Reduction in drug-induced DNA double-strand breaks associated with β1 integrin–mediated adhesion correlates with drug resistance in U937 cells

Abstract

We previously showed that adhesion of myeloma cells to fibronectin (FN) by means of ␤1 integrins causes resistance to certain cytotoxic drugs. The study described here found that adhesion of U937 human histiocytic lymphoma cells to FN provides a survival advantage with respect to damage induced by the topoisomerase (topo) II inhibitors mitoxantrone, doxorubicin, and etoposide. Apoptosis induced by a topo II inhibitor is thought to be initiated by DNA damage. The neutral comet assay was used to determine whether initial drug-induced DNA damage correlated with cellular-adhesionmediated drug resistance. Cellular adhesion by means of ␤1 integrins resulted in a 40% to 60% reduction in mitoxantroneand etoposide-induced DNA doublestrand breaks. When the mechanisms regulating the initial drug-induced DNA damage were examined, a ␤1 integrinmediated reduction in drug-induced DNA double-strand breaks was found to correlate with reduced topo II activity and decreased salt-extractable nuclear topo II␤ protein levels. Confocal studies showed changes in the nuclear localization of topo II␤; however, alterations in the nuclear-to-cytoplasmic ratio of topo II␤ in FN-adhered cells were not significantly different. Furthermore, after a high level of salt extraction of nuclear proteins, higher levels of topo II␤-associated DNA binding were observed in FN-adhered cells than in cells in suspension. Together, these data suggest that topo II␤ is more tightly bound to the nucleus of FN-adhered cells. IntroductionStudies have found that cellular adhesion by means of ␤1 integrins inhibits cell death induced by DNA cross-linking agents and topoisomerase (topo) II inhibitors. 1,2 The mechanisms of resistance associated with ␤1-mediated adhesion are unknown. It is thought that DNA cross-linking agents and topo II inhibitors initiate cellular death by inducing DNA damage. Thus, cellular adhesion by means of ␤1 integrins could confer resistance by either decreasing drug-induced DNA damage or increasing cellular tolerance to such damage. To address this issue, we examined the effects of ␤1-mediated cellular adhesion to fibronectin (FN) on DNA damage induced by pharmacologic inhibitors of topo II. If ␤1 integrin-mediated adhesion reduces DNA damage induced by this class of drugs, then alterations in the putative target (topo II) may represent one mechanism of cellular-adhesion-mediated drug resistance (CAM-DR).Topo II is an adenosine triphosphate (ATP)-dependent enzyme that reversibly cuts double-stranded DNA and is transiently linked to the 5Ј end of the break site by phosphotyrosyl bonds. Mammalian cells contain 2 isoforms of topo II (topo II␣ and topo II␤, which are 170 kd and 180 kd, respectively). These 2 isoforms are encoded by separate human genes and differ with respect to molecular mass, sequence specificity for DNA cleavage, regulation of expression, and tissue distribution. 3,4 Many topo II inhibitors stabilize this normally transiently bound DNA-protein complex and form what is referred to as the cleavable complex. 5 Stabilization o...

ECM overrides DNA damage-induced cell cycle arrest and apoptosis in small-cell lung cancer cells through β1 integrin-dependent activation of PI3-kinase

“…30 Furthermore, integrin-mediated adhesion in hemapoeitic cancer cells reduced DNA double-strand breaks and apoptosis caused by chemotherapeutic agents such as mitoxantrone and etoposide. 31 This primarily a5b1-mediated reduction in DNA damage correlated with decreased topoisomerase-II enzymatic activity. However, the a5b1 integrin is not found in SCLC cells and is reduced in other carcinomas.…”

Irradiation differentially affects substratum-dependent survival, adhesion, and invasion of glioblastoma cell lines

“…The examination of various transformed cell lines and primary cell strains has raised evidence for chemo-and radioprotective effects of specific ECM proteins (Vlodavsky et al, 1980;Fuks et al, 1992;Rose et al, 1999;Hazlehurst et al, 2001;Cordes et al, 2002Cordes et al, in press (a), 2003Cordes and Meineke, 2003;. As reported (Bouterfa et al, 1999;Oz et al, 2000;Mahesparan et al, 2003), FN, laminin, collagen-IV and -I, decorin, tenascin and vitronectin are ECM proteins produced and secreted by cells of malignant tumours of glial origin in vitro and in vivo.…”

“…Each heterodimer consists of one a-and one b-chain. Integrins are involved in the regulation of several cell functions such as differentiation, cell cycle progression, apoptotic pathways and DNA repair mechanisms (Adams and Watt, 1990;Assoian, 1997;Hazlehurst et al, 2001). To date avb3 (Brooks et al, 1996;Rooprai et al, 1999), a5b1 (Ohnishi et al, 1998) and other b1-integrin combinations (Paulus et al, 1993;Chintala et al, 1996;Friedlander et al, 1996) have been identified to participate in cell migration and invasion.…”

Bone marrow stromal-derived soluble factors and direct cell contact contribute to de novo drug resistance of myeloma cells by distinct mechanisms

“…18,19,24,[32][33][34] MM is a latent disease of the BM, where the tumor cell phenotype likely reflects a combination of multiple interactions including cell-cell, cell-ECM, and cellgrowth factor. In this study, we demonstrate that the presence of BMSCs during treatment of H929 or RPMI 8226 cells with mitoxantrone significantly reduces apoptosis in myeloma cell lines.…”