Depletion of T Cell Epitopes in Lysostaphin Mitigates Anti-Drug Antibody Response and Enhances Antibacterial Efficacy In Vivo

Abstract

SUMMARY The enzyme lysostaphin possesses potent anti-staphylococcal activity and represents a promising antibacterial drug candidate; however, its immunogenicity poses a barrier to clinical translation. Here, structure-based biomolecular design enabled widespread depletion of lysostaphin’s DRB1*0401 restricted T cell epitopes, and resulting deimmunized variants exhibited striking reductions in anti-drug antibody responses upon administration to humanized HLA-transgenic mice. This reduced immunogenicity translated into improved efficacy in the form of protection against repeated challenges with methicillin-resistant Staphylococcus aureus, or MRSA. In contrast, while wild type lysostaphin was efficacious against the initial MRSA infection, it failed to clear subsequent bacterial challenges that were coincident with escalating anti-drug antibody titers. These results extend the existing deimmunization literature, in which reduced immunogenicity and retained efficacy are assessed independently of each other. By correlating in vivo efficacy with longitudinal measures of anti-drug antibody development, we provide the first direct evidence that T cell epitope depletion manifests enhanced biotherapeutic efficacy.

EpiSweep: Computationally Driven Reengineering of Therapeutic Proteins to Reduce Immunogenicity While Maintaining Function

“…The methods are illustrated with case study application to a domain from a therapeutic protein that we have been developing [24,35,25]: the cell wall binding domain (CWBD) of Staphylococcus simulans lysostaphin (LST). Lysostaphin is a potent anti-staphylococcal enzyme, effective even against drug-resistant S. aureus strains [36].…”

“…Here, we provide a step by step guide to the application of the EpiSweep suite of deimmunization algorithms, as introduced in our series of algorithmic papers [11,18,19,13,21] and prospectively applied in our series of experimental papers [12,22,14,23–25]. To assess immunogenicity, the software utilizes any pocket profile-based epitope predictor; we illustrate here with the publicly available ProPred matrices [26].…”

Computationally optimized deimmunization libraries yield highly mutated enzymes with low immunogenicity and enhanced activity

“…EpiSweep is an integrated deimmunization software suite that is generic to the epitope score, supports both sequence-and structure-based analysis of mutational effects on function, and designs either individual variants or combinatorial libraries (52). Although structure-based library designs have been described elsewhere (11,51), the results presented here demonstrate the use of a sequence score to drive the design of a combinatorial deimmunization library and demonstrate the scaling of the approach to a massive library size. In short, the sequence score is derived from a multiple sequence alignment of P99βL homologs obtained by PSI-BLAST; it includes both one-body (conservation) and two-body (coupling) terms (15).…”

“…Mutagenic deletion of T-cell epitopes has been demonstrated for a number of therapeutic proteins (2)(3)(4)(5)(6)(7), and in some cases a correlation with reduced antidrug antibody titers has been shown (8)(9)(10)(11). More recently, a direct tripartite link from elimination of MHC II-binding peptides to reduction in antidrug antibodies to improvement in therapeutic efficacy was demonstrated for a reengineered antibacterial enzyme designed to treat methicillin-resistant Staphylococcus aureus infections (11). Given that antibody humanization methods likewise reduce T-cell recognition, there is mounting evidence that T-cell epitope deletion is a clinically useful strategy for addressing biotherapeutic immunogenicity.…”

Chemically modified peptides and proteins - critical considerations for oral delivery

“…presents a potential risk of increased immunogenicity. Assessment of the immunogenicity of such a new peptide or protein is highly complicated; although considerable improvements in immunogenicity prediction algorithms have recently been achieved 30,53 and some cellular based assay are available to test immunogenicity, 50 the ultimate answer will come from the human clinical trials and clinical practice.…”