Angelina Pavlic (ESR 4) - Targeted treatment of vascular calcification

Vascular calcification is an independent risk factor for the development of cardiovascular disorders and currently no therapies are available to treat or prevent it. My project is based around identifying the molecular machinery that modulates vascular calcification with a focus on molecular targets that drive vesicle release. One of these targets: Sphingomyelin phosphodiesterase-3 (SMPD3) is currently recognized as a key signaling molecule in extracellular vesicle release and calcification.

The objective of my project will be to identify inhibitors of SMPD3 activity, by applying 3D homology modeling in a high throughput structure based virtual screen to find small molecule inhibitors. The CRISPR/Cas9 system will subsequently be used to generate a testing system for the in vitro screening of the small molecules. Cell-based assays will also be used as a readout along with confocal imaging of SMPD3-GFP trafficking to determine the effect of small molecules. The best scored compounds will be tested in mouse models of vascular calcification (apoE^-/- and LDLR^-/-) to validate the potential of selected compounds in vivo. Our in-depth research combining computational modeling, virtual ligand screening and molecular, cellular and animal biology will propel our understanding of vascular calcification and reveal potential molecular targets that can be inhibited using pharmacological interventions.

The 2 confocal images on the right show the localisation of SMPD3 in HEK293 cells. In the upper panel HEK293 cells have been transfected with EGFP-SMPD3 (green). The nuclei have been stained with DAPI, in blue, and the F-actin in red (Phalloidin Texas-Red). In the lower panel HEK293 cells have been transfected with EGFP-SMPD3 (green), the plasma-membrane is stained in red (PKH26). The nuclei have been stained with DAPI, in blue. Both images indicate that EGFP-SMPD3 appears to mostly colocalize with the plasma membrane.