Abstract 5629: Application of a novel bispecific antibody-based scaffold for optimal redirected T-cell killing of cancer cells

Abstract

Abstract Despite significant progress, improved treatment modalities are required to eradicate cancer. Cytotoxic T lymphocytes are among the most potent cell populations capable of killing target cells and considerable effort has been dedicated to developing strategies aimed at redirecting T cells toward cancer, including the generation of bispecific molecules capable of co-engaging a tumor target together with an activating receptor on T lymphocytes. Limitations of the existing technology, however, including shortcomings in manufacturability, stability, specificity and potency, have often hampered progress in harnessing the power of T cells with bi-specific molecules. We have applied a novel antibody-based scaffold, termed DART (Dual-Affinity Re-Targeting), to redirect T-cell killing toward tumor cells in the absence of cognate interactions. DART molecules are structurally comprised of a covalently linked “diabody” generated through co-expression in mammalian cells of two separate Fv-encoding chains. To evaluate the ability of the platform to redirect T-cell killing, DARTs were designed to target hematological malignancies by comprising a B-cell lineage specificity (CD19) together with T-cell recognition (CD3 or TCR), while a DART comprising specificities for the HER2 target and the TCR was used to evaluate applicability to solid tumors. All three DART molecules assembled efficiently and were evaluated in a series of redirected T-cell killing assays to evaluate potency and mechanistic aspects of cytotoxicity. Both the CD19xCD3 and CD19xTCR DART molecules demonstrated efficient co-ligation of B-cell lymphoma cells with human T cells and mediated equivalently potent redirected T-cell killing against a panel of CD19-positive B-cell lymphoma cell lines but not CD19-ve cell lines. In similar fashion, the TCRxHER2 DART revealed potent T-cell mediated killing of a panel of HER2 expressing cell lines, including those expressing low HER2 levels (1+ by HercepTest), but was inactive against HER2-negative lines. Analyses of the mechanism of T-cell mediated killing by the DART molecules revealed a dependence on the granzyme B/perforin pathways, with potent killing observed during short-term incubation of the effector populations with target cells and at low E:T ratios. Furthermore, activation of human T cells by DART molecules proceeded strictly in a target-dependent manner. Finally, analyses in xenograft models reconstituted with human peripheral blood mononuclear cells, demonstrated efficient tumor regression by TCR-based DART molecules. In conclusion, T-cell-directed DARTs represent a robust and flexible platform to recruit T-cell populations for cancer cell killing and may offer potential application toward tumor eradication in cancer treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5629.