Granzyme B Peptide as a Non-invasive Theranostic Agent
Mentor: Umar Mahmood, M.D. (Massachusetts General Hospital)
Granzyme B is a serine-protease released by CD8+ T cells and natural killer cells during the cellular immune response, and represents one of the two dominant mechanisms by which T cells mediate cancer cell death. CD8+ T cells are the body’s main immune response to cancerous cells. Our lab has previously designed a PET imaging agent that was specific for Granzyme B. The Granzyme B specific PET imaging agent (GZP) was modified to induce irreversible binding to Granzyme B and allows for labeling with a suitable positron emitting PET radiometal. This imaging agent has been shown to have increased accumulation in responding tumors in preclinical models in which rodents have been treated with immunomodulatory therapy. In CT26 and MC38 synegenic cancer models, it was noted that GZP uptake correlates closely with tumoral Granzyme B expression. We demonstrated that GZP could be used as a predictive imaging biomarker for successful immunotherapy. Human specific Granzyme B agent was also developed in our lab. The agents bind favorably to biopsy specimen obtained from patients undergoing checkpoint inhibitor immunotherapy. Using PET imaging, it was found that Granzyme B expression from those on-treatment specimens was significantly different between patients who responded and those who did not respond to the treatment. It was thus concluded that GZP represents an useful biomarker for active cytotoxic immune response. For my project, I am focused not only on the diagnostic, but also the therapeutic potential of Granzyme B peptide. The goal of this research is to exploit the Grazyme B targeting moiety that has previously shown to accumulate in tumor foci with CD8+ T cells and replace the positron-emitting imaging radionuclide with toxic beta-emitting radiometal. This proposed mechanism involves direct tumor cell killing and should improve the tumor reponse rate to immunomodulatory therapy. Tumor cells (CT26 and MC38) derived from murine colon cancer will be injected into immunocompetent mice model (Balb/c and C57BL/6J) subcutaneously. Immunotherapy (anti-PD1 and anti-CTLA-4) will be given intraperitoneally on days 3, 6 and 9 following tumor implantation. On day 12, Granzyme B peptide will be radiolabeled with 90Y and will be intraveniously injected into the mice; tumor growth rate will be monitored every other day to verify therapeutic response and body weight will be measured to evaluate any overall toxicity.
Figure 1: Granzyme B PET Imaging Differentiates Treated Responders and Non-Responders: Mice were treated with anti-PD-1 and anti-CTLA-4 therapy or vihicle were imaged 12 days post-inoculation. This image demonstrates the activity of Granzyme B in mice that reponded and did not respond to the treatment. Tumors are labeled T. Figure 2: Chemical structure of the Granzyme B peptide attached to a chelating agent NOTA. 68Ga attaches to NOTA for PET imaging purpose. 90Y will replace 68Ga as the toxic beta-emitting radiometal in this project. Source: