Understanding Mechanisms of Resistance to Anti-angiogenic Treatments

DESCRIPTION (provided by applicant): At present survival from malignant glioblastoma (GBM, brain tumor) is usually less than a year. GBM are characterized by release of vascular endothelial growth factor (VEGF), an important regulator and promoter of new vessel formation (angiogenesis) and vascular permeability. Because of initial positive responses noted in clinical trials, it was thought that anti-angiogenic therapy targeting VEGF or VEGF receptors (VEGFRs) would become an effective tool for controlling GBM. However, it is now well recognized that the positive responses are temporary and that the tumors develop resistance and eventually progress, and in some cases with a more aggressive and invasive phenotype. To overcome the failure, it is essential to understand the basic mechanism of resistance to anti-angiogenic therapy. We hypothesize that anti-angiogenic treatment using VEGFR inhibitors (such as vetanalib) will cause hypoxia due to vascular loss and decreased endothelial cell (EC) survival, which will enhance compensatory increases in other pro-angiogenic factors such as stromal-cell derived factor 1 (SDF-1). SDF-1 in turn will induce the mobilization, migration and accumulation of endothelial progenitor cells (EPCs) around or into the tumor. Bone marrow (BM) derived precursor cells are known to promote angiogenesis and pro-growth responses and may be a mechanism for resistance. However it is not known whether EPCs accumulate in the tumor during anti VEGF therapy, or whether decreasing BM cells or neutralizing SDF-1 helps overcome resistance to anti- angiogenic therapy. It would be advantageous to use novel compounds that target multiple sites of angiogenesis to overcome anti-angiogenic resistance. HET0016 is one of the compounds that target multiple sites of angiogenesis. In this proposal we'll make orthotopic human glioma tumor model in nude rat. Tumor bearing animals will be treated either with vetanalib, or HET0016 alone or in combination. Dynamic contrast enhanced MRI (DCE-MRI) will be performed to determine permeability transfer constant (Ktrans), distribution (tumor blood) volume, tumor volume, enhancement pattern, and diffusion parameters in the tumors under basal and treated conditions. Western blot and RT-PCR analysis will determine the expression level of different angiogenic factors/receptors while immunohistochemistry will assess the vascular density and morphology. We expect that treatment resistance tumor will show changes in size, permeability, tumor blood volume, and other MRI parameters. The data may give direct evidence of compensatory/refractory angiogenesis, explain why certain tumors become refractory to anti VEGFR therapy and identify HET0016 as a possible adjuvant therapy in surmounting the tumor resistance to anti-angiogenic therapy. Previous studies by our group showed that transplanted EPCs migrated and incorporated in the tumor angiogenesis due to chemotactic cytokines released from the tumors (such as HIF-1 mediated SDF-1 release). To determine whether decreasing available BM cells or blocking accessible SDF-1, will diminish or even overcome resistance to anti VEGFR or HET0016 therapy. We will use sub lethal irradiation to lower the ability of the BM to release pro-angiogenic cells. In a set of experiments we will also block SDF-1 using specific antibodies or CXCR4 antagonists and thus reduce the ability of EPCs to migrate to the tumor. The initial migration and incorporation of intravenously administered EPCs will be detected by SPECT using In-111 labeled cells and the long term incorporation of the administered cells will be detected by cellular MRI using magnetically labeled cells. These data will be validated by immunohistochemistry. We expect the results of this experimental proposal will shade lights on the mechanisms of resistance to anti- angiogenic treatments and allow clinician to change the strategy of future treatment for GBM or other solid tumors. PUBLIC HEALTH RELEVANCE: It is likely that multiple mechanisms can be brought into play in different tumors to explain evasive and intrinsic resistance to anti-angiogenic therapy. Mobilization and homing of monocytic progenitor cells to the tumor, which contribute to vasculogenesis and release multiple angiogenic and growth factors, may be an important contributor to this resistance. The use of accurate and sensitive non-invasive techniques to document changes in the tumor in the context of anti-angiogenic therapy and EPC's involvement represent a novel aspect. The use of cell labeling will allow to track cells non-invasively, an approach that might be possible to apply in a clinical setting. The use of a new agent, HET0016, in addition to vetanalib (PTK787), to get around the resistance to anti-angiogenic therapy is, we believe, innovative. Even a partial beneficial response would be highly relevant.