22). The expression and function of adhesion molecules are regulated through intracellular signaling induced by several cellular stimuli, a process designated “inside-to-out signaling.” Among these molecules, the expression of ICAM-1 is tightly regulated by locally produced inflammatory cytokines such as IL-1, tumor necrosis factor , IL-6, and IFN- (23, 24). The ICAM-1/LFA-1 pathway regulates important cell-cell interactions such as leukocyte adhesion and migration, espe- cially the killing of tumor cells by natural killer cells and CTLs (25, 26). Although various tumor cells are known to highly express ICAM-1, a potent ligand for LFA-1 on CTL in vitro, many tumor cells remain viable against killing by CTL in vivo. However, the regulation of ICAM-1 expression on cancer cells is still unclear. In the present study, we examined the role of 1 integrin-mediated signaling in the regulation of cell surface adhesion molecules using lung cancer cells. Our results showed that the engagement of 1 integrin by a specific Ab or ligand matrices reduced ICAM-1 expres- sion through tyrosine kinases and FAK, which subsequently resulted in reduced adhesion of lung cancer cells to T cells and which pro- tected cancer cells from CTL-mediated cytotoxicity. MATERIALS AND METHODS The study protocol was approved by the Human Ethics Review Committee of the University of Occupational and Environmental Health, Japan ( Kitak- yushu, Japan), and a signed consent form was obtained from each subject prior to taking tissue samples used in the present study. Tumor Cell Lines. Eleven human lung cancer cell clones were used in the present study; A904L, C831L (lung large cell carcinoma), A110L, C422L (lung adenocarcinoma), and B1203L (lung squamous cell carcinoma). These cell lines were established in our laboratory as described previously (27–31). PC-9 and A549 were derived from lung adenocarcinoma (30) and QG56 was derived from lung squamous cell carcinoma (29). PC-1, PC-6 (32), and QG90 (28) were derived from lung small cell carcinoma. All of the cell lines were Received 1/26/00; accepted 12/27/00. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan. 2 To whom requests for reprints should be addressed, at The First Department of Internal Medicine, University of Occupational and Environmental Health, Japan, School of Medicine, 1-1 Iseigaoka, Yahatanishi-ku, Kitakyushu 807-8555, Japan. Phone: 81-93- 603-1611, extension 2426; Fax: 81-93-691-9334; E-mail: tanaka@med.uoeh-u.ac.jp. 3 The abbreviations used are: FAK, focal adhesion kinase; FAT, focal adhesion targeting domain; FRNK, FAK-related nonkinase; ICAM-1, intercellular adhesion mol- ecule-1; LFA-1, leukocyte function-associated antigen-1; Ab, antibody; mAb, monoclonal Ab; PI, phosphatidylinositol; RLNL, regional lymph node lymphocyte; FACS, fluores- cence-activated cell sorting; IL, interleukin; HSA, human serum albumin; sICAM-1, soluble ICAM-1; PMA, phorbol 12-myristate 13-acetate. 2022 on May 11, 2017. © 2001 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from