Module preservation Brains were removed from each animal after euthanasia

ed, modelling studies were employed to suggest the location of this allosteric site. The models predict a contribution from Leu173, located in the second extracellular loop, and in differing models, contributions from Arg 7.35 of the orthosteric binding site that, as PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19809862 noted earlier, is central to the binding/function of the short-chain fatty acids. A series of hydrophobic residues located in TMDs II, III, VI and VII were also suggested to contribute to the binding of the p-chlorophenyl element of the ligands. Recently, Smith et al. have begun to explore this issue experimentally, taking advantage of the orthosteric binding pocket mutants studied by Stoddart et al.. Although able to confirm both the direct agonist and positive allosteric effects of chloro-a–N-2-thiazolylbenzeneacetamide, as well as demonstrating the selectivity of this ligand for FFA2 over FFA3 and FFA1, Smith et al. were unable to provide evidence to support a key role for Arg 7.35 in the recognition/function of this ligand because chloro-a–N-2-thiazolylbenzeneacetamide was at least as potent and efficacious in causing inhibition of cAMP production at Arg 7.35 Ala FFA2 as at the wild-type receptor. The second extracellular loop differs in length and is relatively diverse between FFA2 and FFA3 and such loops present a substantial challenge to model. However, at least based on a simple alignment, Leu173 in FFA2 is conserved as Leu178 in FFA3, although in each receptor UNC0642 site charged amino acids located close to this residue are not well conserved. As Sum et al. have recently provided evidence that negatively charged amino acids in extracellular loop 2 play an important role in generating an inactive state of FFA1 by co-ordinating Arg 5.39 and Arg 7.35, it may be that similar interactions contribute to the ground state of FFA2 and FFA3, but this remains to be tested. Selective ligands for FFA3, whether orthosteric or ago-allosteric, would be of great value to understanding the biology and function of this receptor. are required. The recent identification of a pair of phenylacetamides as selective ligands at FFA2 that act as both secondsite agonists and positive allosteric modulators of the endogenous free fatty acids at this receptor is a helpful first step, but much remains to be achieved. Acknowledgements The Biotechnology and Biosciences Research Council grant BB/E019455/1 supports a small section of the work described herein. L.A.S. was funded by a BBSRC CASE studentship and N.J.S. is an NHMRC/NHF of Australia C.J. Martin Overseas Fellow. Conflict of interest The authors state no conflict of interest. Angiogenesis plays an important role in the growth of solid tumours; and inhibition of angiogenesis prevents tumour growth and/or progression in experimental models of cancer. Angiogenesis is regulated by many factors, such as VEGF and its receptors, which play important roles in regulating neovascularization and tumour angiogenesis. VEGF binds to transmembrane receptors expressed on vascular endothelial cells and lymphatic vessels, and regulates numerous functions including EC migration, proliferation, protease expression, microvascular integrin expression, as well as capillary tube formation. The VEGFR family has five members, of which the main subtypes are VEGFR1, VEGFR2 and VEGFR3. VEGFR1-3 are exclusively located on the surface of ECs in normal tissues and are up-regulated only during embryonic and tumour angiogenesis. Moreover, VEGFR2 is the major effector of angiogenesis