Anti-TNF Alpha Antibody Humira with pH-dependent Binding Characteristics: A constant-pH Molecular Dynamics, Gaussian Accelerated Molecular Dynamics, and In Vitro Study

Abstract

Humira is a monoclonal antibody that binds to TNF alpha, inactivates TNF alpha receptors, and inhibits inflammation. Neonatal Fc receptors can mediate the transcytosis of Humira–TNF alpha complex structures and process them toward degradation pathways, which reduces the therapeutic effect of Humira. Allowing the Humira–TNF alpha complex structures to dissociate to Humira and soluble TNF alpha in the early endosome to enable Humira recycling is crucial. We used the cytoplasmic pH (7.4), the early endosomal pH (6.0), and pKa of histidine side chains (6.0–6.4) to mutate the residues of complementarity-determining regions with histidine. Our engineered Humira (W1-Humira) can bind to TNF alpha in plasma at neutral pH and dissociate from the TNF alpha in the endosome at acidic pH. We used the constant-pH molecular dynamics, Gaussian accelerated molecular dynamics, two-dimensional potential mean force profiles, and in vitro methods to investigate the characteristics of W1-Humira. Our results revealed that the proposed Humira can bind TNF alpha with pH-dependent affinity in vitro. The W1-Humira was weaker than wild-type Humira at neutral pH in vitro, and our prediction results were close to the in vitro results. Furthermore, our approach displayed a high accuracy in antibody pH-dependent binding characteristics prediction, which may facilitate antibody drug design. Advancements in computational methods and computing power may further aid in addressing the challenges in antibody drug design.

The Role of Structural Bioinformatics in Understanding Tumor Necrosis Factor α-Interacting Protein Mechanisms in Chronic Inflammatory Diseases: A Review

“…In turn, Hong and colleagues [134] carried out an analysis in 2021 using Adalimumab, commercially known as Humira. Taking into account that neonatal Fc receptors can mediate the transcytosis of the Humira-TNF-α complex structures and process them in degradation pathways, which reduces the therapeutic effect of Humira and allows the Humira-TNF-α complex structures to dissociate into Humira and soluble TNF-α in the early endosome to allow recycling of Humira.…”

“…Furthermore, the presented approach demonstrated high accuracy in predicting pH-dependent antibody binding characteristics, which could facilitate the design of antibody drugs. The authors highlighted that advances in computational methods and computational power could further help address challenges in antibody drug design [134].…”

A stepwise mutagenesis approach using histidine and acidic amino acid to engineer highly pH-dependent protein switches

“…These antibodies bind to overexpressed antigens, and the antibody–antigen complexes are dissociated in acidic endosomes; the antigens then undergo lysosomal degradation. Related studies have also focused on improving pharmacokinetics (Hong et al 2021 ; Traxlmayr et al 2014 ). In contrast, engineered pH-dependent antibodies that selectively bind at acidic pH (i.e., they have weakened binding affinity at physiological pH) have rarely been studied.…”