# Altering VP1 and VP2 expression in trans affects the transduction efficiency of AAV9

## TL;DR

This paper shows that changing the amounts of VP1 and VP2 proteins in AAV9 capsids can improve how well they deliver genes into cells.

## Contribution

The study introduces a modified transfection protocol to independently control VP1 and VP2 expression, affecting AAV9 transduction efficiency.

## Key findings

- Modifying VP1 and VP2 expression ratios alters the capsid composition of AAV9.
- Increasing VP1 or VP2 in the capsid improves transduction efficiency in HEK293 cells.
- The findings enhance understanding of AAV biology for potential gene therapy applications.

## Abstract

Currently, adeno-associated virus (AAV) is one of the most reliable carrier for gene delivery in both proliferating and non-proliferating cells. Stable and long-lasting transgene expression has made this viral vector a key platform for the development of advanced therapy. Nevertheless, the widespread clinical use of AAV-based drugs remains limited due to their immunogenicity, low capsid capacity, and restricted tissue tropism. Tissue tropism depends largely on the the transduction efficiency of AAV capsids. In this study, we modified the standard three-plasmid transfection protocol to provide independent expression of VP1 or VP2 proteins from separate plasmids. Adjusting the ratio of these plasmids in the transfection mixture enabled alteration of the stoichiometric composition of the capsids, as SDS-PAGE and mass spectrometry confirmed. Increasing the amount of VP1 or VP2 in the capsid composition enhanced transduction efficiency, as demonstrated in vitro experiments on HEK293 cells. Obtained results contribute to a more comprehensive understanding of the AAV biology and have perspective of application in gene therapy.