Antigenic modulation limits the effector cell mechanisms employed by type I anti-CD20 monoclonal antibodies

Thomas R.W Tipton,Ali Roghanian,Robert J Oldham,Matthew J Carter,Kerry L Cox,C Ian Mockridge,Ruth R French,Lekh N Dahal,Patrick J Duriez,Philip G Hargreaves,Mark S Cragg,Stephen A Beers

doi:10.1182/blood-2014-07-588376

Abstract

AbstractFollowing the success of rituximab, 2 other anti-CD20 monoclonal antibodies (mAbs), ofatumumab and obinutuzumab, have entered clinical use. Ofatumumab has enhanced capacity for complement-dependent cytotoxicity, whereas obinutuzumab, a type II mAb, lacks the ability to redistribute into lipid rafts and is glycoengineered for augmented antibody-dependent cellular cytotoxicity (ADCC). We previously showed that type I mAbs such as rituximab have a propensity to undergo enhanced antigenic modulation compared with type II. Here we assessed the key effector mechanisms affected, comparing type I and II antibodies of various isotypes in ADCC and antibody-dependent cellular-phagocytosis (ADCP) assays. Rituximab and ofatumumab depleted both normal and leukemic human CD20-expressing B cells in the mouse less effectively than glycoengineered and wild-type forms of obinutuzumab, particularly when human immunoglobulin G1 (hIgG1) mAbs were compared. In contrast to mouse IgG2a, hIgG1 mAbs were ineffective in ADCC assays with murine natural killer cells as effectors, whereas ADCP was equivalent for mouse IgG2a and hIgG1. However, rituximab's ability to elicit both ADCC and ADCP was reduced by antigenic modulation, whereas type II antibodies remained unaffected. These data demonstrate that ADCP and ADCC are impaired by antigenic modulation and that ADCP is the main effector function employed in vivo.

Highlights

Rituximab is the archetypal anti-CD20 monoclonal antibody, licensed in 1997 and used alongside chemotherapy to treat nonHodgkin lymphoma.[1]
We show that modulation affects both Fc g receptor (FcgR)-dependent effector mechanisms engaged by monoclonal antibody (mAb) antibody-dependent cellular cytotoxicity (ADCC) and antibodydependent cellular phagocytosis (ADCP), explaining the propensity for type II antibodies to outperform type I in vivo in both normal B-cell depletion and a model of chronic lymphocytic leukemia (CLL)
Based on the observation that hIgG1 was efficacious in depleting hCD20 B cells in vivo but showed little ADCC activity because of its minimal binding to mFcgRIII, we demonstrate that ADCP is the dominant effector mechanism responsible for target cell depletion in the mouse

Summary

Introduction

Rituximab is the archetypal anti-CD20 monoclonal antibody (mAb), licensed in 1997 and used alongside chemotherapy to treat nonHodgkin lymphoma.[1] It is proposed to operate through 4 effector functions: programmed cell death, complement-dependent cytotoxicity (CDC), and the 2 Fc g receptor (FcgR)-dependent mechanisms antibody-dependent cellular cytotoxicity (ADCC) and antibodydependent cellular phagocytosis (ADCP).[1,2,3]. Following on from the success of rituximab are the next-generation anti-CD20 mAbs ofatumumab and obinutuzumab. Ofatumumab was approved for chronic lymphocytic leukemia (CLL) treatment in 2009 and shows enhanced CDC, likely through its low off-rate and cell-surface proximal epitope.[4,5] Obinutuzumab, licensed in 2013 for first-line CLL treatment (in combination with chlorambucil), has a glycoengineered Fc region, resulting in higher affinity binding to human FcgRIIIa (hFcgRIIIa) and b, enhancing hFcgRIIIdependent effector functions.[6,7,8,9]. We have previously demonstrated that antigenic modulation, whereby CD20 antibody:antigen complexes are internalized after type I mAb binding, will contribute to CD20 loss and that this process is accelerated by hFcgRIIb.[11,12,13]

Methods

Results

Conclusion