A Cross Company Perspective on the Assessment of Therapeutic Protein Biotransformation

Abstract

Unlike with new chemical entities, the biotransformation of therapeutic proteins (TPs) has not been routinely investigated or included in regulatory filings. Nevertheless, there is an expanding pool of evidence suggesting that a more in-depth understanding of biotransformation could better aid the discovery and development of increasingly diverse modalities. For instance, such biotransformation analysis of TPs affords important information on molecular stability, which in turn may shed light on any potential impact on binding affinity, potency, pharmacokinetics, efficacy, safety, or bioanalysis. This perspective summarizes the current practices in studying biotransformation of TPs and related findings in the biopharmaceutical industry. Various TP case studies are discussed, and a fit-for-purpose approach is recommended when investigating their biotransformation. In addition, we provide a decision tree to guide the biotransformation characterization for selected modalities. By raising the awareness of this important topic, which remains relatively underexplored in the development of TPs (Bolleddula et al., 2022), we hope that current and developing practices can pave the way for establishing a consensus on the biotransformation assessment of TPs. SIGNIFICANCE STATEMENTThis article provides a comprehensive perspective of the current practices for exploring the biotransformation of therapeutic proteins across the drug development industry. We, the participants of the Innovation and Quality therapeutic protein absorption distribution metabolism excretion working group, recommend and summarize appropriate approaches for conducting biotransformation studies to support internal decision making based on the data generated in discovery and development.This paper received no external funding. No author has an actual or perceived conflict of interest with the contents of this paper.dx.

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top-down and middle-down mass spectrometry

“…With a growing interest in multispecifics, even more complex and diverse formats are now being designed such as CODV-Ig (Cross-over dual variable-Ig like), mAb-scFv or tandem-Nbs. Since the complexity of these new biotherapeutic constructs potentially makes them more prone to in vivo biotransformation [7][8][9], there is nowadays a crucial need to precisely characterize and quantify biotransformation products. Biotransformation of a therapeutic protein includes proteolytic cleavages and post-translational modifications (PTMs) such as deamidation, oxidation or glycation among others.…”

Translatability of In Vitro Stress for Predicting Deamidation and Oxidation Biotransformation on Biotherapeutics

“…12−15 Therefore, characterizing such liabilities is becoming an emerging need for biologics drug discovery. 9,16 Deamidation and isomerization on biomolecules can be generated in vitro using stress incubations under physiologically relevant temperature and pH conditions. Likewise, oxidation can be induced by multiple in vitro conditions including oxidizing agents (AAPH, peroxides), ultraviolet (UV) irradiation, and elevated temperatures.…”

“…Biotransformation leading to modifications on single residues, − such as deamidation, isomerization, and oxidation, can modify the primary sequences and higher-order structures of proteins. − In drug discovery and development, such in vivo degradations are generally most concerning when occurring on target binding domains such as complementarity determining regions (CDRs) for monoclonal antibodies (mAbs). Degradations occurring on other regions such as ones interacting with neonatal Fc receptor (FcRn) and Fc gamma receptors may also be critical.…”

“…Oxidation most commonly occurs on residues such as cysteine, methionine, and tryptophan (and to a lesser extent on histidine and tyrosine). The aforementioned liabilities can lead to loss of efficacy/potency, poor pharmacokinetics, and/or toxicity/immunogenicity for protein therapeutics. − Therefore, characterizing such liabilities is becoming an emerging need for biologics drug discovery. , …”

Special Section on Pharmacokinetics and ADME of Biological Therapeutics–Editorial

“…In "A Cross Company Perspective on the Assessment of Therapeutic Protein Biotransformation" (Walles et al, 2022), Walles and colleagues noted that a more in-depth understanding of biotransformation is needed for increasingly diverse modalities. The authors, who are participants of the IQ TP-ADME working group, summarized the current practices in studying TP biotransformation and related learnings in the biopharmaceutical industry.…”