Transamniotic Delivery of Hematopoietic Stem Cells Genetically Modified to Carry a Human Hemoglobin Subunit Beta Gene (HBB) in a Healthy Rodent Model.

Background: We sought to determine whether transamniotic stem cell therapy (TRASCET) could be a viable alternative for the fetal administration of genetically modified hematopoietic stem cells (HSCs) carrying a human hemoglobin subunit beta gene (hHBB) in a healthy syngeneic rat model.

Methods: Time-dated pregnant Lewis dams underwent volume-matched intra-amniotic injections in all their fetuses (n = 61) of a suspension of donor HSCs genetically modified with either both a hHBB gene and a firefly luciferase reporter gene (n = 42) or the firefly luciferase reporter gene alone to control for HBB-derived protein interspecies homology (n = 19) on gestational day 17 (E17; term = E21). Donor HSCs consisted of syngeneic cells phenotyped by flow cytometry with successful hHBB transduction confirmed by ELISA prior to administration in vivo. At term, fetal samples from five anatomical sites relevant to hematopoiesis were screened for the presence of human hemoglobin subunit beta by ELISA and by digital droplet PCR (ddPCR).

Results: When controlled by HSCs without hHBB injections, human hemoglobin subunit beta production was documented at term in the fetal bone marrow and spleen (p < 0.001 and p = 0.028 respectively). Positive hHBB expression by ddPCR was detected in the spleen (54 %), bone marrow (46 %), blood (46 %), liver (23 %), and thymus (15 %).

Conclusions: Genetically modified hematopoietic stem cells carrying a human hemoglobin subunit beta gene can reach fetal hematopoietic sites after simple intra-amniotic injection in a healthy syngeneic rat model. Transamniotic hematopoietic stem cell-based gene therapy could become a novel strategy for the perinatal management of select hemoglobinopathies.

Level of evidence: N/A (animal and laboratory study).

Type of study: animal and laboratory study.