Anti-angiogenic effects and regression of localized murine AML produced by anti-VEGF and anti-Flk-1 antibodies

Reducing the blood supply of tumors is one modality to combat cancer. The objective of this study was to evaluate such an approach in the treatment of localized murine AML (acute myelogenous leukemia). For this purpose we designed an experimental model in which leukemic cells were embedded in 1% agar discs before subcutaneous implantation in C57Bl female mice. The C-1498 AML cell line (Frederick Inst., NCI, MD, USA) was used. Thirty experimental mice received on alternate days injections of 5 x 2.5 microg anti-VEGF (vascular endothelial growth factor) and 5 x 2.5 microg anti-Flk-1 (VEGFR2) antibodies to the site of cell implantation over a period of 10 d. Fifteen control mice received daily PBS injections. All mice were sacrificed 16 d after AML implantation. Of the 30 experimental animals, macroscopic examination showed in 21 animals (70%) small sized, pale tumors (0.5 g); in six mice (20%) the tumors were replaced completely by necrotic tissue, while in three mice (10%), there were large (2.5 g), highly vascularized tumors. In all 15 control mice large highly vascularized tumors were seen. A separate group of mice was studied for total survival following AML implantation. While 12 mice in the control group not treated with antibodies survived for 16 d post-implantation, survival was prolonged in 15 antibody treated mice by approximate 30 d to a total survival time of 48 d. Tumor specimens were processed for histology, immunohistochemistry (IHC) for CD31 endothelial cell antigen, and tube-like formation assay. The small, pale tumors of antibody treated animals consisted of degenerate hyaline material with remnant nests of leukemic cells, whereas large tumors showed sheets of leukemic cells and numerous blood vessels. Specimens processed for CD31 antigen showed scarce or absence of blood vessels in the small, pale tumors in contrast to intensive staining from a rich network of blood vessels in the large, highly vascularized tumors. Tube-like formation assays disclosed rudimentary Grade 1 endothelial cell tubes in the small, pale tumors as opposed to polygonal Grade 4 tube formation in control animals. In conclusion, this murine model of localized AML allows assessment of anti-angiogenic tumor regression. Anti-angiogenic antibodies against VEGF and Flk-1 have therapeutic effects in murine AML.