In vivo phage display: identification of organ-specific peptides using deep sequencing and differential profiling across tissues

Abstract

In vivo phage display is widely used for identification of organ- or disease-specific homing peptides. However, the current in vivo phage biopanning approaches fail to assess biodistribution of specific peptide phages across tissues during the screen, thus necessitating laborious and time-consuming post-screening validation studies on individual peptide phages. Here, we adopted bioinformatics tools used for RNA sequencing for analysis of high-throughput sequencing (HTS) data to estimate the representation of individual peptides during biopanning in vivo. The data from in vivo phage screen were analyzed using differential binding—relative representation of each peptide in the target organ versus in a panel of control organs. Application of this approach in a model study using low-diversity peptide T7 phage library with spiked-in brain homing phage demonstrated brain-specific differential binding of brain homing phage and resulted in identification of novel lung- and brain-specific homing peptides. Our study provides a broadly applicable approach to streamline in vivo peptide phage biopanning and to increase its reproducibility and success rate.

Cationic Liposomes as Vectors for Nucleic Acid and Hydrophobic Drug Therapeutics

“…The in vivo phage display approach has yielded a variety of homing peptides specific for normal organs as well as for tumor vasculature and tumor cells [ 198 , 201 , 202 , 203 , 204 , 205 , 206 , 207 , 208 , 209 , 210 ]. Recent advances in the approach, most notably the application of high-throughput sequencing and development of bioinformatics tools to analyze the resulting data, allow for streamlined, robust, and reproducible identification of homing peptides [ 211 , 212 ] and mapping of the vascular diversity.…”

Exposure to Bacteriophages T4 and M13 Increases Integrin Gene Expression and Impairs Migration of Human PC-3 Prostate Cancer Cells

“…Bacteriophages for targeting cancer cells, both in vitro and in vivo, has emerged as a promising tool since Kantoch and Mordarski demonstrated phage binding to cancer cells [38]. Furthermore, the molecular heterogeneity of the vascular endothelium was demonstrated, as well as the fact that organ targeting is possible using phage peptides [39][40][41][42], which has considerably affected current cancer research and treatment [43,44]. Moreover, modified bacteriophages, including engineered phages and hybrid phages, are considered tremendously important for cancer cell targeting and drug delivery, as reported by independent research groups [45][46][47][48].…”

Circulation of Fluorescently Labelled Phage in a Murine Model

“…The bioavailability of phages in the living system is shaped by many factors. Some of them depend on phage characteristics e.g., phage capsid morphology, stability [5][6][7][8]; however, in the engineered phages, the multivalent architecture of landscape phages displayed proteins [9,10] that may affect the biodistribution of phage clones able to specifically recognise non-bacterial targets, such as neoplastic ones. Other factors affecting the phage fate in the living stystem are related to the application, i.e., route of administration, timespan, dose [11,12].…”