Cancer Therapy Vol 2, 271-278, 2004

Integrins and the cellular radiation response

Review Article

Emil Lisiak and Nils Cordes*

Bundeswehr Institute of Radiobiology, Neuherbergstrasse 11, 80937 Munich, Germany

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*Correspondence: Dr. Nils Cordes, M.D., Bundeswehr Institute of Radiobiology, Neuherbergstrasse 11, 80937 Munich, Germany; Phone: ++89 – 3168 – 3634; Fax: ++89 – 3168 – 2635; E-mail: cordes@radiation-biology.de

Key words: Integrins, radiosensitivity, ECM, tumor cells

Abbreviations: cell adhesion-mediated radioresistance, (CAM-RR); cell adhesion-mediated drug resistance, (CAM-DR); extracellular matrix, (ECM); focal adhesion kinase, (FAK); integrin-linked kinase, (ILK); ionizing radiation, (IR); mitogen-activated protein kinase, (MAPK); phosphatidylinositol-3- kinase, (PI3K); protein kinase B/AKT, (AKT)

Received: 28 July 2004; Accepted: 12 August 2004; electronically published: August 2004

Summary

Resistance of tumor cells against ionizing radiation or cytotoxic drugs is a widespread phenomenon. In addition to specific genetic alterations, microenvironmental components such as soluble growth factors and cytokines, and extracellular matrix (ECM) proteins serve as critical determinants of the cellular response to genotoxic injury. Based on recent observations, engagement of cells with ECM proteins via the integrin family of cell adhesion molecules modulates the cellular radiation survival and growth response. In dependence on cell type and cellular context, identification of specific differences between integrin signaling and integrin-growth factor receptor crosstalk may provide new facettes for innovative oncological therapies and important molecular insights into oncogenic transformation and anchorage-independent growth.

I. Introduction

During the past decade enormous progress has been made in understanding the molecular differences between normal and cancer cells. Cancer cells have defects in regulatory circuits that govern cell survival, proliferation, differentiation, and homeostasis (Jacks and Weinberg, 2002). Consequently, these defects facilitate improved insensitivity to anti-growth factors or cytotoxic agents such as ionizing radiation (IR) or drugs, evasion of programmed cell death, ability of unlimited replicative potential, relative self-sufficiency in growth signals, undirected invasion into normal tissue and formation of metastasis (Hanahan and Weinberg, 2000). Many proteins and genes are known to be involved in these cancer phenomena and modification of these processes seem critical for the development of new anti-cancer strategies. In preclinical and clinical trials, some of the new molecularly targeted treatments based on either single administration of protein kinase inhibitors or blockade of growth factor receptors or administration of these compounds in combination with chemo- or radiotherapy have indeed shown promising results (Brunner et al, 2003; Foon et al, 2004).

More recently, the influence of the cellular micromilieu has started to be investigated in the context of uncontrolled proliferation, invasion of tumors, and modification of the cellular radiation response (Park et al, 2000; Barcellos-Hoff, 2001; Henning et al, 2004). In addition to microenvironmental factors related to the vasculature or oxygen, the extracellular matrix (ECM) has been found to be a strong modifier of the reaction of transformed and normal cells to ionizing radiation (Rose et al, 1999; Cordes and Meineke, 2003; Cordes, 2004; Cordes and van Beuningen, 2004). Attachment of cells to ECM components triggers biochemical- and mechanical-induced signaling for stimulation of cytoplasmic protein kinases, growth factor receptors, ion channels, and organization of the actin cytoskeleton.

Cell-matrix binding is primarily mediated by integrins, a family of 18 a and 8 ß transmembrane glycoproteins (Hynes, 2002). To date, 24 different aß-heterodimers were identified. Most integrins recognize several ECM proteins and individual matrix proteins bind to several integrins. Activation of integrin-mediated intracellular pathways regulate signaling elements in proliferation, survival, migration, tissue organization and many more (Assoian, 1997; Giancotti and Ruoslahti, 1999; Schwartz, 2001; Watt, 2002; Zahir and Weaver, 2004; Brakebusch and Fässler, 2003). Integrin signaling and assembly of the cytoskeleton are intimately linked. As integrins bind to ECM, they associate with cytoskeletal and signaling complexes to form focal adhesions (Burridge, 1988). In this manner integrins serve as structural integrators between the ECM and the cytoskeleton, the property for which integrins are named. Via diverse cytoplasmic signaling molecules, including focal adhesion kinase (FAK), Src-family kinases and integrin-linked kinase (ILK), the stimuli generated through ligation of integrin receptors are transduced for regulating cellular events (Hannigan et al, 1996; Schaller, 2001).

