Novel immunotherapies using optimized tumor cell-penetrating antibodies - ABSTRACT Using recombinant antibodies targeting common oncogenic mutations, we have recently demonstrated that dimeric IgA penetrates human epithelial cancer cells through PIGR-dependent transcytosis, neutralizing mutated oncodrivers and expelling them outside the tumor cell. Accordingly, targeting KRASG12D with dimeric IgA abrogated the growth of different carcinomas. Moving forward, our primary objective is to translate tumor cell-penetrating Abs into the clinic. However, challenges arise due to the low yield associated with dimeric IgA production and the comparatively extended half-life of IgG antibodies. To expedite clinical translation and maximize therapeutic effectiveness, we have engineered new IgG4 antibodies incorporating a PIGR-binding peptide at the C-terminus of the Fc domain. PIGR-binding IgGs penetrate cancer cells and elicit Ag-specific control of tumor growth. Our central hypothesis is that our mutant KRAS-specific recombinant IgGs control KRAS-driven carcinomas that became resistant to small molecule KRAS inhibitors, with longer half-lives and production yields than recombinant dIgA, while making tumor cells sensitive to T cell-mediated killing. This hypothesis will be tested in these Aims: In Specific Aim 1, we will elucidate the mechanisms governing the effectiveness of our new PIGR-binding mutation-specific IgGs. Besides understanding how these Abs neutralize antigenic mutations inside tumor cells, we will select a candidate for clinical development, based on effectiveness, the capacity to target multiple mutations, and production yields. In Specific Aim 2, we will define how transcytosis of PIGR-binding IgG impacts T cell- dependent anti-tumor immunity. We will define mechanistically-driven and safe interventions that synergize with PIGR-binding Abs, possibly including immune checkpoint inhibitors. In Specific Aim 3, we will determine whether tumor cell-penetrating Abs overcome resistance to inhibitors across various mutations. We will establish a range of human cancers sensitive to our new immunotherapy, including tumors that became resistant to inhibitors. These studies will not only bridge existing knowledge gaps regarding transcytosing antibodies, but also pave the way for a new generation of immunotherapies that effectively combat aggressive human cancers driven by mutational hotspots, and perhaps other cytosolic proteins.
