GFP-on mouse model for interrogation of in vivo gene editing

The development of Cas9 technology has spurred advances in in vivo treatments, offering promising avenues for targeted gene editing within living organisms. However, the challenge of efficient and specific delivery remains a significant hurdle in harnessing the full potential of Cas9-based therapies. Here, we have generated a GFP-on reporter mouse model, harboring Q81X mutation in the EGFP sequence, a nonsense mutation that can be efficiently corrected with an SpCas9 adenine base editor (ABE). The GFP-on mouse model was validated using dual adeno-associated virus serotype 9 (AAV9) encoding ABE8e and EGFPQ81X guide RNA (gRNA) and VSV-G pseudotyped engineered virus-like particles (eVLPs) containing the same ABE8e protein and gRNA. Following intravenous administration of AAV9-SpABE8e and SpABE8e-eVLPs into GFP-on mice, EGFP expression was observed in tissues and organs consistent with the cellular tropism of AAV9 and VSVG-pseudotyped particles. Intrahepatic delivery of AAV9-SpABE8e into fetuses showed restoration of fluorescence in AAV9-targeted organs lasting up to 6 months in treated mice. These findings illustrate the successful establishment of an EGFP point mutant mouse model, ideal for swiftly testing gene editing tools, assessing various delivery strategies, and screening tissue-specific targeting. In summary, this GFP-on mouse model is an ideal tool for optimizing in vivo gene editing.