658. Increasing Fetal Hemoglobin Expression With TALEN Induced Small Deletions in Peripheral Blood Stem Cells

Abstract

Hemoglobinopathies are common and morbid inherited conditions caused by mutations in hemoglobin genes that generate pathologic globin molecules or reduced expression of wild type hemoglobin. A subset of individuals with hemoglobinopathy-associated beta globin mutations have a mild clinical phenotype and are found to carry additional disease modifying mutations including small deletions or point mutations that induce fetal hemoglobin expression. This condition is referred to as hereditary persistence of fetal hemoglobin. Recent progress in the arenas of gene editing and hematopoietic stem cell (HSC) expansion has raised the possibility of treating hemoglobinopathies through direct modification of genes in HSCs to increase the expression of fetal hemoglobin in the erythroid lineage. To date, most attempts to induce fetal hemoglobin have targeted the expression of transcription factors implicated in regulation of fetal hemoglobin production, such as Bcl11a, an approach which could lead to unintended and potentially deleterious effects on cellular phenotype. Here, we are evaluating the application of mRNA transfection of TALENs to directly modify transcription factor binding sites within the hemoglobin locus in HSCs to achieve efficient, stable expression of fetal hemoglobin. We have designed TALENs to induce small deletions in the beta hemoglobin promoter at the binding site of KLF1, postulated to play a role in localizing the active chromatin hub to and preferentially expressing adult beta hemoglobin, as well as other globin locus target regions implicated in the regulation of fetal hemoglobin production. By combining TALEN-mediated gene modification with novel HSC expansion approaches involving the recently described small molecule UM729, we hope to realize both efficient HSC modification and expansion with a sufficient level of fetal hemoglobin production in erythrocytes to achieve a functional cure for a wide range of beta hemoglobinopathies.