Shiga-like toxin-VEGF fusion proteins are selectively cytotoxic to endothelial cells overexpressing VEGFR-2

Growing endothelial cells at sites of angiogenesis may be more sensitive th an quiescent endothelial cells to toxin-VEGF fusion proteins, because they express higher numbers of VEGF receptors. We have constructed, expressed an d purified a protein containing the catalytic A-subunit of Shiga-like toxin I fused to VEGF(121) (SLT-VEGF/L). SLT-VEGF/L inhibits protein synthesis i n a cell-free translation system and induces VEGFR-2 tyrosine autophosphory lation in cells overexpressing VEGFR-2 indicating that both SLT and VEGF mo ieties are properly folded in the fusion protein. SLT-VEGF/L selectively in hibits growth of porcine endothelial cells expressing 2-3 x 10(5) VEGFR-2/c ell with an IC50 of 0.1 nM, and rapidly induces apoptosis at concentrations > 1 nM. Similar results are observed with human transformed embryonic kidn ey cells, 293, engineered to express 2.5 x 10(6) VEGFR-2/cell. In contrast, SLT-VEGF/L does not affect three different types of endothelial cells (PAE /KDRlow, HUVE, MSI) expressing between 5 x 10(3) and 5 x 10(4) VEGFR-2/cell , and quiescent endothelial cells overexpressing VEGFR-2. Growth inhibition and induction of apoptosis by SLT-VEGF/L require intrinsic N-glycosidase a ctivity of the SLT moiety, but occur without significant inhibition of prot ein synthesis, The selective cytotoxicity of SLT-VEGF proteins against grow ing endothelial cells overexpressing VEGFR-2 suggests that they may be usef ul in targeting similar cells at sites of angiogenesis. (C) 2001 Elsevier S cience BY. All rights reserved.