Generation and Optimization of a Chimeric Antigen Receptor Against CD22: A New Immunotherapeutic Agent for Treating B Lineage Leukemia and Lymphoma

Abstract 4313

CD22 is expressed on a number of hematologic malignancies. A recombinant immunotoxin composed of an anti-CD22 Fv fused to a 38 kDa fragment of Pseudomonas exotoxin A (Moxetumomab pasudodtox, HA22) is currently being evaluated in the treatment of B cell malignancies. We used the Fv sequence to construct a series of chimeric antigen receptors (CARs) to determine the optimal affinity, domain structure, and signaling required for optimal anti-leukemic activity. The original anti-CD22 binding domain, BL22, or a newer generation high-affinity domain, HA22, were fused to transmembrane and signaling sequences derived from the TCR zeta-chain, CD28, and CD137. In some constructs we extended the binding domain away from the membrane using constant domains from IgG (CH2CH3). Retroviral gene vectors were used to transduce activated primary T cells with CAR constructs. We found: HA22 Fv, 2 as opposed to 3 signaling motifs, and non-CH2CH3 containing CARs were superior in short-term in vitro tumor cell cytolysis assays, indicating that Fv affinity, signaling, and 3-D structure of the CAR all impact the anti-leukemic effectiveness of CARs. Finally, direct comparison to CD19-specific CAR showed an equivalent or superior killing activity in 2 out of 3 ALL lines tested. Using Scatchard analysis to define the number of CD22 moleules on the surface, the ALL line with the lowest number of CD22 molecules is also the least able to be killed by CAR-transduced T cells. The supeior lytic activity of CD19-CAR for this single cell line was due to preservation of high levels of CD19 at the cell surface. We are currently testing the in vivo activity of anti-CD22 CAR in an immunodeficient mouse model bearing human cell lines and primary patient-derived ALL. Our results indicate that CD22-CAR should be developed for the immunotherapy of CD22+ hematologic malignancies.