**4. Sprouting angiogenesis**

*Current Cancer Treatment*

**2. The role of VEGF in tumour angiogenesis**

and will be referred to as simply 'VEGF' from herein.

**3. Signalling in the VEGF pathway**

Angiogenesis is considered to be a fundamental event in tumour progression and metastatic dissemination and is [2] regulated by numerous endogenous factors that stimulate or inhibit neovascularisation [3]. One of the most studied pathways is the vascular endothelial growth factor (VEGF) family of ligands and their receptors [5]. In humans and mice, the VEGF family consists of 5 members: VEGF-A, -B, -C, -D and placental growth factor (PIGF). These ligands demonstrate variable specificity for the three VEGF receptors (VEGFR1, VEGFR2, VEGFR3) [3, 5]. The predominant member of the VEGF family involved in tumourigenesis is VEGF-A

One of the most important stimuli for tumour angiogenesis is hypoxia, which can occur when a rapidly growing tumour exceeds the ability of the local vasculature to supply its needs. Hypoxia-inducible factor 1 (HIF-1) is a heterodimeric transcription factor, made up of two DNA binding proteins (HIF-1α and HIF-1β), which induces the transcription of many genes, including VEGF [6]. In the presence of adequate oxygen concentrations, HIF-1α is ubiquitinated and subsequently degraded by the proteasome. However, under hypoxic conditions, HIF-1α is stabilised and persistently dimerises with the other subunit, HIF-1β, to form the HIF-1 heterodimer. The stabilised HIF-1 is then able to bind the VEGF promoter, leading to persistent transcription of the VEGF gene [7]. The expression of VEGF is also stimulated by oncogenes, including Ras, c-Src, Bcr-Abl and p53 [8]. A multitude of studies have shown that VEGF is overexpressed in the majority of solid tumours and that it is a key driver of sprouting angiogenesis [9]. Furthermore, it has been demonstrated in multiple xenograft models that VEGF inhibition supresses tumour growth [10].

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**Figure 1.**

*Signalling from VEGFR2. The signalling cascades downstream of VEGFR2 have been the best studied and are illustrated here. VEGF binding to VEGFR2 induces dimerisation of VEGFR2 and phosphorylation of tyrosine residues (indicated by the four-digit numbers in the illustration). Pathways activated include the Ras-Raf-MEK-ERK pathway, PLCγ/PKC pathway and the PI3K/Akt pathway. Activation of downstream signalling from VEGF receptors exerts control over multiple processes required for angiogenesis including endothelial cell* 

Binding of VEGF to the extracellular domain of VEGFR2 causes receptor dimerisation and phosphorylation of the receptor on tyrosine residues within the

*proliferation, migration, tube formation and vascular permeability.*

According to the established dogma, VEGF released by tumours stimulates the growth of new vessels in the following way. The VEGF diffuses through the tissue and activates endothelial cells located in local blood vessels. Firstly, VEGF receptor activation induces the selection of sprouting endothelial cells. Proteinases such as urokinase-type plasminogen activator, uPA, and members of the matrix metalloproteinase (MMP) family mediates the dissolution of the vascular basement membrane and extracellular matrix to facilitate the infiltration of sprouting endothelial cells into the surrounding tissue [16]. Next, endothelial proliferation, migration and branching allows for the formation of new vessels. This is followed by sprout fusion and lumen formation where vessels fuse together to form a network. Finally, there is perfusion and maturation. This is where the stabilisation of new blood vessels forms a functionally perfused system, which is mediated by the recruitment of pericytes to surround the newly formed endothelial tubes; recruitment of pericytes prevents further endothelial cell proliferation and migration and also suppresses vessel leakage [17].
