Perspectives on Engineering Biobetter Therapeutic Proteins with Greater Stability in Inflammatory Environments

Abstract

Rapid progression of genetic engineering technology has accelerated the development and availability of protein-based biopharmaceuticals for clinical use. However, their unique and complex structures render them susceptible to a plethora of post-translational modifications (PTMs) as well as chemical degradative processes they may encounter in distinct in vivo environments. Among the more common degradative reactions that can impact the structure and function of therapeutic proteins are oxidation, proteolysis, phosphorylation, and deamidation. Several PTMs such as oxidation further render modified proteins vulnerable to aggregation or proteolytic degradation by enzymatic or non-enzymatic mechanisms (Torosantucci et al., Pharm Res 31(3):541–553, 2014). Degradation of a therapeutic protein in vivo becomes problematic when the structural modification alters its intended function and safety or efficacy profile (Foye, Lippincott Williams & Wilkins, Philadelphia, 2008). As a result of protein aggregation or degradation, therapeutic activity can be decreased, increased, or altered to have off-target effects. Protein degradation or aggregation could be facilitated under inflammatory circumstances in diverse clinical settings such as cancer, chronic inflammatory diseases, organ transplants, infectious diseases, and cardiovascular disorders, and can exacerbate an inflammatory response with unintended consequences (Chennamsetty et al., Proc Natl Acad Sci U S A 106(29):11937–11942, 2009; Hermeling et al., Pharm Res, 21(6):897–903, 2004). Therefore, characterizing and controlling the degradation or aggregation profiles for a therapeutic protein, especially for indications where the physiological environment presents additional opportunity for instability, is an essential component of a drug manufacturer’s control strategy. Evaluating a manufacturer’s control strategy is a key risk assessment tool for the regulation of investigational and licensed biologic drugs for human use. Better risk assessment can result from (1) greater characterization of critical structural modifications that can influence therapeutic protein function (2), application of sensitive and suitable methods to objectively measure protein modifications, and (3) use of relevant preclinical models or human tissue samples for predicting the in vivo impact of the inflammatory environment on such protein alterations. The increasing number of novel investigational drugs and the simultaneous demand for safer and more effective drugs warrants the need to examine the mechanisms by which therapeutic proteins are modified in vitro and in vivo, as well as apply modern analytical and genetic engineering techniques to design biobetter biologic drugs with improved safety and efficacy profiles. This chapter will examine factors known to alter the stability of therapeutic proteins in vivo, potential interactions of susceptible proteins with the inflammatory environment, and review some challenges and potential strategies for designing biobetters.