New drug screening tool: combination of microdialysis and in vivo phage display for the identification of macrocyclic homing and tissue penetrating peptides for drug delivery

This screening technology is based on a combination of in vivo phage display (using macrocyclic peptide library), microdialysis and next-generation sequencing for the discovery of peptides that, besides site-specific vascular homing, promote tissue/cell penetration.

In vivo phage display was combined with microdialysis-based parenchymal recovery and high-throughput sequencing to select for peptides that, besides vascular homing, facilitate extravasation and tissue penetration. The drug screening technology has been validated: The proof-of-concept studies demonstrated remarkable selection of homing and cell-penetrating macrocyclic peptides with extravascular phage rescue using a custom-made (semi-permeable microdialysis membrane allowing T7 bacteriophage to enter the dialysate) microdialysis catheter implanted in the wound tissue. We first demonstrated in skin wounds that the method can selectively separate known homing peptides from those with additional tissue-penetrating ability. Subsequent in vivo screening of the macrocyclic peptide library (C X7 C macrocyclic peptide library) by microdialysis identified novel peptides by high-throughput sequencing (HTS) of the peptide-encoding region of the phage genomic DNA. Phage clones displaying homing peptides capable of penetrating wound tissue, selectively accumulate in the dialysate (collected by semipermeable microdialysis probe inserted in the wound), and identified new macrocyclic peptides with homing and tissue penetrating properties. We demonstrate in proof-of-concept studies that hese peptides home and extravasate to extravascular granulation tissue in vascularized and diabetic wounds and cross the blood-brain (-retina) barrier in retinopathy.