II. Integrin signaling in normal and cancer cells

Normal cells such as fibroblasts, endothelial or epithelial cells need both adhesion to ECM proteins and stimulation of growth factors in order to survive and proliferate. Matrix protein binding by integrins initiates interactions of FAK with Src, which links FAK to survival and proliferation signaling via mitogen-activated protein kinase (MAPK) (Schlaepfer et al, 1997). Integrin-mediated regulation of survival furthermore involves integrin-linked kinase (ILK) and growth factor receptor-related phosphatidylinositol-3-kinase (PI3K) (Delcommenne et al, 1998). These events modulate apoptotic responses via phosphorylation of protein kinase B/AKT (AKT) and inhibition of the proapoptotic molecules caspase-9 and the Bcl-2 family member Bad (Datta et al, 1999). AKT can also activate NF-kß, which induces the expression of a set of survival factors such as osteoprotegerin and prevents release of cytochrome C from mitochondria (Hofbauer and Heufelder, 2001; Wang et al, 2001). To note, extensive investigations revealed strong synergistic cooperation between integrin- and growth factor receptor-mediated signaling, which triggered and optimized signal transduction (Moro et al, 2002; Yamada and Even-Ram, 2002) (Figure 1).

Integrins were first studied in platelets and thrombocytes (Davignon et al, 1981; Hansen and Clemmensen, 1982). Nowadays e.g. endothelial cells are intensively under investigation to identify potential therapeutic targets for inhibition of angiogenesis in tumors (Wary, 2004). In fact, both ß3 and ß1 integrin subunits are expressed by endothelial cells and regulate critical adhesive interactions with a variety of ECM proteins including fibronectin, vitronectin, laminin, collagens and fibrinogen (Ingber, 2002). Cytokine- and growth factor-stimulated

Figure 1. Schematic diagram of converging and mutually modifying integrin-RTK signaling in focal adhesions. The ligand binding, e.g. extracellular matrix proteins or growth factors, to their receptors activates downstream protein kinases. Stimulation and inhibition of the different signaling pathways as well as the crosstalk between integrin and growth factor receptor pathways optimize the regulation of cell survival, cell cycle progression, apoptosis, DNA repair, adhesion, migration, protein synthesis, and gene transcription. Further structural and signaling molecules of these membranous multiprotein signaling complexes (actin, gelsolin, talin, a-actinin) are not displayed. However, these components are substantial for the regulation of diverse cellular functions. Additionally, most of the delineated signaling events are not located at the cell membrane where they usually occur to avoid overcrowding. RTK, receptor-tyrosine-kinase; GF, growth factor.

endothelial cells express a wide spectrum of cell adhesion receptors as compared to quiescent cells (Ingber, 1992). In particular, the function of avß3 integrins was elucidated in survival and apoptotic processes with regard to the identification of a specific target molecule for innovative anti-tumor therapies (Kumar et al, 2001). This integrin is only minimally expressed on quiescent blood vessels, but highly upregulated on cells undergoing angiogenesis (Ingber, 1992). Recent studies provided promising evidence that avß3 integrins may also serve as a useful diagnostic indicator. In animal studies, anti-avß3 antibodies coupled to a gadolinium-containing liposome or iodine radioisotopes specifically targeted to tumor-associated blood vessels and, thus, proved useful in MRI and scintigraphy (Blankenberg et al, 2002). However, blockade of this integrin disturbed angiogenic processes in the chicken chorioallantoic membrane, mouse retina and rabbit cornea (Friedlander et al, 1995; Luna et al, 1996; Kumar et al, 2001). Moreover, avß3 or ß1 integrin knockout mice developed serious vascular malformations (Hodivala-Dilke et al, 1999). These observations led to the conclusion that avß3 integrin facilitates the survival of stimulated endothelial cells. In fact, systemic administration of avß3 antagonists to animals with ongoing angiogenesis showed blood vessels containing high levels of apoptotic endothelial cells. Similar effects were observed in vitro by comparing endothelial cells of different origins. Testing of a first anti-angiogenic compound, which is a specific avß3 integrin antibody, has recently started in clinical oncological trials (Eskens et al, 2003).

