AAV4-Cre

Gene therapy using adeno-associated virus (AAV) vectors requires knowledge of their tropism in vivo. Here, researchers analyzed the tropism of ten naturally occurring AAV serotypes (AAV3B, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAVrh8, AAVrh10, and AAVrh74) after systemic delivery into male and female mice. Transgenes expressing ZsGreen and Cre recombinase were used to identify transduced cells in a cell-dependent manner based on fluorescence. Cre-driven tdTomato fluorescence activation provided excellent sensitivity for transduced cells. All serotypes, except AAV3B and AAV4, had high liver tropism. Fluorescence activation revealed transduction of unexpected tissues, including adrenal glands, testes, and ovaries. Rare transduced cells within tissues were also readily observed. Biodistribution of the AAV genome correlated with fluorescence, with the exception of immune tissues. The study found that AAV4 has pan-endothelial tropism and also targets pancreatic β cells.

Here, researchers generated AAV4 vectors expressing Cre recombinase. Intravenous injection of AAV4-Cre into Ai9 mice resulted in abundant tdTomato-positive signals in the lungs outside the airways (Figure 1A). Flow cytometry showed that the majority of tdTomato signals originated from endothelial cells based on CD31+/CD45- expression. Notably, >85% of lung endothelial cells were transduced by AAV4 in both male and female mice. In contrast, only about 20% of non-immune non-endothelial cells in the lung appeared to be transduced by AAV4 (Figure 1B). Repeating the experiment using an AAV4 vector expressing Cre demonstrated poor AAV4 transduction of the liver, showing only sparse TdTomato hepatocytes and possible signal in isolated liver sinusoidal endothelial cells. In stark contrast, impressive endothelial cell transduction was observed in the stomach, bladder, adrenal glands, pancreas, brain, small intestine, and thymus (Figure 1C). Furthermore, co-staining with insulin antibodies confirmed that AAV4 could effectively transduce β cells (Figure 1D).

Figure 1. AAV4 exhibits a unique tropism for endothelial cells and pancreatic beta cells. (Walkey C J, et al., 2024)