# PNA Nanoparticles for Gene Editing In Vivo

> **NIH NIH U01** · YALE UNIVERSITY · 2020 · $404,237

## Abstract

There is substantial interest in gene editing as a potential means to treat human genetic disorders such as
thalassemia and sickle cell disease. Much effort has been focused on targeted nucleases such as
CRISPR/Cas9 and zinc-finger nucleases (ZFNs), based on work showing that site-directed DNA damage
strongly promotes homologous recombination (HR). However, clinical application of targeted nucleases is
challenged by the risk of off-target cleavage events in the genome. As an alternative, in work recently
published in Nature Communications, the Ly, Saltzman, and Glazer labs have shown that γ-substituted triplex-
forming peptide nucleic acids (PNAs) and donor DNAs delivered intravenously (IV) via poly(lactic-co-glycolic)
acid (PLGA) nanoparticles (NPs) into a mouse model of human β-thalassemia produced almost complete
amelioration of the disease, with clinically relevant β-globin gene correction frequencies in hematopoietic stem
cells (HSCs) of up to 7%. The mice showed alleviation of anemia, improvement in RBC morphologies, and
reversal of splenomegaly and extramedullary hematopoiesis, with extremely low off-target effects in the
genome, a key advantage of this technology. The other key advantage is that the components can be
synthesized chemically and formulated into nanoparticles for simple IV administration. However, synthesis of
γPNAs is complicated and expensive, and they are not commercially available, limiting the ability of
investigators to exploit this technology. In line with RFA-RM-18-024, “Expanding the Human Genome
Engineering Repertoire”, this multi-PI proposal by Ly, Saltzman, and Glazer seeks to advance PNA/NP-based
gene editing by simplifying and scaling up PNA synthesis, by incorporating next generation PNA chemistry to
boost binding affinity, increase selectivity, and enhance potency, and by strategically exploiting cellular DNA
repair pathways. The Specific Aims are: (1) To scale up PNA production and augment DNA binding, in order to
expedite the translation of PNAs for therapeutic gene editing and enable widespread adoption of the
technology. We will devise an enantioselective strategy for scaling up the production of monomers, and we will
synthesize and test γPNAs with modified nucleobases to achieve improved DNA binding properties and to
overcome the homopurine sequence restriction for triplex formation. (2) To develop strategies to manipulate
DNA repair to enhance the efficiency of PNA-mediated gene editing, based on promising preliminary results
with a novel DNA repair inhibitor. (3) To provide a robust platform of assays to evaluate the advancements
from Aims 1-2 and to generalize this approach to multiple genes. We will continue to exploit facile mouse- and
cell-based assays for correction of the human β-globin gene at the IVS2-654 thalassemia mutation. We expect
this work to provide the basis for designing even more potent PNAs applicable to gene editing for many human
genetic disorders.

## Key facts

- **NIH application ID:** 9971452
- **Project number:** 5U01AI145965-02
- **Recipient organization:** YALE UNIVERSITY
- **Principal Investigator:** PETER M GLAZER
- **Activity code:** U01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $404,237
- **Award type:** 5
- **Project period:** 2019-07-05 → 2023-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9971452

## Citation

> US National Institutes of Health, RePORTER application 9971452, PNA Nanoparticles for Gene Editing In Vivo (5U01AI145965-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9971452. Licensed CC0.

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