In contrast to normal cells, cancer cells are often able to proliferate and survive in the absence of adhesion to ECM proteins (Schwartz, 1997). These characteristics are important for cancer progression and metastases formation. Intense studies performed in a wide spectrum of cancer cells including melanoma, breast, prostate or colon gave rise to the hypothesis that resistance of cancer cells to medical treatment is partly due to alteration or loss of cell-matrix interactions (Lewis et al, 2002). Because of difficulties in demonstrating a positive or negative correlation between tumor progression and integrin expression, which was due to the heterogeneity of tumors and to the fact that changes in the expression level of a single integrin subunit always had to be judged against the background of the expression levels of all other integrins, the results are still controversial. For example, a2ß1 integrin expression seems to promote invasion in pancreatic carcinomas but is downregulated in bladder and colon cancer (Orian-Rousseau et al, 1998; Arao et al, 2000). On the other hand, a strong correlation between the enhancement of avß3 integrin expression and the metastatic potential of melanoma cells was detected, thus rating expression of avß3 as prognostic factor (Hieken et al, 1996). Contrary, expression of ß8 integrin was only observed in normal airway epithelium but not in lung cancer cells (Fjellbirkeland et al, 2003). Tumor progression and ability to form distant metastases has been correlated with the elevation of integrin-dependent protein kinases activity such as FAK or ILK (Gabarra-Niecko et al, 2003; Persad and Dedhar, 2003). A major observation in these studies was a strong converging communication between oncogenic Ras and Raf with components of integrin signaling cascades (Kinbara et al, 2003). Ras and Raf activate prosurvival pathways and are able to induce a significant degree of anchorage-independent growth.

Considering the role of integrins in malignant disease, this family of cell adhesion molecules presents as attractive drug target but the wide range of functions in different cell types is daunting. Understanding of integrin-mediated signaling, function and cross-talk with growth factor receptor-regulated pathways is crucial for the optimization and development of innovative chemo- and radiotherapeutic approaches. Nevertheless, a handful of therapeutic approaches have been initiated that implicate engineered anti-integrin antibodies or peptidomimetics with clinical success (Mousa, 2002). Most of the compounds tested with high degree of specificity were anti-avß3 or anti-RGD peptides for blocking the RGD receptor subset of integrins (Tucker, 2002). While a5ß1 integrins are predominantly involved in survival signalling and anti-RGD peptides possess anti-migratory potential, avß3-integrins mainly promote angiogenic events (Tucker, 2003). Interestingly, neither systemic increase in bleeding nor impairment of wound healing repair have yet been observed. These findings provide first successful results that can serve as basis for further detailed preclinical and clinical trials.

III. Cell-extracellular matrix interactions modulate the cellular behavior after exposure to ionizing radiation

Experimental data demonstrated that cancer cells are often more resistant against cell killing by conventional antimitotic drugs in the presence of ECM proteins compared to standard in vitro culture conditions. Cell adhesion-mediated drug resistance (CAM-DR) is thought to be partially caused by integrin signaling (Damiano et al, 1999; Cordes et al, 2004). Sethi et al, (1999) showed that adhesion of small-cell lung cancer cells to ECM protects from the chemotherapeutic agents doxorubicin, etoposide, cyclophosphamide, and cisplatin. This effect is mediated by ß1 integrins, which activate intracellular protein kinases in response to therapy-induced DNA-damage. PI3K was uncovered to participate in drug resistance of breast cancer cells (Clark et al, 2002) and increased drug resistance could also be correlated with overexpressed a4ß1 integrin in human myeloma cells (Damiano et al, 1999).

Similar findings were seen in cell adhesion-mediated radioresistance (CAM-RR) studies and the specific impact of integrin-mediated signaling on decreased cellular radiosensitivity will be outlined in detail below.

A. ECM-dependent and integrin-mediated cell survival after radiation exposure

The cytoprotective effects of physiological substrata were detected both in irradiated normal cells like skin and lung fibroblasts, endothelial cells, and keratinocytes as well as in transformed cells of different origin (lung, pancreas, skin, brain, breast, melanoma, colon) (Figure 2; Rose et al, 1999; Cordes and Meineke, 2003; Cordes and van Beuningen, 2003; Cordes and van Beuningen, 2004). Clonogenic survival experiments provided evidence that the interaction of cells with the physiological integrin ligands fibronectin, laminin, collagen-III and vitronectin before and at the time of irradiation essentially promotes survival compared to cells irradiated on culture plastic. Interestingly, this effect demonstrated only marginal variation between the different matrix proteins, complex matrix compositions like Matrigel (matrix extract from the Englebreth-Holm-Swarm mouse sarkoma) or matrix produced by bovine and human endothelial cells. Except from a variety of human glioblastoma cell lines (Cordes et al, 2003), our studies clearly show that the transformed cell lines used are all susceptible to matrix signals and that these signals strongly impact on the cellular resistance to IR. In agreement with our studies on different ILK overexpressing mutants (Cordes, 2004), Lewis et al, (2002) and Truong et al, (2003) observed adhesion to matrix as one of the major determinants for the ability to execute apoptosis after induction of genotoxic stress.

