In vivo Phage Display: A promising selection strategy for the improvement of antibody targeting and drug delivery properties

Abstract

The discovery of hybridoma technology, described by Kohler and Milstein in 1975, and the resulting ability to generate monoclonal antibodies (mAbs) initiated a new era in antibody research and clinical development. However, limitations of the hybridoma technology as a routine antibody generation method in conjunction with high immunogenicity responses have led to the development of alternative approaches for the streamlined identification of most effective antibodies. Within this context, display selection technologies such as phage display, ribosome display, yeast display, bacterial display, and mammalian cell surface display have been widely promoted over the past three decades as ideal alternatives to traditional hybridoma methods. The display of antibodies on phages is probably the most widespread and powerful of these methods and, since its invention in late 1980s, significant technological advancements in the design, construction, and selection of antibody libraries have been made, and several fully human antibodies generated by phage display are currently approved or in various clinical development stages. With evolving novel disease targets and the emerging of a new generation of therapeutic antibodies, such as bispecific antibodies, antibody drug conjugates (ADCs), and chimeric antigen receptor T (CAR-T) cell therapies, it is clear that phage display is expected to continue to play a central role in antibody development. Nevertheless, for non-standard and more demanding cases aiming to generate best-in-class therapeutic antibodies against challenging targets and unmet medical needs, in vivo phage display selections by which phage libraries are directly injected into animals or humans for isolating and identifying the phages bound to specific tissues offer an advantage over conventional in vitro phage display screening procedures. Thus, in the present review, we will first summarize a general overview of the antibody therapeutic market, the different types of antibody fragments, and novel engineered variants that have already been explored. Then, we will discuss the state-of-the-art of in vivo phage display methodologies as a promising emerging selection strategy for improvement antibody targeting and drug delivery properties.

Generation and Next-Generation Sequencing-Based Characterization of a Large Human Combinatorial Antibody Library

“…Further, the mAbs derived from animals also have a higher immunogenicity risk, a major hurdle in the development of therapeutic antibodies [ 6 ]. To overcome this hurdle, a human naïve or synthetic antibody library has been constructed to isolate target-specific human antibodies [ 43 , 44 , 45 ]. Although vaccination in humanized mouse models is also used as another reliable method for the generation of human mAbs, based on our previous work and similar reports by others, phage display-based antibody selection from an established antibody library can considered a faster and more efficient approach to isolating human recombinant mAbs than the antibody selection from the immune library [ 8 , 46 , 47 ].…”

Rabbit derived VL single-domains as promising scaffolds to generate antibody–drug conjugates

“…This enables the reduction of the off-target tissue and protein interactions by eliminating non-specific ligands, enriching the recovery of specific ligands that specifically target the tumor. Despite its advantages, in vivo phage display selection remains relatively unexplored 44 , 45 . To date, few studies on cancer models have been reported.…”

Generation of peptides using phage display technology for cancer diagnosis and molecular imaging

“…This powerful selection technique tends to simulate ideal living circumstances, improving the identification of specific ligands against challenging receptors in their native conformation [ 14 ]. In vivo screening offers advantages over conventional in vitro and in situ screening methods where the phage-displayed peptide candidates with desired features, including specificity, pharmacokinetics and stability can be selected in the complicated milieu of living animals [ 28 ]. Pasqualini and Ruoslahti were the first to demonstrate this technique in 1996, involving the identification of peptides homing to the vascular endothelium of renal and cerebral tissues in vivo [ 29 ].…”

“…The phages are allowed to circulate for a few hours, promoting the binding of the displayed peptide to the specific targets that are expressed on the organ or tumour surface. Subsequently, the animals will be perfused and sacrificed to wash the unbound phages, followed by the collection of the interested organ to recover the bound phages [28]. Using this approach, various peptides such as CSSTRESAC [30], VAV3 [31], SP5-52 [32] and A54 [33] that target breast cancer cells, glioblastoma, lung tumour blood vessels and liver cancer cells, respectively, were identified in recent studies.…”