Date of Graduation

8-2014

Document Type

Dissertation (PhD)

Program Affiliation

Biomedical Sciences

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

George Calin, M.D., Ph.D.

Committee Member

Wadih Arap, M.D., Ph.D.

Committee Member

Steven Curley, M.D.

Committee Member

Mikhail Kolonin, Ph.D.

Committee Member

Renata Pasqualini, Ph.D.

Committee Member

M. James You, M.D., Ph.D.

Abstract

In vivo phage display screenings by intravenous injection of a random phage-displayed peptide library allow for the selection of peptides that localize to specific vascular beds. At the University of Texas MD Anderson Cancer Center, we have had the opportunity to perform phage display screenings in cancer patients in order to select for cancer specific targets directly in humans. These targets serve to define biochemical diversity of endothelial cell surfaces and can be validated and explored towards the design of vascular-targeted pharmacology. In the most recent patient screen, samples were recovered from hepatocellular carcinoma (HCC) as well as 26 additional tissues. High-throughput sequencing and multidimensional bioinformatics analysis of recovered peptides led to the identification of extensive panels of motifs that are predicted to distinctly localize to tissue-specific vascular beds. Utilizing peptide affinity purification and phage based binding assays, we have shown that the HCC targeting peptide (SGVGAASL) identified from this patient screen, selectively binds to HCC in vitro as well as in vivo facilitated by a receptor mediated interaction with the giantin protein. FACS and protein fractionational experiments showed that the giantin polypeptide, normally considered an intracellular protein, is uniquely expressed on the surface of HCC cell lines as well as activated endothelial cells. shRNA mediated depletion of giantin expression lead to a loss of proliferation and adhesion in cancer cells. Finally, an extensive study of giantin expression in patient HCC tissue uncovered a unique expression pattern on the surface of tumor-associated vasculature. Collectively, these data support a functional role for giantin on the surface of HCC tumor endothelium that could potentially be exploited for delivery of imaging and therapeutic agents. Ultimately, this work serves as the foundation of a high-throughput integrative platform for discovery and validation of tissue-specific motifs towards a comprehensive understanding of the vascular landscape in humans.

Keywords

hepatocellular carcinoma, phage display, angiogenesis, human vasculature, next generation sequencing

Available for download on Saturday, July 22, 2017

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