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NIH grant 1R21EB039470-01: Targeting repeat expansion instability with de novo DNA slip-out binding proteins
Institute: {'code': 'EB', 'abbreviation': 'NIBIB', 'name': 'National Institute of Biomedical Imaging and Bioengineering', 'admin_org_id': '12500035', 'admin_funding_url': ''}. PI: Cameron Glasscock. Award: $246208.
Start: 2026-08-22T00:00:00.
PROJECT SUMMARY/ABSTRACT
DNA repeat expansions, characterized by the abnormal expansion of genetically unstable, short, repetitive
DNA sequences, cause over 60 REDs including Huntington's disease (HD), myotonic dystrophy, and
spinocerebral ataxia, that result in a range of neurological and other health problems. While RED
pathogenesis can arise from a variety of downstream mechanisms, including production of aberrant RNA
accumulation and mutant protein aggregation, evidence is accumulating that a shared characteristic for
multiple REDs is the somatic instability (SI) of DNA repeats that underlies disease progression. Emerging
evidence for HD, the archetypal RED, suggests that disease onset occurs upon crossing a toxic threshold
of repeat length as patients age, a likely feature of RED pathogenesis more broadly. Hence, we expect that
a general therapeutic approach for REDs that rely on SI should target the mechanisms underlying the genetic
instability. However, there are few approaches under exploration that can halt SI and that don't also alter
normal processes by knocking down repair enzymes. This proposal's objective is to pioneer a novel
gene therapy approach for halting somatic instability by developing new molecular tools that target
DNA structure. By developing such tools, the aims of this proposal will overcome a critical limitation of
existing genome targeting technologies that only recognize linear DNA sequence.
We will pursue two parallel approaches, using HD as a paradigm RED, that serve to de-risk our overarching
goal. In Aim 1, we will leverage AI-based computational methods for design of DNA-binding proteins that
selectively target DNA hairpin or bulge structures that form at slip-outs of pathogenic expanded repeats. We
will fuse these proteins with strand-selective nickases to test our hypothesis that slip-out selective nicking
can induce repeat contractions. In Aim 2, as a secondary approach, we will engineer slip-out selective
cytosine base editors to slow SI by stabilizing the DNA duplex through repeat interruption. To do this, we will
leverage the natural properties of cytidine deaminase enzymes that act on ssDNA substrates. Finally, in
Aim 3, we will stablish impact of slip-out selective molecular tools to induce contractions or slow SI in vivo
in an HD knock-in mouse model. The expected outcome of this study will be development of a new
technologies for structure-specific DNA targeting and a proof-of-principle demonstration of their use to
induce repeat contractions or slow SI in HD. The high-risk-high impact nature of this proposal is aligned with
the mission of NIBIB, the Biomolecular Technologies program area, and the explicit goals of the Trailblazer
R21 mechanism for Early-Stage Investigators because of its focus on developing an exploratory molecular
technology relevant to a broad spectrum of diseases.document fetched 2026-08-30 02:46