HuMab-7D8, a monoclonal antibody directed against the membrane-proximal small loop epitope of CD20 can effectively eliminate CD20low expressing tumor cells that resist rituximab-mediated lysis

Abstract

The online version of this article has a Supplementary Appendix. BackgroundIncorporation of the chimeric CD20 monoclonal antibody rituximab in the treatment schedule of patients with non-Hodgkin's lymphoma has significantly improved outcome. Despite this success, about half of the patients do not respond to treatment or suffer from a relapse and additional therapy is required. A low CD20-expression level may in part be responsible for resistance against rituximab. We therefore investigated whether the CD20-expression level related resistance to rituximab could be overcome by a new group of CD20 mAbs (HuMab-7D8 and ofatumumab) targeting a unique membrane-proximal epitope on the CD20 molecule. Design and MethodsBy retroviral transduction of the CD20 gene into CD20-negative cells and clonal selection of transduced cells a system was developed in which the CD20-expression level is the only variable. These CD20 transduced cells were used to study the impact of rituximab and HuMab-7D8 mediated complement-dependent cytotoxicity. To study the in vivo efficacy of these mAbs an in vivo imaging system was generated by retroviral expression of the luciferase gene in the CD20-positive cells. ResultsWe show that HuMab-7D8 efficiently killed CD20 low cells that are not susceptible to rituximabinduced killing in vitro. In a mouse xenograft model, we observed a comparable increase in survival time between HuMab-7D8 and rituximab-treated mice. Most significantly, however, HuMab-7D8 eradicated all CD20-expressing cells both in the periphery as well as in the bone marrow whereas after rituximab treatment CD20 low cells survived. ConclusionsCells that are insensitive to in vitro and in vivo killing by rituximab as the result of their low CD20-expression profile may be efficiently killed by an antibody against the membrane-proximal epitope on CD20. Such antibodies should, therefore, be explored to overcome rituximab resistance in the clinic. © F e r r a t a S t o r t i F o u n d a t i o n

Ofatumumab Is More Efficient than Rituximab in Lysing B Chronic Lymphocytic Leukemia Cells in Whole Blood and in Combination with Chemotherapy

“…We could confirm significant CDC even of low CD20 expressing B-CLL by OFA, in contrast to virtually no lysis of these targets by RTX. This finding is in agreement with previous data showing that RTX-resistant cells may be sensitive to OFA (13,31). Furthermore, we could show that neoplastic samples expressing intermediate to high CD20 showed relatively high lysis ranging from 50-99%.…”

Antibodies that efficiently form hexamers upon antigen binding can induce complement-dependent cytotoxicity under complement-limiting conditions

“…However, for CLL B cells with lower levels of CD20, there is a considerable drop-off for CDC mediated by IgG1-7D8 and OFA, but Hx-7D8 still promotes substantial CDC. Finally, across all of the CLL B cells examined, the CDC activities of IgG1-7D8 and OFA are quite similar, in agreement with previous reports (12,53). Dose-response tests for CDC in C9-dpl sera differentiate Hx-7D8 from wild-type complement-fixing mAbs IgG1-7D8 and W6/32…”

Impact of Epidermal Growth Factor Receptor (EGFR) Cell Surface Expression Levels on Effector Mechanisms of EGFR Antibodies

“…However, data for Fab-mediated MoAs differed between that study for trastuzumab and our data for EGFR Abs using BHK-EGFR + cell lines. Also, with regard to the therapeutic CD20 mAbs rituximab and ofatumumab, CDC and ADCC activity was demonstrated to correlate positively with CD20 cell surface expression levels (25,26).…”