B. Adhesion of cells to ECM proteins modulates radiation-induced cell cycle arrest

Cell survival and cell cycling are closely linked cell functions. After irradiation, cells are blocked in the G1- or G2-phase of the cell cycle, which is hypothesized to provide time for DNA-repair (O`Connor, 1997). These cellular phenomena have been intensively investigated in radiobiology and are regulated by cyclins, cyclin-dependent kinases (CDK) and their inhibitors (Bernhard et al, 1999) (Figure 3). Unfortunately, only few studies have considered an impact of cell-matrix interactions on the regulation of cell cycling in irradiated cells (Gadbois et al, 1997; Dimitrijevic-Bussod et al, 1999). Recently, we showed that adhesion of normal lung fibroblasts to fibronectin enabled pronounced radiation-induced accumulation of cells in the G2/M phase of the cell cycle relative to culture plastic (Cordes and van Beuningen, 2004). Whether improved ECM-dependent survival is caused by optimization of DNA repair mechanisms at fibronectin presence is currently not known but studies to clarify this aspect are ongoing in our laboratory. Confirmatory observations were reported by Gadbois et al. (1997) and Dimitrijevic-Bussod et al. (1999) who showed that fibronectin and collagen-IV significantly reduced the portion of cells arrested in G1 and S and that radiation-induced cell cycle arrest of human normal fibroblasts is substratum-dependent.

C. Radiation-dependent modification of integrin cell surface presentation, cell motility and invasion

In vitro experiments using primary and permanent cell cultures of diverse origin demonstrated that IR is able to induce significant up-regulation of cell surface ß1-, ß3-,

Figure 2. Fibronectin-mediated radiation resistance. In contrast to polystyrene (Poly), growth of cells on the matrix protein fibronectin (FN) improves survival of irradiated human normal and transformed cells. Radiation doses ³ 2 Gy for normal skin (HSF1) and lung (CCD32) fibroblasts and for radiation doses ³ 4 Gy for A172 (glioblastoma) and A549 (lung carcinoma) cells grown on FN resulted in a p < 0.05 compared to cells grown on polystyrene. * p < 0.05.

Figure 3. Schematic diagram delineating the regulation of the G2 checkpoint response after genotoxic stress (e.g. ionizing radiation) in vitro. During G2/M-phase transition, the activity of various cell cycle proteins is tightly regulated by phosphorylation, dephosphorylation and protein interactions, mechanisms that are modulated by cell-matrix interactions. Chk1 and Chk2 phosphorylate Cdc25C, which is proposed to be critical for G2 checkpoint regulation. After binding of 14-3-3, the 14-3-3-Cdc25C complex is degraded in the cytoplasm thus preventing the activation of the Cdc25C substrate and G2-phase driving protein kinase Cdk1 (p34^cdc2) by dephosphorylation. Furthermore, Chk1 and Chk2 are able to phosphorylate p53 on serine-20, a site implicated in p53 stabilization through p53-MDM2 interactions. While subsequent induction of CDK-inhibitor p21 prevents cleavage of the pRb-E2F-complex via inhibition of cyclin D1/CDK4/CDK6 activation responsible for G1-phase arrest, p53 is also required to sustain but not to initiate a G2 arrest.

and a5-integrin expression within 48 to 96 h (Meineke et al, 2002; Cordes et al, 2002). This effect presented always dose- and frequently matrix-dependent. Interestingly, in most cell lines there was a transient decrease in ß1 or ß3 integrin cell surface presentation detectable during the first 6 h following IR. Evaluation of the adhesive functionality of the newly expressed integrin receptors revealed a strong dose-dependent elevation of cell adhesion to various matrix proteins. To assess the importance of single integrin subunits in this process, blocking experiments were carried out and identified both ß1- and ß3-integrins to play an essential role in adhesion of irradiated cells. Similar observations were reported by other investigators in endothelial cells (Hallahan et al, 2003). One elegant therapeutic approach used enhancement of avß3 integrin expression on sublethally irradiated endothelial cells undergoing angiogenesis to deliver drugs or immuno-radioisotopes directly into the irradiated endothelium of a malign tumor (Hallahan et al, 2003). This might be a reasonable strategy for tumor-specific integrin trageting in selected tumor entities.

