Immune Therapy for Chronic Lymphocytic Leukemia Induces the Antibody Response Against a Novel Tumor-Associated Antigen, the Orphan Tyrosine Kinase Receptor ROR1.

Transduction of Chronic lymphocytic leukemia (CLL) CLL cells with replication-defective adenovirus encoding CD154 (Ad-CD154) enhances the capacity of such cells to function as antigen presenting cells. In a clinical trial patients received multiple infusions of autologous CLL cells transduced ex vivo with Ad-CD154 to examine the safety and efficiency of Ad-CD154 gene therapy. Treated patients were found to mount antibody responses against the recombinant adenovirus and cellular immune responses against autologous CLL cells. We examined the antibody response generated in some of the treated patients against CLL cells. Pre and post treatment sera were incubated with CLL cells followed by detection with human anti-IgG antibody via flow cytometry. We found that 3 out of 6 patients examined developed IgG antibodies after therapy that could bind CLL cells. Microarray analyses of CLL samples identified a relatively small number of genes that were differentially expressed in CLL cells compared with normal B cell subsets or neoplastic B cells of other B cell malignancies. These CLL signature genes are candidates TAAs of CLL. One such a CLL signature gene encodes ROR1, a surface receptor tyrosine kinase. Surface expression of ROR1 was on CLL cells, but not on normal lymphocytes. We used two methods to detect the anti-ROR1 activity. One was the flow cytometric analysis using Chinese hamster ovary cell transfected with ROR1 cDNA. Another strategy used an ELISA with plates coated with recombinant ROR1protein. Both methods revealed that patients who developed anti-CLL autoantibodies following gene therapy also developed IgG antibodies reactive with ROR1. As such, ROR1 appears to be a candidate TAA that may be targeted by immune responses induced by Ad-CD154 gene therapy.