Abstract C180: A novel bispecific platform for potent redirected killing of B cell lymphoma
Despite significant advances in the treatment of B cell lymphoma, in many cases the disease remains untreatable or displays recurrence after initial response to therapy. The recent clinical success of bi specific molecules capable of redirecting T lymphocyte mediated cytotoxicity (Bi specific T cell Engagers, BiTEs) against B cell lymphoma provides a feasible therapeutic option. We have developed an alternate bi specific platform of Dual Affinity Re Targeting (DART) molecules that is well suited for clinical applications. Compared to BiTEs, which comprise a pair of single chain Fv domains in a single polypeptide, DARTs contain two separate chains associated into a heterodimeric, covalently linked, 'diabody' structure. We show here that DARTs are very stable in vitro and can be built to recruit and retarget different effector cells, including T cells or CD16 (Fc RIII) positive cells, against B cell lineage targets. A CD3xCD19 DART capable of recruiting cytotoxic T cells against CD19 positive targets mediated extremely potent killing of human peripheral blood B lymphocytes or B cell lymphoma lines in vitro, with an EC50 that compares favorably to that of a BiTE with identical Fv sequences. Redirected killing of Daudi, Raji, or Nalm6 lymphoma lines was observed in the picomolar range using either freshly purified T cells or peripheral blood mononuclear cells (PBMC) as effector populations. In contrast, no killing was observed against a CD19 negative cell line (RPMI 8226) with either resting or stimulated effector populations, indicating a requirement for CD19 engagement for T cell mediated killing by the CD3xCD19 DART molecule. Furthermore, a DART constructed with a humanized version of a pan anti TCR beta antibody (TCRxCD19 DART) was as efficient against CD19 positive targets as the CD3 based DART. A CD16xCD32B DART was also constructed to redirect CD16 positive NK and/or mononuclear phagocyte effector cells against CD32B positive B cells, displaying potent killing against Daudi, Raji or RPMI 8226 cells but not against a CD32B negative cell line (Ramos) Both TCR and CD16 based DARTs show potent activity in vivo in several mouse models, including autologous B cell depletion in huCD32B transgenic mice, tumor reduction in human Raji Burkitt9s lymphoma cell xenografts and Raji cell implants in NOD/SCID mice reconstituted with human PBMC. In conclusion, DARTs represent a robust and flexible platform to recruit different cell populations via various cell surface targets for re directed killing applications against B cell malignancies. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C180.