[R21] Developing Improved Reporter Mouse Models for In Vivo Gene Editing
Ente: NIH Office of the Director
Scadenza: 2028-06-30
Importo max: 486.750 EUR
Paese: US
Descrizione
PROJECT SUMMARY
CRISPR-Cas9 technologies have revolutionized genetic engineering and have led to a renaissance in
genetic therapy research. The initial CRISPR discovery was that of an RNA-guided endonuclease capable of
creating double-strand DNA breaks which can be precisely corrected using DNA donor-mediated homology-
directed repair (HDR). While an advance, this approach is complicated by low efficiency and non-therapeutic
side products arising from non-homologous end joining (NHEJ). Among the most promising of various newer
approaches are A- and C-base editing, which allow for precise genome modification of adenine and cytosine
residues, respectively, without the generation (and drawbacks) of double-strand DNA breaks. Testing the safety
and efficacy of such approaches relies on preclinical studies often performed using mouse models. To this point,
mouse gene editing reporter strains serve many roles in characterizing gene editors. For example, gene editing
reporter strains can be used to track the timing and location of gene editing, assess gene editing efficiency,
optimize gene editor delivery, discriminate between repair mechanisms, and lay the groundwork for clinical
studies through preclinical testing of emerging therapeutics.
During Phase 1 of the Somatic Cell Genome Editing Consortium, The Jackson Laboratory established a
Gene Editing and Testing Center to design, engineer, and deploy new reporter models. We created three new
mouse gene editing reporter strains that can effectively indicate HDR, NHEJ, and A-base editing activity at single-
cell resolution through the activation of a fluorescent marker. These models are now available for public
distribution and are well-utilized by the therapeutic gene editing community. Despite these successes, gaps
remain in the mouse gene editing and characterization toolbox limiting the number of editing outcomes that can
be reliably measured, in vivo. This proposal aims to address these needs by constructing and characterizing two
mouse gene editing reporter strains. The first strain will enable detection of C-base editor activity, an important
editing technology with growing therapeutic interest thanks to the development of C-base editor variants with
high specificity. The second strain is a multifunctional reporter that addresses the specific need for a reporter
that can faithfully detect most NHEJ events, but that also reports, HDR, A-base editing, and Cre-directed
recombination. In all, this multifunctional reporter assesses a number of important activities for evaluating new
gene editing platforms. Each of these reporters will be functionally characterized across three levels of biological
complexity, qualifying them for further use in preclinical testing.
Istituzione: JACKSON LABORATORY
PI: DAVID ERIC BERGSTROM
Progetto: 1R21OD040605-01
Settori: NIH Office of the Director
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