Cellular Delivery of Cas9/sgRNA RNPs by hydrophobic Oligoamino Amides


Dorrer, Sarah  


Gene therapy is gaining increasing interest in its universal application in the treatment of cancer and other genetic diseases. Therefore, genetic engineering has become a fundamental and powerful tool aimed at: genetic, epigenetic, and transcriptomic abnormalities that cause disease. The discovery of CRISPR-Cas9 is very promising due to its ability to target specific and precise genome in vitro and in vivo. One of the biggest problems with CRISPR-Cas9 transfection via RNP is the decreased in vivo efficiency caused by enzyme degradation before reaching the destination, as well as low cell permeability. Therefore, there is a need to develop a transport system to ensure adequate protection of the load and cell uptake, although these systems must have other properties such as low immunogenicity and toxicity. Until a few years ago, peptide-based carriers, such as oligo (ethylamino) amides (OAA), showed great potential as administration platforms. Due to the easy and rapid synthesis of defined OAA sequences, these are suitable for various types of therapy, such as the delivery of DNA, RNA, proteins, and drugs. Therefore, a library containing new lipo-OAA has been established in this project. Synthesized building blocks differ in hydrophobicity by modifying the acid domain (Stp-Analogs) or the spine (Gtt-Analogs). The efficiency of the Cas 9 / sgRNA RNP administration agents was evaluated by cellular tests targeting Hela GFP Tubulin cells. It has been shown that more hydrophobic building blocks compared to 1445 (Stp) increase transfection efficiency. Even at a ten times greater dilution, a transfection efficiency> 90% was obtained at 1633 (Htp). A promising result in high-efficiency genome editing using OAA would be of great importance for future individual therapies.



Cuenca González, Ana Belén
Lächelt, Ulrich


IQS SE - Master’s Degree in Pharmaceutical Chemistry