Chapter 080. Cancer Cell Biology and Angiogenesis (Part 16) Normalization of tumor blood vessels due to inhibition of VEGF signaling. A. Blood vessels in normal tissues exhibit a regular hierarchical branching pattern that delivers blood to tissues in a spatially and temporally efficient manner to meet the metabolic needs of the tissue (top). At the microscopic level, tight junctions are maintained between endothelial cells (EC), which are adherent to a thick and evenly distributed basement membrane (BM). Pericytes form a surrounding layer that provides trophic signals to the EC and helps maintain proper vessel tone. Vascular permeability is regulated, interstitial fluid pressure is low, and oxygen tension and pH are physiologic. B. Tumors have abnormal vessels with tortuous branching and dilated, irregular interconnecting branches, causing uneven blood flow with areas of hypoxia and acidosis. This harsh environment selects genetic events that result in resistant tumor variants, such as the loss of p53. High levels of VEGF (secreted by tumor cells) disrupt gap junction communication, tight junctions, and adherens junctions between EC via src-mediated phosphorylation of proteins such as connexin 43, zonula occludens- 1, VE-cadherin, and α/β-catenins. Tumor vessels have thin, irregular BM, and pericytes are sparse or absent. Together, these molecular abnormalities result in a vasculature that is permeable to serum macromolecules, leading to high tumor interstitial pressure, which can prevent the delivery of drugs to the tumor cells. This is made worse by the binding and activation of platelets at sites of exposed BM, with release of stored VEGF and microvessel clot formation, creating more abnormal blood flow and regions of hypoxia. C. In experimental systems, treatment with bevacizumab or blocking antibodies to VEGFR2 leads to changes in the tumor vasculature that has been termed vessel normalization. During the first week of treatment, abnormal vessels are eliminated or pruned (dotted lines), leaving a more normal branching pattern. ECs partially regain features such as cell-cell junctions, adherence to a more normal BM, and pericyte coverage. These changes lead to a decrease in vascular permeability, reduced interstitial pressure, and a transient increase in blood flow within the tumor. Note that in murine models, this normalization period lasts only for ~5–6 days. D. After continued anti-VEGF/VEGFR therapy (which is often combined with chemo- or radiotherapy), ECs die, leading to tumor cell death (either due to direct effects of the chemotherapy or lack of blood flow). Unlike normal blood vessels, the vascular lining of tumor vessels is not a homogeneous layer of ECs but often consists of a mosaic of ECs and tumor cells; the concept of cancer cell–derived vascular channels, which may be lined by ECM secreted by the tumor cells, is referred to as vascular mimickry. It is unclear whether tumor cells actually form structural elements of vascular channels or represent tumor cells in transit into or out of the vessel. However, the former is supported by evidence that in some human colon cancers, tumor cells can comprise up to 15% of vessel walls. The ECs of angiogenic blood vessels are unlike quiescent ECs found in adult vessels, where only 0.01% of ECs are dividing. During tumor angiogenesis, ECs are highly proliferative and express a number of plasma membrane proteins that are characteristic of activated endothelium, including growth factor receptors and adhesion molecules such as integrins. Mechanisms of Tumor Vessel Formation Tumors utilize a number of mechanisms to promote their vascularization, and in each case they subvert normal angiogenic processes to suit this purpose (Fig. 80-8). Primary or metastatic tumor cells sometimes arise in proximity to host blood vessels and grow around these vessels, parasitizing nutrients by coopting the local blood supply. However, most tumor blood vessels arise by the process of sprouting, in which tumors secrete trophic angiogenic molecules, the most potent being VEGF, that induce the proliferation and migration of host ECs into the tumor. Sprouting in normal and pathogenic angiogenesis is regulated by three families of transmembrane RTKs expressed on ECs and their ligands (VEGFs, angiopoietins, ephrins; Fig. 80-9), which are produced by tumor cells, inflammatory cells, or stromal cells in the tumor microenvironment. When tumor cells arise in or metastasize to an avascular area, they grow to a size limited by hypoxia and nutrient deprivation. Hypoxia, a key regulator of tumor angiogenesis, causes the transcriptional induction of the gene encoding VEGF by a process that involves stabilization of HIF-1α. Under normoxic conditions, HIF-1α levels are maintained at a low level by proteasome-mediated destruction regulated by a ubiquitin E3-ligase encoded by the VHL tumor- suppressor locus. However, under hypoxic conditions, HIF-1α is not hydroxylated and association with VHL does not occur; therefore HIF-1 levels increase, and target genes including VEGF, nitric oxide synthetase (NOS), and Ang2 are induced. Loss of the VHL genes, as occurs in familial and sporadic renal cell carcinomas, results in HIF-1α stabilization and induction of VEGF. Most tumors have hypoxic regions due to poor blood flow, and tumor cells in these areas stain positive for HIF-1α expression; in renal cancers with VHL deletion, all of the tumor cells express high levels of HIF-1α, and VEGF-induced angiogenesis leads to high microvascular density (hence the term hypernephroma). . Chapter 080. Cancer Cell Biology and Angiogenesis (Part 16) Normalization of tumor blood vessels due to inhibition of VEGF. but often consists of a mosaic of ECs and tumor cells; the concept of cancer cell derived vascular channels, which may be lined by ECM secreted by the tumor cells, is referred to as vascular mimickry proliferation and migration of host ECs into the tumor. Sprouting in normal and pathogenic angiogenesis is regulated by three families of transmembrane RTKs expressed on ECs and their ligands (VEGFs,