Replacing Antibodies: Engineering New Binding Proteins

Abstract

Nature's reliance on proteins to carry out nearly all biological processes has led to the evolution of biomolecules that exhibit a seemingly endless range of functions. Much research has been devoted toward advancing this process in the laboratory in order to create new proteins with improved or unique capabilities. The protein-engineering field has rapidly evolved from pioneering studies in engineering protein stability and activity to an application-driven powerhouse on the forefront of emerging technologies in biomedical engineering and biotechnology. A classic protein-engineering technique in the medical field has focused on manipulating antibodies and antibody fragments for various applications. New classes of alternative scaffolds have recently challenged this paradigm, and these structures have been successfully engineered for applications including targeted cancer therapy, regulated drug delivery, in vivo imaging, and a host of others. This review aims to capture recent advances in the engineering of nonimmunoglobulin scaffolds as well as some of the applications for these molecular recognition elements in the biomedical field.

Targeted Drug Delivery with an Integrin-Binding Knottin–Fc–MMAF Conjugate Produced by Cell-Free Protein Synthesis

“…The 2.5F-Fc-MMAF protein is approximately 60 kDa in size, about 60% smaller than a traditional ADC, and thus presents a new tool to explore the effects of protein size on tumor penetration in future studies (61). Our work also highlights the potential for development of other tumor targeting drug conjugates based on so-called "alternative scaffolds," including, but not limited to anticalins, affibodies, fibronectin domains, and designed ankyrin repeat proteins (62,63). In this way, combining the array of available tumor-targeting proteins with emerging linker, warhead, and conjugation technologies provides numerous opportunities for the development of next-generation cancer therapeutics.…”

Genetically modified proteins: functional improvement and chimeragenesis

“…43,83 For-example, the Bacillus subtilis chimeric bifunctional enzyme laccase/β-1,3-1,4-glucanase that catalyzes the hydrolysis of both plant cell wall β-glucans and oxidation of aromatic compounds was created by an insertion fusion of the bglS and cotA genes (Table 1). 83 The hydrolytic activity of the chimera was 20% higher against 87,88,89,90 However, the C-terminal end of luciferases is a part of the active site and thus tagging an epitope to it invades the optical intensities for the broad use in bioassays. The problem of the limited functionality of the hybrid epitope was solved by creating a copepod luciferase Aluc30 of Renilla reniformis, embedding epitopes in the middle of the N-terminal region with a high optical intensity (Table 1).…”

Peptide Aptamers: Development and Applications

“…This fast growing class of antibody mimetics is finding its way into the field of biomedical and bioanalytical applications dominated by antibodies and nucleic acids aptamers. Most of the efforts directed at developing alternative scaffold designs are concentrated in the area of therapeutic and diagnostic applications [15, 63, 222, 223]. Below are a few examples.…”

“…Affibodies are considered to be the protein scaffold design most fit for in vivo imaging [223]. Derived from protein A, they tend to generate an immune response after being repeatedly administered to patients, making them poor therapeutic agents.…”