Biophysical Characteristic Analysis of Tolerogenic Nanoparticle Containing CRISPR CAS-9

Maureen Wang

Pharmacy building where Maureen carried out the project.

Pharmacy building where Maureen carried out the project.

Undergraduate Student Project

Introduction

Have you ever thought about gene editing? With CRISPR technology, gene editing is a lot faster, cheaper, and more deficient.

My name is Maureen Wang and I am a Pharmaceutical Sciences major at SUNY University at Buffalo. I am currently doing undergraduate research at the School of Pharmacy and Pharmaceutical Sciences. In the past semester, I got to work in Dr. Balu-Iyer's lab in the Pharmaceutical Sciences department. With the help of others in the lab, I was able to design and start my undergraduate project on the biophysical properties of tolerogenic properties of Cas-9 encapsulated in Lyso-Phosphatidylserine.

CRISPR is a powerful tool for editing one's genome; however, because of antibodies the therapeutic effects and safety of the drug is decreased. By looking into the biophysical characteristics of Cas-9 protein can help better understand the structure of the protein and encapsulate in lipid to increase therapeutic effects.

Abstract

Clustered regularly interspaced short palindromic repeats, or CRISPR, carries out targeted genetic changes in eukaryotic cells. However, the presence of anti-Cas9 antibodies lead to a decrease in efficacy. Phosphatidylserine (PS) and Lyso-Phosphatidylserine (Lyso-PS) can prevent immunogenicity by converting the immunogenic protein to a tolerogen during therapeutic use. The objective is to reverse anti-Cas9 antibodies by establishing a stable nanoparticle formulation. The protein and lipid was prepared and sized. Circular dichroism (CD) and Fluorescence was performed to determine the ideal conformation for loading Cas9 protein into the nanoparticle through incubation. Florescence emission scan was performed to analyze the nanoparticle conformation. Anticipated results would show the sizing of the encapsulated protein in lipid and its encapsulation efficiency. Fluorescence test would be performed to determine the lipid:protein conformation. The hypothesis is supported if the results of the characterization show consistent nanoparticle sizing, high encapsulation efficiency, and minimal shift in fluorescence emission scan.

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