Abstract
Similar to other integrin-targeting strategies, disintegrins have previously shown good efficacy in animal cancer models with favorable pharmacological attributes and translational potential. Nonetheless, these polypeptides are notoriously difficult to produce recombinantly due to their particular structure requiring the correct pairing of multiple disulfide bonds for biological activity. Here, we show that a sequence-engineered disintegrin (called vicrostatin or VCN) can be reliably produced in large scale amounts directly in the oxidative cytoplasm of Origami B E. coli. Through multiple integrin ligation (i.e., αvβ3, αvβ5, and α5β1), VCN targets both endothelial and cancer cells significantly inhibiting their motility through a reconstituted basement membrane. Interestingly, in a manner distinct from other integrin ligands but reminiscent of some ECM-derived endogenous anti-angiogenic fragments previously described in the literature, VCN profoundly disrupts the actin cytoskeleton of endothelial cells (EC) inducing a rapid disassembly of stress fibers and actin reorganization, ultimately interfering with EC's ability to invade and form tubes (tubulogenesis). Moreover, here we show for the first time that the addition of a disintegrin to tubulogenic EC sandwiched in vitro between two Matrigel layers negatively impacts their survival despite the presence of abundant haptotactic cues. A liposomal formulation of VCN (LVCN) was further evaluated in vivo in two animal cancer models with different growth characteristics. Our data demonstrate that LVCN is well tolerated while exerting a significant delay in tumor growth and an increase in the survival of treated animals. These results can be partially explained by potent tumor anti-angiogenic and pro-apoptotic effects induced by LVCN.
Highlights
Despite the tremendous progress made in the last decades in deciphering the molecular intricacies of various signaling circuitries that operate aberrantly in cancers and the therapeutic advancement seen with some of the newer anti-cancer modalities recently approved by the FDA, such as humanized monoclonal antibodies directed at VEGF-A and receptor tyrosine kinases [1,2] or non-peptide tyrosine kinase inhibitors [3,4], the 5-year prognosis for most solid tumors remains reserved
Another advantage of expressing heterologous proteins fused to thioredoxin A (TrxA) is the high solubility of this bacterial protein, the result of which is that TrxA internally chaperones the recombinant protein fused to it keeping it in solution and allowing for higher levels of foreign protein accumulation in the cytoplasm [40]
Tumor angiogenesis involves a still poorly understood crosstalk between transformed epithelial cells and quiescent endothelial cells originating from preexisting vessels
Summary
Despite the tremendous progress made in the last decades in deciphering the molecular intricacies of various signaling circuitries that operate aberrantly in cancers and the therapeutic advancement seen with some of the newer anti-cancer modalities recently approved by the FDA, such as humanized monoclonal antibodies directed at VEGF-A (vascular endothelial growth factor A) and receptor tyrosine kinases [1,2] or non-peptide tyrosine kinase inhibitors [3,4], the 5-year prognosis for most solid tumors remains reserved. The precise roles, played by different integrin subunits in various aspects of tumor progression and why some integrins appear to be especially supportive of tumor progression [9] are still not fully understood Despite these limitations, due to their pivotal roles in cancer biology, integrins represent attractive therapeutic targets. Due to their pivotal roles in cancer biology, integrins represent attractive therapeutic targets It doesn’t seem to be essential for the formation of vasculature during development [10], nor during physiological angiogenesis associated with wound healing or tissue repair [11,12], the b3 integrin appears to be critically involved in the regulation of pathological angiogenesis [13]. We provide further evidence in support of the above therapeutic paradigm by showing that the efficient disruption by a member of the disintegrin family of multiple integrin pathways upregulated in cancer is followed by significant tumor anti-angiogenic and proapoptotic effects
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