Development and Potent Anti-Tumor Activity of a Fully Humanized Anti-TAG-72-IL-2 Fusion Protein for Therapy of Solid Tumors

<b>Background/Objectives:</b> Immunocytokines (ICKs) are antibody-cytokine fusion proteins that deliver cytokines directly to tumors to increase immune responses, which are otherwise absent or limited by the delivery of antibodies alone. Tumor-associated glycoprotein-72 (TAG72) is overexpressed in numerous solid tumors. <b>Methods:</b> An anti-TAG72-IL-2 fusion protein was expressed in mammalian cells and tested in vitro for binding and bioactivity, and in vivo in two models. <b>Results:</b> In vitro studies showed high antigen specificity against TAG-72-positive tumor cell lines and IL-2 activity in CD25 (IL-2R)-positive reporter cells. To study the anti-tumor activity of huCC49-IL-2 in an immunocompetent model, the TAG-72 expression was established in murine mammary and colorectal cells by transfection with murine st6galnac-I gene (mSTn). Four daily doses of anti-TAG72-IL-2 monotherapy for TAG-72-expressing orthotopic murine mammary tumors in immunocompetent mice resulted in minimal whole-body toxicity and significant tumor growth reduction mediated by tumor infiltration of IFNγ⁺ CD8⁺ T cells. When mammary tumors were pretreated with image-guided fractionated radiation therapy (IGRT) followed by anti-TAG72-IL-2 therapy, an improved tumor growth inhibition was observed along with an increased tumor infiltration of IFNγ⁺ CD8⁺ T cells and a significant reduction in Foxp3⁺ CD4⁺ cells. Anti-TAG72-IL-2 monotherapy in TAG-72⁺ colorectal tumors led to a significant tumor reduction but also cures (4/7), with a rejection of rechallenges with both TAG-72-positive and -negative MC38 cells, thus demonstrating evidence of immune memory and antigen spreading. <b>Conclusions:</b> antiTAG-72-IL-2 therapy showed strong anti-tumor effects driven by activated CD8⁺ T cells making it a promising approach to the treatment of TAG-72⁺ tumors.