Identification of peptide coatings that enhance diffusive transport of nanoparticles through the tumor microenvironment

Rashmi Mohanty; Xinquan Liu; Jae You Kim; Xiujuan Peng; Sahil Bhandari; Jasmim Leal; Dhivya Arasappan; Dennis C. Wylie; Tony Dong; Debadyuti Ghosh

We used combinatorial screening (Figure 1a ) with next-generation sequencing and analysis to identify peptides that can achieve diffusive transport through the ECM network present in tumors. A cysteine constrained 9-mer random peptide-presenting T7 phage library was engineered to effectively serve as a collection of peptide-based surface chemistries with a combination of amino acids possessing varying physicochemical properties. The phage library was incubated with a transwell coated with ECM components collagen I (7.8 mg/ml) and hyaluronic acid (HA) (0.84 mg/ml), which are predominant in the tumor microenvironment of pancreatic cancers [65] [66] [67] . The charged ECM components serve as an electrostatic interaction filter that has been shown to hinder diffusion of charged molecules 15, 67 . A 1 mm thickness of tumor ECM was prepared in the insert of a 24-well transwell. The phage library was added at the top of the ECM to allow it to diffuse through the tumor ECM model at 37 ºC. After 1 hour of incubation, phage that permeated through the ECM was collected, and a fraction of the output was quantified by standard double-layer plaque assay. The remaining fraction was amplified in E. coli to make more copies needed for the subsequent round of screening. This screening was iterated for three additional rounds and the overall selection was performed in replicate. Selection of peptides using phage libraries is typically reported from one or two screenings 68, 69 .