# Efficient Sendai virus mediated CRISPR/Cas9 gene editing to protect hematopoietic stem cells from HIV

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA LOS ANGELES · 2021 · $625,091

## Abstract

Project summary/abstract
The long-term objective of this proposal is to achieve highly efficient gene editing and maximize engraftment of
genetically engineered HIV-resistant hematopoietic stem/progenitor cells (HSPCs) to achieve HIV cure. HSPC
based gene therapy that results in HIV-resistant progenies from engrafted patient derived blood stem cells can
provide long-term protection against HIV with the promising possibility of achieving a cure. However, previous
HSPC based gene therapy strategies did not provide clear therapeutic effects. Recently, the ease and versatility
of the CRISPR/Cas9-mediated gene editing technology has spurred an immense interest in using it to edit the
CCR5 gene in HSPC based anti-HIV gene therapy strategies. Nonetheless, a successful cure by an anti-HIV
HSPC based gene therapy still has to overcome 2 major barriers: 1) insufficient levels of CCR5 gene modification
in HSPC and 2) poor engraftment of gene modified HSPCs. Firstly, to maximize the levels of CCR5 gene
modification in HSPCs without the residual genotoxicity associated with DNA based delivery platforms, we
developed a novel RNA-based Sendai virus (SeV) vector for highly efficient CRISPR/Cas9-mediated gene
editing of human HSPC. Our SeV-Cas9 transduces and edits these HSPCs with unprecedented efficiency (up
to ~80% at the CCR5 locus). Secondly, to improve the engraftment of gene modified HSPCs, we have developed
a novel in vivo chemoselection strategy that employs 6-thioguanine (6TG), a clinically available prodrug that
requires hypoxanthine-guanine phosphoribosyl-transferase (HPRT) for activity. Editing of HPRT allows for pre-
conditioning and post-transplant in vivo chemoselection of HPRT-deficient HSPC in the humanized bone
marrow, liver and thymus transplanted (hu BLT) mouse model. We hypothesize that efficient SeV delivered
CRISPR/Cas9 mediated editing of CCR5 and HPRT genes in HSPC followed by in vivo selection with 6TG will
maximize the engraftment of HIV-resistant HSPC. In toto, our strategy functionally maximizes ex vivo HPSC
gene modification and in vivo engraftment efficiency, which will continuously provide sufficient numbers of HIV-
resistant progenies that might ultimately replace HIV latently infected cells in hu BLT mice. To achieve our overall
goal, and to speed the clinical translation of our gene therapy strategy, we propose the following Specific Aims:
 AIM 1. Develop a clinically relevant SeV vector for highly efficient CRISPR/Cas9 mediated gene
 modification of HSPC.
 AIM 2. Maximize the efficiency of engraftment by a safe and effective pre-conditioning and selection strategy
 for CCR5 and HPRT modified HSPC in hu BLT mice.
 AIM 3. Investigate HIV inhibition by engraftment of CCR5 and HPRT modified HSPC in hu BLT mice.

## Key facts

- **NIH application ID:** 10171759
- **Project number:** 5R01AI138921-04
- **Recipient organization:** UNIVERSITY OF CALIFORNIA LOS ANGELES
- **Principal Investigator:** Dong Sung An
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $625,091
- **Award type:** 5
- **Project period:** 2018-06-11 → 2023-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10171759

## Citation

> US National Institutes of Health, RePORTER application 10171759, Efficient Sendai virus mediated CRISPR/Cas9 gene editing to protect hematopoietic stem cells from HIV (5R01AI138921-04). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10171759. Licensed CC0.

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