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Final Program


CRS Abstract Guide

Session: Technical Session VII

Nanomedicine and Nanoscale Delivery VIII

Friday, July 12, 2024
8:00 AM - 10:00 AM CET
Location: Sala Europa
Title: Controlling the protein corona of siRNA nanocarriers to reach undruggable oncoproteins: the KRAS case.

Introduction: RNA interference offers great applicability in addressing undruggable cancer targets such as oncogenic KRAS, however, the delivery challenge toward target tissues remains. Nanotechnology provides peerless opportunities to tackle this challenge, yet understanding the biological identity of nanocarriers (NCs) upon protein corona formation is crucial to develop translatable drug delivery systems.

Methods: A combination of Design-of-Experiment (DoE) and rational design was employed to engineer NCs. NCs were characterized by dynamic light scattering, RNA encapsulation assays and cryogenic electron microscopy. Over 30 NCs encapsulating siGFP were evaluated for their silencing and toxicity in GFP-expressing HeLa cells via flow cytometry. The protein corona of the lead NC1 was quantified by SWATH-MS/MS and compared to variations thereof with altered helper and PEG lipid identity and PEG density. NC1 was functionalized with a cancer targeting peptide via click-chemistry. The capacity of therapeutic siKRAS-carrying NC1 to silence KRAS and downstream effectors was analyzed by Western Blot in KRASmutant A549 cells. The proliferation of siKRAS-treated A549 cells was
studied in a clonogenic assay.

Results: Lead NC1 exhibited a size of 70 nm, PDI of 0.1, neutral charge and RNA encapsulation of 85%, low toxicity and prominent silencing of model GFP. Protein corona analysis of NC1 and its variants demonstrated the influence of helper and PEG lipid identity and PEG density on corona fingerprint. NC1 attracted a corona enriched in vitronectin, targeting its cognate receptor expressed by cancer cells. Lead candidates elicited dose-dependent silencing of KRAS and downstream pERK proteins. Functional evaluation of NC1 siKRAS revealed suppression of up to 90% on lung cancer colony formation, outperforming benchmark LNPs.
Conclusions/Impact: We designed, characterized, and evaluated a versatile class of NCs for the delivery of siRNA. KRAS knockdown translated into dramatic inhibition of cancer cell proliferation. The protein corona analysis linked the composition of NCs with their corona fingerprint, ultimately governing their biological fate. The ongoing in vivo efficacy study in an orthotopic A549 lung cancer model in combination with pan-ERBB inhibitor Afatinib will determine the antitumoral capacity of targeting peptide.
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conjugated NC1-siKRAS.