TIGER: A tdTomato in-vivo genome-editing reporter mouse for investigating precision-editor delivery approaches

In vivo genome editing has the potential to address many inherited and
environmental disorders. However, a major hurdle for the clinical
translation of genome editing is safe, efficient delivery to
disease-relevant tissues. A modality-agnostic reporter animal model that
facilitates rapid, precise, and quantifiable assessment of functional
delivery and editing could greatly enhance the evaluation and translation
of delivery technologies. Here, we present the development of the TIGER
(tdTomato in vivo genome-editing reporter) mouse, a reporter strain that
harbors an integrated and constitutively expressed mutated tdTomato gene
in the Polr2a locus. The mutations (Q115X, Q357X) abolish fluorescence,
but successful adenine base editing (ABE) or prime editing (PE) restores
tdTomato fluorescence. This mouse model facilitates the tissue- and
cell-type-specific assessment of genome editing agent delivery. We
describe several editing strategies validated in vitro and demonstrate
efficient ABE and PE in vivo using viral and non-viral delivery vectors
targeting four cell types within the mouse eye: the retinal pigment
epithelium (RPE), photoreceptors, Müller glia, and the trabecular
meshwork. We show direct editing characterization in the ocular tissues
via in vivo and ex vivo two-photon confocal microscopy, and verify the
spectral and fluorescence lifetime properties of tdTomato reporter in
other mouse tissues. Additionally, we demonstrate successful AAV-mediated
prime editing of extraocular tissues, including hepatocytes, skeletal
muscle, and brain neurons, by intravenous injection. Thus, the TIGER mouse
facilitates the direct development, comparison, and optimization of
delivery platforms for efficient and productive ABE or PE broadly
applicable in vivo across multiple tissues tested in this study.