# Mapping Gamma Globin Regulatory Elements

> **NIH NIH F32** · ST. JUDE CHILDREN'S RESEARCH HOSPITAL · 2020 · $69,306

## Abstract

PROJECT SUMMARY/ABSTRACT
I am studying developmental globin gene expression in order to better understand and treat sickle cell disease
(SCD) and β-thalassemia, common disorders that cause substantial morbidity and early mortality around the
world. Both diseases become symptomatic between birth and 6 months age, as the γ-globin (HBG1/2) genes
switch to β-globin (HBB) causing fetal hemoglobin (HbF, α2γ2) to be replaced by adult hemoglobin (HbA, α2β2).
Elevated postnatal levels of HbF, including a benign condition termed "hereditary persistence of fetal hemoglobin
(HPFH)", alleviates SCD and β-thalassemia. We are seeking to reverse the γ-to-β-globin gene switch in order to
induce HbF therapeutically. Our preliminary studies show that CRISPR/Cas9-mediated genome editing of
primary human hematopoietic stem and progenitor cells (HPSCs) can recreate an HPFH-associated 13-
nucleotide deletion in the HBG1 promoter by removing nucleotides -102 to -114 upstream of the transcriptional
start site. In vitro differentiation of the gene-edited HSPCs generates erythroid progeny with HbF elevated to
potentially therapeutic levels. Now, I will fine-map the -102 to -114 region using genome editing-mediated
homology directed repair and cellular assays for Hb switching in order to better define the relevant nucleotide
motif(s) and their interacting transcription factors (Aim 1). Because HBG1 and HBG2 are nearly identical,
CRISPR/Cas9 gene editing of the -102 promoter regions introduces double-stranded DNA breaks in both genes
simultaneously, which could cause deletion of the 5 kb intervening sequence, including HBG2. Preliminary
studies indicate that approximately 20% of alleles harbor the 5 kb deletion after editing of human CD34+ HSPCs.
Surprisingly, however, erythroid clones with this deletion exhibit increased γ-globin production, despite loss of
HBG2. Epigenetic analysis and functional testing of targeted deletions within the deleted HBG1-HBG2 intergenic
region identified a potential DNA regulatory element that represses HBG1 transcription. I will better define this
element by mutational analysis using cellular assays for globin switching and epigenetic studies to map
associated transcription factors and effects on three-dimensional chromatin structure (Aim 2). By characterizing
the cis elements and the corresponding trans-acting factors that regulate γ-to-β globin switching, I hope to
develop new ways to induce HbF for treating SCD and β-thalassemia.

## Key facts

- **NIH application ID:** 9971525
- **Project number:** 5F32DK118822-03
- **Recipient organization:** ST. JUDE CHILDREN'S RESEARCH HOSPITAL
- **Principal Investigator:** Phillip A Doerfler
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $69,306
- **Award type:** 5
- **Project period:** 2018-07-05 → 2021-07-04

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9971525

## Citation

> US National Institutes of Health, RePORTER application 9971525, Mapping Gamma Globin Regulatory Elements (5F32DK118822-03). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9971525. Licensed CC0.

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