Vascular attack by 5,6-dimethylxanthenone-4-acetic acid combined with B7.1(CD80)-mediated immunotherapy overcomes immune resistance and leads to theeradication of large tumors and multiple tumor foci

The promise of cancer immunotherapy is that it will not only eradicate prim ary tumors but will generate systemic antitumor immunity capable of destroy ing distant metastases, A major problem that must first be surmounted relat es to the immune resistance of large tumors. Here we reveal that immune res istance can be overcome by combining immunotherapy with a concerted attack on the tumor vasculature. The functionally related antitumor drugs 5,6-dime thylxanthenone-4-acetic acid (DMXAA) and flavone acetic acid (FAA), which c ause tumor vasculature collapse and tumor necrosis, were used to attack the tumor vasculature, whereas the T-cell costimulator B7.1 (CD80), which cost imulates T-cell proliferation via the CD28 pathway, was used to stimulate a ntitumor immunity. The injection of cDNA (60-180 mug) encoding B7.1 into la rge EL-4 tumors (0.8 cm in diameter) established in C57BL/6 mice, followed 24 h later by i.p. administration of either DMXAA (25 mg/kg) or FAA (300 mg /kg), resulted in complete tumor eradication within 2-6 weeks, In contrast, monotherapies were ineffective. Both vascular attack and B7.1 immunotherap y led to up-regulation of heat shock protein 70 on stressed and dying tumor cells, potentially augmenting immunotherapy, Remarkably, large tumors took on the appearance of a wound that rapidly ameliorated, leaving perfectly h ealed skin. Combined therapy was mediated by CD8(+) T cells and natural kil ler cells, accompanied by heightened and prolonged antitumor cytolytic acti vity (P < 0.001), and by a marked increase in tumor cell apoptosis, Cured a nimals completely rejected a challenge of 1 x 10(7) parental EL-4 tumor cel ls but not a challenge of 1 x 10(4) Lewis lung carcinoma cells, demonstrati ng that antitumor immunity was tumor specific. Adoptive transfer of 2 x 10( 8) splenocytes from treated mice into recipients bearing established (0.8 c m in diameter) tumors resulted in rapid and complete tumor rejection within 3 weeks. Although DMXAA and B7.1 monotherapies are complicated by a narrow range of effective doses, combined therapy was less dosage dependent. Thus , a broad range of amounts of B7.1 cDNA were effective in combination with 25 mg/kg DMXAA. In contrast, DMXAA, which has a very narrow range of high a ctive doses, was effective at a low dose (18 mg/kg) when administered with a large amount (180 <mu>g) of B7.1 cDNA, Importantly, combinational therapy generated heightened antitumor immunity, such that gene transfer of B7.1 i nto one tumor, followed by systemic DMXAA treatment, led to the complete re jection of multiple untreated tumor nodules established in the opposing fla nk. These findings have important implications for the future direction and utility of cancer immunotherapies aimed at harnessing patients' immune res ponses to their own tumors.