The overall goal of the proposed research is to reveal how variants and homologs of bovine pancreatic ribonuclease (RNase A) promote tumor cell death. Onconase (ONC), which is an amphibian homolog of RNase A, is now undergoing Phase III human clinical trials for the treatment of malignant mesothelioma and has completed Phase II human clinical trials for the treatment of metastatic kidney cancer, refractory breast cancer, and prostate cancer. In contrast to ONC, RNase A is not cytotoxic. In the previous funding period, variants of RNase A and its human homolog were created that, like ONC, were able to evade the ribonuclease inhibitor protein (RI) that resides in the cytosol of mammalian cells. These variants were toxic to tumor cells. This finding portends the development of a new class of cancer chemotherapeutics based on mammalian ribonucleases.
The Specific Aims of this proposal are (1) to enhance the antitumoral activity of RNase A by the disruption of """"""""hot spots"""""""" identified by computational analysis of the Rl.RNase A complex, (2) to identify new cytotoxic ribonucleases from unbiased libraries using a genetic screen, (3) to reveal fundamental information about the subcellular routing of ribonucleases by using a new class of pro-fluorophores, (4) to create a ribonuclease zymogen that unleashes its cytotoxic activity in the presence of a matdx metalloprotease, and (5) to determine the three-dimensional structure of cytotoxic variants and homologs of RNase A, and thereby enhance the understanding of their antitumoral activity Significance. The proposed research will provide a detailed biochemical understanding of the antitumoral activity of ribonucleases, and could lead to new cancer chemotherapeutics based on variants and homologs of RNase A.

National Institute of Health (NIH)
National Cancer Institute (NCI)
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Biochemistry Study Section (BIO)
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Wolpert, Mary K
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University of Wisconsin Madison
Schools of Earth Sciences/Natur
United States
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Hoang, Trish T; Tanrikulu, I Caglar; Vatland, Quinn A et al. (2018) A Human Ribonuclease Variant and ERK-Pathway Inhibitors Exhibit Highly Synergistic Toxicity for Cancer Cells. Mol Cancer Ther 17:2622-2632
Thomas, Sydney P; Hoang, Trish T; Ressler, Valerie T et al. (2018) Human angiogenin is a potent cytotoxin in the absence of ribonuclease inhibitor. RNA 24:1018-1027
Chyan, Wen; Raines, Ronald T (2018) Enzyme-Activated Fluorogenic Probes for Live-Cell and in Vivo Imaging. ACS Chem Biol 13:1810-1823
Lomax, Jo E; Eller, Chelcie H; Raines, Ronald T (2017) Comparative functional analysis of ribonuclease 1 homologs: molecular insights into evolving vertebrate physiology. Biochem J 474:2219-2233
Chyan, Wen; Kilgore, Henry R; Gold, Brian et al. (2017) Electronic and Steric Optimization of Fluorogenic Probes for Biomolecular Imaging. J Org Chem 82:4297-4304
Hoang, Trish T; Raines, Ronald T (2017) Molecular basis for the autonomous promotion of cell proliferation by angiogenin. Nucleic Acids Res 45:818-831
Hoang, Trish T; Smith, Thomas P; Raines, Ronald T (2017) A Boronic Acid Conjugate of Angiogenin that Shows ROS-Responsive Neuroprotective Activity. Angew Chem Int Ed Engl 56:2619-2622
Arnold, Ulrich; Raines, Ronald T (2016) Replacing a single atom accelerates the folding of a protein and increases its thermostability. Org Biomol Chem 14:6780-5
Thomas, Sydney P; Kim, Eunji; Kim, Jin-Soo et al. (2016) Knockout of the Ribonuclease Inhibitor Gene Leaves Human Cells Vulnerable to Secretory Ribonucleases. Biochemistry 55:6359-6362
Andersen, Kristen A; Smith, Thomas P; Lomax, Jo E et al. (2016) Boronic Acid for the Traceless Delivery of Proteins into Cells. ACS Chem Biol 11:319-23

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