Anti-angiogenic cancer therapies possess immune-stimulatory properties by counteracting pro-angiogenic molecular mechanisms. We report that tumor endothelial cells ubiquitously overexpress and secrete the intermediate filament protein vimentin through type III unconventional secretion mechanisms. Extracellular vimentin is pro-angiogenic and functionally mimics VEGF action, while concomitantly acting as inhibitor of leukocyte-endothelial interactions. Antibody targeting of extracellular vimentin shows inhibition of angiogenesis in vitro and in vivo. Effective and safe inhibition of angiogenesis and tumor growth in several preclinical and clinical studies is demonstrated using a vaccination strategy against extracellular vimentin. Targeting vimentin induces a pro-inflammatory condition in the tumor, exemplified by induction of the endothelial adhesion molecule ICAM1, suppression of PD-L1, and altered immune cell profiles. Our findings show that extracellular vimentin contributes to immune suppression and functions as a vascular immune checkpoint molecule. Targeting of extracellular vimentin presents therefore an anti-angiogenic immunotherapy strategy against cancer.

Linking decyl-triphenyl-phosphonium to fluorescein yields a fluorescent probe that accumulates in energized mitochondria, facilitates proton transfer across membranes and stimulates mitochondrial respiration. This features a mitochondria-targeted uncoupler, being of potential interest for therapeutic use against oxidative stress-related diseases.

Chemokines are a group of chemotaxis proteins that regulate cell trafficking and play important roles in immune responses and inflammation. Ticks are blood-sucking parasites that secrete numerous immune-modulatory agents in their saliva to evade host immune responses. Evasin-3 is a small salivary protein that belongs to a class of chemokine-binding proteins isolated from the brown dog tick, Rhipicephalus sanguineus . Evasin-3 has been shown to have a high affinity for chemokines CXCL1 and CXCL8 and to diminish inflammation in mice. In the present study, solution NMR spectroscopy was used to investigate the structure of Evasin-3 and its CXCL8–Evasin-3 complex. Evasin-3 is found to disrupt the glycosaminoglycan-binding site of CXCL8 and inhibit the interaction of CXCL8 with CXCR2. Structural data were used to design two novel CXCL8-binding peptides. The linear tEv3 17–56 and cyclic tcEv3 16–56 dPG Evasin-3 variants were chemically synthesized by solid-phase peptide synthesis. The affinity of these newly synthesized variants to CXCL8 was measured by surface plasmon resonance biosensor analysis. The K d values of tEv3 17–56 and tcEv3 16–56 dPG were 27 and 13 n m , respectively. Both compounds effectively inhibited CXCL8-induced migration of polymorphonuclear neutrophils. The present results suggest utility of synthetic Evasin-3 variants as scaffolds for designing and fine-tuning new chemokine-binding agents that suppress immune responses and inflammation.

Ticks, as blood sucking parasites, have developed a complex strategy to evade and suppress host immune responses during feeding. The crucial part of this strategy is expression of a broad family of salivary proteins, called Evasins, to neutralize chemokines responsible for cell trafficking and recruitment. However, structural information about Evasins is still scarce and little is known about the structural determinants of their binding mechanism to chemokines. Here, we studied the structurally uncharacterized Evasin-4, which neutralizes a broad range of CC-motif chemokines, including the chemokine CC-motif ligand 5 (CCL5) involved in atherogenesis. Crystal structures of Evasin-4 and E66S CCL5—an obligatory dimeric variant of CCL5—were determined to a resolution of 1.3-1.8 Å. The Evasin-4 crystal structure revealed an L-shaped architecture formed by an N- and C-terminal subdomain consisting of eight β-strands and an α-helix that adopts a substantially different position compared to closely related Evasin-1. Further investigation into E66S CCL5/Evasin-4 complex formation with NMR spectroscopy showed that residues of the N-terminus are involved in binding to CCL5. The peptide derived from the N-terminal region of Evasin-4 possessed nM affinity to CCL5 and inhibited CCL5 activity in monocyte migration assays. This suggests that Evasin-4 derivatives could be used as starting point for the development of anti-inflammatory drugs.

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