# Developing a novel ex vivo platform to support hematopoietic cells and characterize the stem cell niche

> **NIH NIH F30** · ALBERT EINSTEIN COLLEGE OF MEDICINE · 2020 · $50,520

## Abstract

PROJECT SUMMARY / ABSTRACT
Current culture methods reduce the ability of hematopoietic stem cells (HSCs) to successfully engraft
in a host. Emerging gene editing technologies such as CRISPR/Cas9 require time in culture to allow
for the correction of disease-causing alleles. There is therefore a need to develop new methods of
culturing HSCs. Coculture of HSCs with bone marrow niche cells such as mesenchymal stem cells
(MSCs) is one possible solution to problems in HSC culture, as these cells provide factors that
support HSCs in vivo. However, MSCs cannot be maintained in culture for extended periods of time
and fairly rapidly lose expression of niche factors. Through a screen of candidate transcription factors,
our lab identified 5 factors that when transduced together restore niche factor expression and allow
for prolonged culture. These factors are Kruppel-like factor 7 (Klf7), Osteoclast stimulating factor
(Ostf1), X-box binding protein (XBP1), Interferon regulatory factor 3 (Irf3), and Irf7, which we
collectively dubbed the KOXII factors. KOXII-transduced MSCs were able to expand both murine and
human funtional HSCs to a much greater extent than mock-transduced MSCs. These cells therefore
may be useful in expanding HSCs ex vivo for gene correction. However, there are regulatory barriers
to the application of murine cells in human therapeutics. The work proposed here will in part focus on
the development and characterization of KOXII-transduced human MSCs. After generating these
cells, I will determine if the KOXII factors affect expression of niche factors in human MSCs. I will also
use flow cytometry and stem cell xenotransplantation to determine if KOXII-transduced MSCs are
more effective at driving HSC expansion than unmodified MSCs. Finally, using CRISPR/Cas9-based
gene editing of HSCs derived from patients with sickle cell disease as a model, I will determine if
coculture of patient cells with KOXII-transduced MSCs can improve the efficiency of gene editing or
increase the yield of properly edited cells over current standard HSC culture methods. In parallel, I
will use murine KOXII-transduced MSCs to more closely examine niche signalling by MSCs. As these
cells can be cultured in relatively large numbers, they are ideal for proteomic studies. In collaboration
with the lab of Jeroen Krijgsveld, I will examine the secretome of these cells in order to identify
proteins whose secretion is upregulated by the KOXII factors. Using both in vitro and in vivo assays, I
will evaluate the effect of these factors on HSC maintenance and proliferation, with the aim of
identifying secreted proteins with previously unappreciated roles in MSC-HSC niche interactions.

## Key facts

- **NIH application ID:** 10068797
- **Project number:** 1F30HL154749-01
- **Recipient organization:** ALBERT EINSTEIN COLLEGE OF MEDICINE
- **Principal Investigator:** Daniel K Borger
- **Activity code:** F30 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $50,520
- **Award type:** 1
- **Project period:** 2020-09-01 → 2024-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10068797

## Citation

> US National Institutes of Health, RePORTER application 10068797, Developing a novel ex vivo platform to support hematopoietic cells and characterize the stem cell niche (1F30HL154749-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10068797. Licensed CC0.

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