Besides adhesion, integrins serve in invasion and migration events of normal but especially cancer cells. The impact of irradiation on these two cell functions is still controversial. Wild-Bode et al, (2001) showed enhanced migration and invasiveness of glioma cells, which correlated with elevated expression of avß3 integrins and activity of matrix-metalloproteinases (MMPs). In contrast, we showed differential invasion-impairing potential of irradiation in glioma cells (Cordes et al, 2003). Radiation-induced ß1 and ß3 integrin cell surface presentation in combination with elevated MMP-2 activity suggested that the invasion-inhibiting effect in irradiated cells is in part due to integrin-mediated local anchorage and disproportional MMP-2-mediated degradation of ECM proteins. Further controversy data on this issue indicated that exposure of endothelial cells to 6 Gy of X-rays promotes migration of endothelial cells in vitro (Sonveaux et al, 2003). Studies on the area vasculosa or the chorioallantoic membrane of the fertilized egg clearly showed that angiogenesis is inhibited after low doses of IR such as 2 Gy but is stimulated after higher doses like 10 Gy (Hatjikondi et al, 1996; Plasswilm et al, 1999).

IV. Discussion

In many cases refractory tumor responses are clinically observable during both chemo- and radiotherapy. The resistance of tumors or tumor cell clones is still a major problem to be solved and integrin-mediated cell-ECM interactions are very likely to play an essential role in this context. It is widely accepted that a combination of molecularly targeted anticancer therapies may be more effective than conventional approaches. The presented data on the role of integrin receptors in CAM-RR could provide a new, challenging, and effective basis for the development of innovative multi-targeted oncological strategies that are able to augment the cytotoxic powers against the primary tumor as well as its metastases. Further investigations on tumor-specific integrin-mediated signaling and cross-talk between integrins and growth factor receptor signaling may contribute to the identification of new molecular targets and the design of interacting substances for adjuvant chemo- and radiotherapy strategies.

References

Arao S, Masumoto A, and Otsuki M (2000) Beta1 integrins play an essential role in adhesion and invasion of pancreatic carcinoma cells. Pancreas 20, 129-137.

Assoian RK (1997) Anchorage-dependent cell cycle progression. J Cell Biol 136, 1-4.

Barcellos-Hoff MH (2001) It takes a tissue to make a tumor: epigenetics, cancer and the microenvironment. J Mammary Gland Biol Neoplasia 6, 213-221.

Beinke C, van Beuningen D, and Cordes N (2003) Ionizing radiation modulates expression and tyrosine phosphorylation of the focal adhesion-associated proteins focal adhesion kinase (FAK) and its substrates p130cas and paxillin in A549 human lung carcinoma cells in vitro. Int J Radiat Biol 79, 721-731.

Bernhard EJ, McKenna WG, and Muschel RJ (1999) Radiosensitivity and the cell cycle. Cancer J Sci Am 5, 194-204.

Blankenberg FG, Mari C, and Strauss HW (2002) Development of radiocontrast agents for vascular imaging: progress to date. Am J Cardiovasc Drugs 2, 357-365.

Brakebusch C, and Fässler R (2003) The integrin-actin connection, an eternal love affair. EMBO J 22, 2324-2333.

Brunner TB, Hahn SM, Gupta AK, Muschel RJ, McKenna WG, and Bernhard EJ (2003) Farnesyltransferase inhibitors: an overview of the results of preclinical and clinical investigations. Cancer Res 63, 5656-5668.

Burridge K, Fath K, Kelly T, Nuckolls G, and Turner C (1988) Focal adhesions: transmembrane junctions between the extracellular matrix and the cytoskeleton. Annu Rev Cell Biol 4, 487-525.

Clark AS, West K, Streicher S, and Dennis PA (2002) Constitutive and inducible Akt activity promotes resistance to chemotherapy, trastuzumab, or tamoxifen in breast cancer cells. Mol Cancer Ther 1, 707-717.

Cordes N (2004) Overexpression of hyperactive integrin-linked kinase leads to increased cellular radiosensitivity. Cancer Res, in press

Cordes N, and Beinke C (2004) Fibronectin alters A549 human lung cancer cell survival after irradiation. Cancer Biol Ther 3, 47-53.

Cordes N, and Meineke V (2003) Cell adhesion-mediated radioresistance (CAM-RR): Extracellular matrix-dependent improvement of cell survival in human tumor and normal cells in vitro. Strahlenther Onkol 179, 337-344.

