Next generation of anti-PD-L1 Atezolizumab with enhanced anti-tumor efficacy in vivo

Maohua Li,Wenzhi Tian,Hunter Sun,Tong Shen,Yingzi Li,Rongqing Zhao,Chenxi Xia,Jianxin Chen,Le Sun,Song Li,Xudong Liu,Wenlin Ren

doi:10.1038/s41598-021-85329-9

Maohua Li, Wenzhi Tian + Show 10 more

Open Access

https://doi.org/10.1038/s41598-021-85329-9

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Abstract

FDA-approved anti-PD-L1 antibody drug Atezolizumab is a human IgG1 without glycosylation by an N297A mutation. Aglycosylation of IgG1 has been used to completely remove the unwanted Fc-mediated functions such as antibody-dependent cytotoxicity (ADCC). However, aglycosylated Atezolizumab is unstable and easy to form aggregates. Here, we report the development of the anti-PD-L1 antibody drug Maxatezo, a glycosylated version of Atezolizumab, with no ADCC activity, better thermo-stability, and significantly improved anti-tumor activity in vivo. Using Atezolizumab as the starting template, we back-mutated A297N to re-install the glycosylation, and inserted a short, flexible amino acid sequence (GGGS) between G237 and G238 in the hinge region of the IgG1 heavy chain. Our data shows that insertion of GGGS, does not alter the anti-PD-L1′s affinity and inhibitory activity, while completely abolishing ADCC activity. Maxatezo has a similar glycosylation profile and expression level (up to 5.4 g/L) as any normal human IgG1. Most importantly, Maxatezo’s thermal stability is much better than Atezolizumab, as evidenced by dramatic increases of Tm1 from 63.55 °C to 71.01 °C and Tagg from 60.7 °C to 71.2 °C. Furthermore, the levels of ADA in mice treated with Maxatezo were significantly lower compared with animals treated with Atezolizumab. Most importantly, at the same dose (10 mg/kg), the tumor growth inhibition rate of Maxatezo was 98%, compared to 68% for Atezolizumab.

Highlights

In recent years, with greater understanding of the mechanism of tumor immune escape and the tumor microenvironment, immune-checkpoint inhibitor-based therapies have shown satisfactory clinical efficacy in a variety of tumors
Therapeutic antibodies have different mechanisms of actions, including, but not limited to (1) neutralizing antibodies that block the target/pathogen’s interaction with their cellular receptors, (2) clearing antibodies that mediate action by antibody-dependent cell phagocytosis (ADCP) to remove the target/pathogen from the body, or (3) targeting antibodies via antibody-dependent cytotoxicity (ADCC) to recruit natural killer (NK) cells and
FcγRI and FcγRIIA are expressed by macrophages, and involved in ADCP function[15,16], while FcγRIIIA is expressed on NK cells and is important for ADCC function[17]

Summary

Introduction

With greater understanding of the mechanism of tumor immune escape and the tumor microenvironment, immune-checkpoint inhibitor-based therapies have shown satisfactory clinical efficacy in a variety of tumors. Quite a few Phase III clinical tries of Atezolizumab did not reach the clinical end point (in 2017, IMvigor[211] for advanced bladder cancer failed in clinical Phase III; in 2018, IMblaze[370] for colorectal cancer failed in clinical Phase III; in 2019, IMspire[170] for melanoma failed in clinical Phase III)[1]. It is well-known in antibody manufacturing that incomplete glycosylation will lead to aggregations of antibodies[2], which in turn will induce strong ADA in treated patients[3]. FcγRI and FcγRIIA are expressed by macrophages, and involved in ADCP function[15,16], while FcγRIIIA is expressed on NK cells and is important for ADCC function[17]

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