Expression of a Glyco-Optimized Anti-CD20 Antibody in the Aquatic Plant Lemna with Enhanced ADCC Activity.

Abstract

Monoclonal antibodies (mAbs) are one of the fastest growing therapeutics in the drug sector. For many antibodies, the structure and extent of the N-glycans on the Fc region of the H-chain plays a significant role in their therapeutic function. A glyco-optimized anti-CD20 antibody (rituximab) was expressed in the clonal aquatic plant Lemna. The optimized glycosylation was accomplished by co-expressing an interfering RNA (RNAi) construct targeting the endogenous alpha-1,3-fucoslytransferase (FucT) and beta-1,2-xylosyltransferase (XylT) genes (Cox et al., 2006). The resulting glyco-optimized rituximab contained a single major G0 N-glycan without any detectable xylose or fucose. In cell-based functional assays, the glyco-optimized rituximab showed similar CD20-binding affinity and apoptotic effects as Rituxan® produced in mammalian cells but with significantly enhanced (up to 100-fold) antibody-dependent cellular cytotoxicity (ADCC). Using FACS based methods, comparable CD20-binding was demonstrated in the B-cell lines Raji, Daudi and Wil2S. Apoptotic measurements used Raji and Daudi cells. In the treatment of non-Hodgkin's B-cell lymphoma (NHL), the patient response rate for Rituxan is only 50–60% and is significantly correlated with a FcgRIIIa receptor polymorphism (Cartron et al., 2002). However, the glyco-optimized rituximab showed enhanced ADCC with effector cells from all FcgRIIIa-158 genotypes. ADCC activity was determined by FACS-based methods using Raji as the target cells. PBMC genotyping was accomplished by PCR analysis (Koene et al., 1997). Up to a ten-fold decrease in complement dependent cytotoxicity (CDC) was also observed using Raji cells. This functional profile may offer the potential for an optimized anti-CD20 antibody therapeutic with improved efficacy and potency while simultaneously decreasing the side effects associated with CDC activity (Clark and Ledbetter, 2005).