Acquired resistance to the antitumor effect of epidermal growth factor receptor-blocking antibodies in vivo: A role for altered tumor angiogenesis

Inhibitors of epidermal growth factor receptor (EGFR) signaling are among t he novel drugs showing great promise for cancer treatment in the clinic, Ho wever, the possibility of acquired resistance to such drugs because of tumo r cell genetic instabilities has not yet been explored. Here we report the experimental derivation and properties of such cell variants obtained from recurrent tumor xenografts of the human A431 squamous cell carcinoma, after two consecutive cycles of therapy with one of three different anti-EGFR mo noclonal antibodies: mR3, hR3, of C225. Initial response to a 2-week period of treatment was generally fetal tumor regression and was not significantl y different among the three antibody groups. However, tumors often reappear ed at the site of inoculation, generally after prolonged latency periods, a nd most of the tumors became refractory to a second round of therapy, Cell lines established from such resistant tumors retained high EGFR expression, normal sensitivity to anti-EGFR antibody or ligand, and unaltered growth r ate when compared with the parental line in vitro, Tn contrast, the A431 ce ll variants exhibited an accelerated growth rate and a significantly attenu ated response to anti-EGFR antibodies in vivo relative to the parental line . Because of the reported suppressive effect of EGFR inhibitors on vascular endothelial growth factor(VEGF) expression, and the demonstrated role of V EGF in the angiogenesis and growth of A431 tumor xenografts, relative VEGF expression was examined. Five of six resistant variants expressed increased levels of VEGF, which paralleled an increase in both angiogenic potential in vitro and tumor angiogenesis in vivo. In addition, elevated expression o f VEGF in variants of A431 cells obtained by gene transfection rendered the cells significantly resistant to anti-EGFR antibodies in vivo. Taken toget her, the results suggest that, at least in the A431 system, variants displa ying acquired resistance to anti-EGFR antibodies can emerge in vivo and can do so, at least in part, by mechanisms involving the selection of tumor ce ll subpopulations with increased angiogenic potential.