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CLT1 targets angiogenic endothelium through CLIC1 and fibronectin

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Abstract

Angiogenesis is important for tumor growth and metastasis. CLT1 (CGLIIQKNEC), a peptide that binds to tumor interstitial spaces in the presence of fibrin-fibronectin, has structural similarity to the anti-angiogenic β-sheet peptides anastellin and anginex. This similarity is reflected in the ability of CLT1 to form co-aggregates with fibronectin that induce an unfolded protein response and cause autophagic cell death in proliferating endothelial cells. CLT1 cytotoxicity is mediated at least in parts by a novel CLT1 binding protein, Chloride Intracellular Channel 1 (CLIC1), which promotes internalization of CLT1-fibronectin co-aggregates in a mechanism that depends on the LIIQK amino acid sequence of CLT1. LIIQK encompasses amino acid residues relevant for CLT1 binding to CLIC1 and in addition, facilitates the formation of CLT1-fibronectin co-aggregates, which in turn promote translocation of CLIC1 to the endothelial cell surface through ligation of integrin αvβ3. Paralleling the in vitro results, we found that CLT1 co-localizes with CLIC1 and fibronectin in angiogenic blood vessels in vivo, and that CLT1 treatment inhibited angiogenesis and tumor growth. Our findings show that CLT1 is a new anti-angiogenic compound, and its mechanism of action is to form co-aggregates with fibronectin, which bind to angiogenic endothelial cells through integrins, become internalized through CLIC1 and elicit a cytotoxic unfolded protein response. The simple structure and high potency of CLT1 make it a potentially useful compound for anti-angiogenic treatments.

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Acknowledgments

We thank Dr. Erkki Ruoslahti for providing anastellin expression clones as well as for helpful discussions and comments on the manuscript. We also thank Dr. Per Basse, Dr. Simon Watkins and the UPCI Imaging Facility for help with confocal microscopy. This work was supported by National Institutes of Health grants CA134330 (JP), P50 CA90386 (pilot project to JP) and P30CA047904 (UPCI CCSG), and Department of Defense grant PC073635-NIA (JP).

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The authors declare that they have no conflict of interest.

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Experiments comply with current U.S. law.

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Author notes

  1. Gunjan Malik

    Present address: Department of Pharmacology, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX, 78229, USA

  2. Brian L. Hood & Thomas P. Conrads

    Present address: Women’s Health Integrated Research Center at Inova Health System, 3289 Woodburn Road, Suite 375, Annandale, VA, 22003, USA

Authors and Affiliations

  1. Department of Urology, University of Pittsburgh School of Medicine, Shadyside Medical Center, Suite G33, 5200 Centre Avenue, Pittsburgh, PA, 15232, USA

    Lynn M. Knowles, Gunjan Malik & Jan Pilch

  2. Prostate and Urological Cancers Program, University of Pittsburgh Cancer Institute, 5150 Centre Avenue, Pittsburgh, PA, 15232, USA

    Jan Pilch

  3. Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, 204 Craft Avenue, Pittsburgh, PA, 15213, USA

    Brian L. Hood & Thomas P. Conrads

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  1. Lynn M. Knowles
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Correspondence to Jan Pilch.

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Knowles, L.M., Malik, G., Hood, B.L. et al. CLT1 targets angiogenic endothelium through CLIC1 and fibronectin. Angiogenesis 15, 115–129 (2012). https://doi.org/10.1007/s10456-011-9247-8

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