Forced enhancer-promoter rewiring to alter gene expression in animal models

Scott A Peslak; Selami Demirci; Vemika Chandra; Byoung Ryu; Saurabh K Bhardwaj; Jing Jiang; Jeremy W Rupon; Robert E Throm; Naoya Uchida; Alexis Leonard; Khaled Essawi; Aylin C Bonifacino; Allen E Krouse; Nathaniel S Linde; Robert E Donahue; Francesca Ferrara; Matthew Wielgosz; Osheiza Abdulmalik; Nicole Hamagami; Paula Germino-Watnick; Anh Le; Rebecca Chu; Malikiya Hinds; Mitchell J Weiss; Wei Tong; John F Tisdale; Gerd A Blobel

doi:10.1016/j.omtn.2023.01.016

Forced enhancer-promoter rewiring to alter gene expression in animal models

Mol Ther Nucleic Acids. 2023 Jan 31:31:452-465. doi: 10.1016/j.omtn.2023.01.016. eCollection 2023 Mar 14.

Authors

Scott A Peslak^{1

2}, Selami Demirci³, Vemika Chandra², Byoung Ryu⁴, Saurabh K Bhardwaj², Jing Jiang^{2

5}, Jeremy W Rupon², Robert E Throm⁴, Naoya Uchida^{3

6}, Alexis Leonard³, Khaled Essawi^{3

7}, Aylin C Bonifacino⁸, Allen E Krouse⁸, Nathaniel S Linde⁸, Robert E Donahue³, Francesca Ferrara⁴, Matthew Wielgosz⁴, Osheiza Abdulmalik², Nicole Hamagami², Paula Germino-Watnick³, Anh Le³, Rebecca Chu³, Malikiya Hinds³, Mitchell J Weiss⁴, Wei Tong², John F Tisdale³, Gerd A Blobel²

Affiliations

¹ Division of Hematology/Oncology, Department of Medicine, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA.
² Division of Hematology, The Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
³ Cellular and Molecular Therapeutics Branch, National Heart, Lung, and Blood Institutes (NHLBI), National Institutes of Health (NIH), Bethesda, MD 20892, USA.
⁴ Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
⁵ CAS Engineering Laboratory for Nanozyme, Institute of Biophysics, Chinese Academy of Sciences, Beijing, People's Republic of China.
⁶ Division of Molecular and Medical Genetics, Center for Gene and Cell Therapy, The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan.
⁷ Department of Medical Laboratory Science, College of Applied Medical Sciences, Jazan University, Jazan, Saudi Arabia.
⁸ Translational Stem Cell Biology Branch, NHLBI, NIH, Bethesda, MD 20814, USA.

Abstract

Transcriptional enhancers can be in physical proximity of their target genes via chromatin looping. The enhancer at the β-globin locus (locus control region [LCR]) contacts the fetal-type (HBG) and adult-type (HBB) β-globin genes during corresponding developmental stages. We have demonstrated previously that forcing proximity between the LCR and HBG genes in cultured adult-stage erythroid cells can activate HBG transcription. Activation of HBG expression in erythroid cells is of benefit to patients with sickle cell disease. Here, using the β-globin locus as a model, we provide proof of concept at the organismal level that forced enhancer rewiring might present a strategy to alter gene expression for therapeutic purposes. Hematopoietic stem and progenitor cells (HSPCs) from mice bearing human β-globin genes were transduced with lentiviral vectors expressing a synthetic transcription factor (ZF-Ldb1) that fosters LCR-HBG contacts. When engrafted into host animals, HSPCs gave rise to adult-type erythroid cells with elevated HBG expression. Vectors containing ZF-Ldb1 were optimized for activity in cultured human and rhesus macaque erythroid cells. Upon transplantation into rhesus macaques, erythroid cells from HSPCs expressing ZF-Ldb1 displayed elevated HBG production. These findings in two animal models suggest that forced redirection of gene-regulatory elements may be used to alter gene expression to treat disease.

Keywords: MT: Oligonucleotides; Therapies and Applications; enhancer-promoter interactions; fetal hemoglobin; forced chromatin looping; hemoglobin switching; preclinical animal models; sickle cell disease; vector optimization.

Abstract

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