Cordes N, and van Beuningen D (2003) Cell adhesion to the extracellular matrix protein fibronectin modulates radiation-dependent G2 phase arrest involving integrin-linked kinase (ILK) and glycogen synthase kinase-3b (GSK-3b) in vitro. Brit J Cancer 88, 1470-1479.

Cordes N, and van Beuningen D (2004) Arrest of human lung fibroblasts in G2 phase after irradiation is regulated by converging phosphatidylinositol-3 kinase and beta1-integrin signaling in vitro. Int J Radiat Oncol Biol Phys 58, 453-462.

Cordes N, Beinke C, Plasswilm L, and van Beuningen D (2004) Irradiation and various cytotoxic drugs enhance tyrosine phosphorylation and b1-integrin clustering in human A549 lung cancer cells in a substratum-dependent manner in vitro. Strahlenther Onkol 180, 157-164.

Cordes N, Blaese MA, Meineke V, and van Beuningen D (2002) Ionizing radiation induces up-regulation of functional b1-integrin in lung tumour cell lines in vitro. Int J Radiat Biol 78, 347-357.

Cordes N, Hansmeier B, Beinke C, Meineke V, and van Beuningen D (2003) Irradiation differentially affects substratum-dependent survival, adhesion, and invasion of glioblastoma cell lines. Brit J Cancer 89, 2122-2132.

Damiano JS, Cress AE, Hazlehurst LA, Shtil AA, and Dalton WS (1999) Cell adhesion mediated drug resistance (CAM-DR): role of integrins and resistance to apoptosis in human myeloma cell lines. Blood 93, 1658-1667.

Datta SR, Brunet A, and Greenberg ME (1999) Cellular survival: a play in three Akts. Genes Dev 13, 2905-2927.

Davignon D, Martz E, Reynolds T, Kurzinger K, and Springer TA (1981) Lymphocyte function-associated antigen 1 (LFA-1): a surface antigen distinct from Lyt-2,3 that participates in T lymphocyte-mediated killing. Proc Natl Acad Sci USA 78, 4535-4539.

Delcommenne M, Tan C, Gray V, Ruel L, Woodgett J, and Dedhar S (1998) Phosphoinositide-3-OH kinase-dependent regulation of glycogen synthase kinase 3 and protein kinase B/Akt by the integrin-linked kinase. Proc Natl Acad Sci USA 95, 11211-11216.

Dimitrijevic-Bussod M, Balzaretti-Maggi VS, and Gadbois DM (1999) Extracellular matrix and radiation G1 cell cycle arrest in human fibroblasts. Cancer Res 59, 4843-4847.

Eskens FA, Dumez H, Hoekstra R, Perschl A, Brindley C, Bottcher S, Wynendaele W, Drevs J, Verweij J, and van Oosterom AT (2003) Phase I and pharmacokinetic study of continuous twice weekly intravenous administration of Cilengitide (EMD 121974), a novel inhibitor of the integrins alphavbeta3 and alphavbeta5 in patients with advanced solid tumours. Eur J Cancer 39, 917-926.

Fjellbirkeland L, Cambier S, Broaddus VC, Hill A, Brunetta P, Dolganov G, Jablons D, and Nishimura SL (2003) Integrin alphavbeta8-mediated activation of transforming growth factor-beta inhibits human airway epithelial proliferation in intact bronchial tissue. Am J Pathol 163, 533-542.

Foon KA, Yang XD, Weiner LM, Belldegrun AS, Figlin RA, Crawford J, Rowinsky EK, Dutcher JP, Vogelzang NJ, Gollub J, Thompson JA, Schwartz G, Bukowski RM, Roskos LK, and Schwab GM (2004) Preclinical and clinical evaluations of ABX-EGF, a fully human anti-epidermal growth factor receptor antibody. Int J Radiat Oncol Biol Phys 58, 984-990.

Friedlander M, Brooks PC, Shaffer RW, Kincaid CM, Varner JA, and Cheresh DA (1995) Definition of two angiogenic pathways by distinct alpha v integrins. Science 270, 1500-1502.

Gabarra-Niecko V, Schaller MD, and Dunty JM (2003) FAK regulates biological processes important for the pathogenesis of cancer. Cancer Metastasis Rev 22, 359-374.

Gadbois DM, Bradbury EM, and Lehnert BE (1997) Control of radiation-induced G1 arrest by cell-substratum interactions. Cancer Res 57, 1151-1156.

