Generation of a canine anti-canine CD20 antibody for canine lymphoma treatment

Abstract

Lymphoma is the most common hematological cancer in dogs. Canine diffuse large B cell lymphoma shows a relatively good response to treatment with multi-agent cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) chemotherapy; however, the 2-year survival rate is as low as 20%. For human B cell type lymphoma, the anti-CD20 chimeric antibody, rituximab, was developed two decades ago. The combination of rituximab and CHOP chemotherapy was highly successful in improving patient prognosis. However, no anti-canine CD20 antibody is available for the treatment of canine lymphoma. During this study, a rat anti-canine CD20 monoclonal antibody was established. We also generated a rat-canine chimeric antibody against canine CD20 designed for clinical application. This chimeric antibody (4E1-7-B) showed in vitro antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against the canine B cell lymphoma cell line CLBL-1. Moreover, to obtain stronger ADCC activity, a defucosylated 4E1-7-B antibody (4E1-7-B_f) was also generated, and it showed tenfold stronger ADCC activity compared with 4E1-7-B. 4E1-7-B_f as well as 4E1-7-B suppressed the growth of CLBL-1 tumors in an immunodeficient xenotransplant mouse model. Finally, a single administration of 4E1-7-B_f induced considerable peripheral B cell depletion in healthy beagles. Thus, 4E1-7-B_f is a good antibody drug candidate for canine B cell type lymphoma. Lymphoma is one of the most common hematological cancers in dogs, and is important as a point of comparison for human lymphoma 1. Canine lymphomas are classified into T cell and B cell types according to their cell of origin. B cell lymphoma, the most common type which is similar to human diffused large B cell lymphoma (DLBCL), has a favorable prognosis compared to T cell lymphoma because the cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) regimen of chemotherapy leads to remission in most cases. However, the 2-year survival rate is only 20%, and most cases recur after remission, finally proving refractory to chemotherapy 2. Therefore, a novel therapy is required. Recently, several novel treatment strategies for canine lymphoma, including low molecular weight inhibitors, have been established 3 , leading to the expectation of an improved prognosis for dogs with lymphoma. Nevertheless, antibody treatment for canine lymphoma has not yet been established. Among antibody therapies for cancer, rituximab, an anti-human CD20 antibody, is the oldest and most widely distributed. Rituximab is a chimeric antibody in which the constant region of the original anti-human CD20 mouse antibody was replaced with a human constant region 4. Rituximab binds to human CD20 molecules and directly induces apoptotic cell death in addition to its function through antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) 5. It has been used alongside multiple drug CHOP chemotherapy (R-CHOP) in the treatment of B cell type lymphoid tumors, includin...

Immunotherapy for Dogs: Still Running Behind Humans

“…Elanco’s anti-CD20 mAb (1E4) has shown to deplete B-cells in healthy Beagles with an in vivo half-life of about 2 weeks, but the mAb is still undergoing clinical efficacy testing ( 32 ). Recently a chimeric (rat) anti-canine CD20 mAb (defucosylated) has been reported ( 33 ) which significantly depleted B-cells in the blood of beagles (n=8) lasting for about 4 weeks. Clinical data are still pending.…”

Immunotherapeutic Strategies for Canine Lymphoma: Changing the Odds Against Non-Hodgkin Lymphoma

“…Furthermore, its defucosylation resulted in a 10-fold higher ADCC activity. The in vivo antitumor activity of this improved mAb version was assessed, revealing a tumoral growth inhibition in a cNHL xenograft mouse model and a peripheral B cell depletion in healthy beagles ( 70 ). Finally, AT-005 ( Aratana Therapeutics ), a caninized mAb targeting CD52 on T cells, has obtained conditional USDA approval for the treatment of T-cell lymphoma and is currently being evaluated in clinical trials ( 62 ).…”

Development of a monoclonal antibody for the detection of anti-canine CD20 chimeric antigen receptor expression on canine CD20 chimeric antigen receptor-transduced T cells

“…However, the clinical efficacy of these antibodies in patient dogs remain unknown. Our laboratory recently developed the novel rat anti-canine CD20 antibody, 4E1-7 [ 12 ]. Based on 4E1-7 antibody, the chimeric anti-canine CD20 antibody, 4E1-7-B, and the chimeric defucosylated anti-canine CD20 antibody, 4E1-7-B_f, were generated, and the latter was found to have dramatically high in vitro antitumor activity [ 12 ], and clinical trials for dogs with B cell lymphoma are under preparation.…”

“…Our laboratory recently developed the novel rat anti-canine CD20 antibody, 4E1-7 [ 12 ]. Based on 4E1-7 antibody, the chimeric anti-canine CD20 antibody, 4E1-7-B, and the chimeric defucosylated anti-canine CD20 antibody, 4E1-7-B_f, were generated, and the latter was found to have dramatically high in vitro antitumor activity [ 12 ], and clinical trials for dogs with B cell lymphoma are under preparation. In this therapy, pharmacokinetic studies are essential to determine individual variability and understand the relationships between antibody concentration and clinical efficacy.…”