Tumor-associated neovascularization: innate immunity and therapeutic intervention

Abstract

Tumor growth and metastasis are dependent on the development of a tumor-associated blood supply. Abrogation of this vascular network would result in tumor stasis. Conventional cancer therapy has traditionally targeted neoplastic cells directly, however, since the recognition of this critical vascular component, research efforts have incorporated a significant focus on the pathogenesis of tumor-associated vascular development. It has been established that this process is dependent on the intricate balance between pro- and anti-angiogenic factors in the tumor microenvironment and a significant number of these factors have since been identified. The mechanisms which ultimately control or shift the balance of these factors, recognized as the angiogenic switch, have been extensively studied yet remain to be clearly elucidated. Further understanding of these mechanisms is critical to the future development of diagnostic assays for early detection of cancer as well as for both preventative and therapeutic intervention strategies. Preventative therapy would inhibit initiation of the angiogenic switch while intervention strategies could shift the environment back to an antiangiogenic state or inhibit the effects of the pro-angiogenic factors. Studies presented here sought to further understand the role of endogenous angiogenic factors in the physiological tumor-bearing state, more specifically focusing on the anti-angiogenic effects of type I and II interferons. To evaluate these potential effects, intradermal tumors were induced in type I and II interferon receptor knock out (IFN-R -/-) mice with transplanted syngeneic methylcholanthrene-induced sarcoma cells. Tumor growth rates and tumor microvessel density (MVD) in both type I and II IFN-R -/- mice exceeded those of wild type (wt) mice, thus supporting an endogenous antiangiogenic role of type I and II interferons. Characterization of tumor infiltrating leukocytes revealed increased numbers of dendritic cells (DCs) in tumors from IFN-R-/- mice suggesting these interferons may elicit their antivascular effects, at least in part, by inhibiting DC differentiation or recruitment to the tumor microenvironment. Initial experiments evaluating circulating endothelial progenitor cells (CEPs) demonstrated a decrease in CEPs in IFNR-/- mice over the course of tumor development suggesting these interferons may also elicit antivascular effects by inhibiting recruitment of progenitor cells to the tumor microenvironment. Additional studies consisted of clinical trials in canines with spontaneously occurring soft tissue sarcoma. The first of the two studies sought to determine if intravenous gene therapy utilizing plasmid DNA encoding the anti-angiogenic, collagen-derived protein endostatin complexed with cationic liposomes (Cationic Liposome-DNA Complex - CLDC) could induce anti-angiogenic and anti-tumor effects. Dogs enrolled in the study were treated with either endostatin encoded or irrelevant DNA. Serial tumor biopsies were obtained to evaluate tumor MVD, transgene expression, and infiltrative leukocytes while patients were monitored clinically for tumor responses. Ultimately, > 50% of dogs in each group 1) demonstrated a significant decrease in tumor MVD at one or more time points over the course of treatment and 2) had an objective tumor response or stable disease. Additionally, transgene expression was not detected in tumor tissue from endostatin treated dogs. These results suggested that intravenous infusions of CLDCs may elicit antiangiogenic and antitumor effects in a gene independent manner. The second clinical trial sought to determine the antiangiogenic and antitumor effects of intradermal xenovaccination with recombinant human vascular endothelial growth factor (rhVEGF). Nine dogs were immunized over a 16-week period. Effects of immunization on the induction of anti-canine and anti-human VEGF antibodies, circulating plasma VEGF levels, tumor MVD, and tumor growth were assessed. The vaccine was well tolerated by all dogs and resulted in the induction of both anti-human and anti-canine VEGF antibodies in dogs which remained in the study long enough to receive five or more vaccinations (n=4). Of these four dogs, 3 had a decrease in circulating VEGF and a >50% reduction in tumor volume, while 2 dogs had a decrease in tumor MVD. Results of this pilot study suggest that multiple immunizations with xenogeneic VEGF in dogs may be a safe and effective therapy for inhibition of tumor growth and angiogenesis.