Garcia-Barros M, Paris F, Cordon-Cardo C, Lyden D, Rafii S, Haimovitz-Friedman A, Fuks Z, and Kolesnick R (2003) Tumor response to radiotherapy regulated by endothelial cell apoptosis. Science 300, 1155-1159.

Giancotti FG, and Ruoslahti E (1999) Integrin signaling. Science 285,1028-1032.

Hallahan D, Geng L, Qu S, Scarfone C, Giorgio T, Donnelly E, Gao X, and Clanton J (2003) Integrin-mediated targeting of drug delivery to irradiated tumor blood vessels. Cancer Cell 3, 63-74.

Hanahan D, and Weinberg RA (2000) The hallmarks of cancer. Cell 100, 57-70.

Hannigan GE, Leung-Hagesteijn C, Fitz-Gibbon L, Coppolino MG, Radeva G, Filmus J, Bell JC, and Dedhar S (1996) Regulation of cell adhesion and anchorage-dependent growth by a new b1-integrin-linked protein kinase. Nature 379, 91-96.

Hansen MS, and Clemmensen I (1982) A fibronectin-binding glycoprotein from human platelet membranes. Biochem J 201, 629-633.

Hatjikondi O, Ravazoula P, Kardamakis D, Dimopoulos J, and Papaioannou S (1996) In vivo experimental evidence that the nitric oxide pathway is involved in the X-ray-induced antiangiogenicity. Br J Cancer 74, 1916-1923.

Henning T, Kraus M, Brischwein M, Otto AM, and Wolf B (2004) Relevance of tumor microenvironment for progression, therapy and drug development. Anticancer Drugs 15, 7-14.

Hieken TJ, Farolan M, Ronan SG, Shilkaitis A, Wild L, and Das Gupta TK (1996) Beta3 integrin expression in melanoma predicts subsequent metastasis. J Surg Res 63, 169-73.

Hodivala-Dilke KM, McHugh KP, Tsakiris DA, Rayburn H, Crowley D, Ullman-Cullere M, Ross FP, Coller BS, Teitelbaum S, and Hynes RO (1999) Beta3-integrin-deficient mice are a model for Glanzmann thrombasthenia showing placental defects and reduced survival. J Clin Invest 103, 229-238.

Hofbauer LC, and Heufelder AE (2001) Role of receptor activator of nuclear factor-kappaB ligand and osteoprotegerin in bone cell biology. J Mol Med 79, 243-253.

Hynes RO (2002) Integrins: Bidirectional, allosteric signaling machines. Cell 110, 673-687.

Ingber DE (1992) Extracellular matrix as a solid-state regulator in angiogenesis: identification of new targets for anti-cancer therapy. Semin Cancer Biol 3, 57-63.

Jacks T, and Weinberg RA (2002) Taking the study of cancer cell survival to a new dimension. Cell 111, 923-925.

Kasahara T, Koguchi E, Funakoshi M, Aizu-Yokota E, and Sonoda Y (2002) Antiapoptotic action of focal adhesion kinase (FAK) against ionizing radiation. Antioxid Redox Signal 4, 491-499.

Kinbara K, Goldfinger LE, Hansen M, Chou FL, and Ginsberg MH (2003) Ras GTPases: integrins' friends or foes? Nat Rev Mol Cell Biol 4, 767-776.

Kumar CC, Malkowski M, Yin Z, Tanghetti E, Yaremko B, Nechuta T, Varner J, Liu M, Smith EM, Neustadt B, Presta M, and Armstrong L (2001) Inhibition of angiogenesis and tumor growth by SCH221153, a dual alpha(v)beta3 and alpha(v)beta5 integrin receptor antagonist. Cancer Res 61, 2232-2238.

Lewis JM, Truong TN, and Schwartz MA (2002) Integrins regulate the apoptotic response to DNA damage through modulation of p53. Proc Natl Acad Sci USA 99, 3627-3632.

Luna J, Tobe T, Mousa SA, Reilly TM, and Campochiaro PA (1996) Antagonists of integrin alpha v beta 3 inhibit retinal neovascularization in a murine model. Lab Invest 75, 563-73.

Meineke, Gilbertz KP, Schilperoort K, Cordes N, Sendler A, Moede T, and van Beuningen D (2002) Ionizing radiation modulates cell surface integrin expression and adhesion of COLO-320 cells to collagen and fibronectin in vitro. Strahlenther Onkol 178, 709-714.

