Abstract
Purpose:
Based on a number of clinical, experimental and genetic data, lipid accumulation, oxidant stress and inflammation appear to partake in the pathogenesis of age-related macular degeneration (AMD). CD36 is an interesting target given its pleiotropic actions on uptake of oxidized lipids, induction of inflammation and oxidant stress, and modulation of angiogenesis through the above actions as well as by being the receptor for the potent anti-angiogenic factor thrombospondin-1; of interest these angiogenic modulatory properties are largely independent of VEGF. Using a rational medicinal chemistry approach we proceeded to discover lead CD36 modulators that could be effective in inhibiting CNV.
Methods:
Based on a prototypical CD36 ligand EP80317,we generated a library of novel azapeptides(YGR100, YGR212-2, YGR212-4, CP-08-24 et CP-07-95, Aza-Lys [negative control]) , that were tested for their biological activities, in vitro and in vivo. Using primary (peritoneal) macrophages stimulated with a ligand for TLR2 (R-FSL1; a danger/pathogen-associated molecular pattern receptor which interacts with CD36), we screened for induction of various cytokines/chemokines with Elisa kit. We verified the physiologic impact of these observations in vivo. In a validated model of AMD in Apo-E-null mice illuminated with 1000 lux blue light for 5 days, we quantified macrophage/microglia (Iba-1-immunopositive) accumulated CD36-dependently in the sub-retina. The angiogenic effect of these CD36 ligand was tested with an established model of neovascularization of choroidal explants ex vivo before to be used in an in vivo model of laser impact-induced CNV.
Results:
All new azapeptides effectively inhibited R-FSL1-induced pro-inflammatory TNF, IL-6 (except for CP-07-95), and MCP-1, but did not affect anti-inflammatory IL-10; as expected, efficacy of azapeptides was not detected on macrophage from CD36-null mice. In vivo, the accumulation of macrophage/microglia in the sub-retina induced by illumination was abrogated by all azapeptides except CP-07-95 and CP-08-24. Ex vivo, we found that YGR100, YGR212-4 and CP-08-24, were effective in markedly reducing choroidal vascular sprouting in choroidal explants.
Conclusions:
Our findings have uncovered novel CD36 modulators YGR100, YGR212-4 and CP-08-24, exhibiting common aza-amino acid motifs, as potential candidates for CNV in ‘wet’ AMD.