High expression of integrin alpha v beta 3 enables uptake of targeted fluorescent probes into ovarian cancer cells and tumors

The cell line OVCAR-4 was recently ranked as one of the most representative cell lines for high grade serous ovarian cancer (HGSOC). However, little work has been done to assess the susceptibility of OVCAR-4 cells and tumors to the more common types of molecular targeting. Proteome profiles suggest OVCAR-4 express high levels of integrin alpha v beta 3 receptors. Using flow cytometry with fluorescent antibodies we determined that OVCAR-4 cells have high number of integrin alpha v beta 3 receptors ([9.8 +/- 2.5] x 10(4)/cell) compared to the well-characterized cell line U87-MG ([5.2 +/- 1.4] x 10(4)/cell). However, OVCAR-4 cells also have roughly three times the surface area of U87-MG cells, so the average alpha v beta 3 receptor density is actually lower (11 +/- 3 versus 18 +/- 6 receptors/mu m(2)). A series of new fluorescent molecular probes was prepared with structures comprised of a deep-red squaraine fluorophore (similar to 680 nm emission) covalently attached to zero, one, or two cyclic pentapeptide cRGD sequences for integrin targeting. Microscopy studies showed that uptake of the divalent probe into cultured OVCAR-4 cells was 2.2 +/- 0.4 higher than the monovalent probe, which in turn was 2.2 +/- 0.4 higher than the untargeted probe. This probe targeting trend was also seen in OVCAR-4 mouse tumor models. The results suggest that clinically relevant OVCAR-4 cells can be targeted using molecular probes based on alpha v beta 3 integrin receptor antagonists such as the cRGD peptide. Furthermore, deep-red fluorescent cRGD-squaraine probes have potential as targeted stains of cancerous tissue associated with HGSOC in surgery and pathology settings. (C) 2018 Published by Elsevier Ltd.