Moro L, Dolce L, Cabodi S, Bergatto E, Erba EB, Smeriglio M, Turco E, Retta SF, Giuffrida MG, Venturino M, Godovac-Zimmermann J, Conti A, Schaefer E, Beguinot L, Tacchetti C, Gaggini P, Silengo L, Tarone G, and Defilippi P (2002) Integrin-induced epidermal growth factor (EGF) receptor activation requires c-Src and p130Cas and leads to phosphorylation of specific EGF receptor tyrosines. J Biol Chem 277, 9405-9414.

Mousa SA (2002) Anti-integrin as novel drug-discovery targets: Potential therapeutic ad diagnostic implications. Curr Opin Chem Biol 6, 534-541.

O´Connor PM (1997) Mammalian G₁ and G₂ phase checkpoints. Cancer Surv 29, 151-182.

Orian-Rousseau V, Aberdam D, Rousselle P, Messent A, Gavrilovic J, Meneguzzi G, Kedinger M, and Simon-Assmann P (1998) Human colonic cancer cells synthesize and adhere to laminin-5. Their adhesion to laminin-5 involves multiple receptors among which is integrin alpha2beta1. J Cell Sci 111, 1993-2004.

Park CC, Bissell MJ, and Barcellos-Hoff MH (2000) The influence of the microenvironment on the malignant phenotype. Mol Med Today 6, 324-9.

Persad S, and Dedhar S (2003) The role of integrin-linked kinase (ILK) in cancer progression. Cancer Metastasis Rev 22, 375-384.

Plasswilm L, Hoper J, Cordes N, and Tannapfel A (1999) Investigation of microvessel density after irradiation. Radiat Res 151, 454-460.

Rose RW, Grant DS, O’Hara MD, and Williamson SK (1999) The role of laminin-1 in the modulation of radiation damage in endothelial cells and differentiation. Radiat Res 152, 14-28.

Schaller MD (2001) Biochemical signals and biological responses elicited by the focal adhesion kinase. Biochem Biophys Acta 1540, 1-21.

Schlaepfer DD, Broome MA, and Hunter T (1997) Fibronectin-stimulated signaling from a focal adhesion kinase-c-Src complex: involvement of the Grb2, p130cas, and Nck adaptor proteins. Mol Cell Biol 17, 1702-1713.

Schwartz MA (1997) Integrins, oncogenes, and anchorage independence. J Cell Biol 139, 575-578.

Schwartz MA (2001) Integrin signaling revisited. Trends Cell Biol 12, 466-470.

Sethi T, Rintoul RC, Moore SM, MacKinnon AC, Salter D, Choo C, Chilvers ER, Dransfield I, Donnelly SC, Strieter R, and Haslett C (1999) Extracellular matrix proteins protect small cell lung cancer cells against apoptosis: a mechanism for small cell lung cancer growth and drug resistance in vivo. Nature Med 5, 662-668.

Sonveaux P, Brouet A, Havaux X, Gregoire V, Dessy C, Balligand JL, and Feron O (2003) Irradiation-induced angiogenesis through the up-regulation of the nitric oxide pathway: implications for tumor radiotherapy. Cancer Res 63, 1012-1019.

Truong T, Sun G, Doorly M, Wang JY, and Schwartz MA (2003) Modulation of DNA damage-induced apoptosis by cell adhesion is independently mediated by p53 and c-Abl. Proc Natl Acad Sci USA 100, 10281-10286.

Tucker GC (2002) Inhibitors of integrins. Curr Opin Pharmacol 2, 394-402.

Tucker GC (2003) Alpha v integrin inhibitors and cancer therapy. Curr Opin Investig Drugs 4, 722-731.

Wang XQ, Sun P, and Paller AS (2001) Inhibition of integrin-linked kinase/protein kinase B/Akt signaling: mechanism for ganglioside-induced apoptosis. J Biol Chem 276, 44504-44511.

Wary KK (2004) Molecular targets for anti-angiogenic therapy. Curr Opin Mol Ther 6, 54-70.

Watt FM (2002) Role of integrins in regulating epidermal adhesion, growth and differentiation. EMBO J 21, 19-26.

Wild-Bode C, Weller M, Rimner A, Dichgans J, and Wick W (2001) Sublethal irradiation promotes migration and invasiveness of glioma cells: implications for radiotherapy of human glioblastoma. Cancer Res 61, 2744-2750.

Yamada KM, and Even-Ram S (2002) Integrin regulation of growth factor receptors. Nat Cell Biol 4, E75-E76.

Zahir N, and Weaver VM (2004) Death in the third dimension: apoptosis regulation and tissue architecture. Curr Opin Genet Dev 14, 71-80.

Dr. Nils Cordes