In Utero Stem Cell Transplantation in Patients with Alpha Thalassemia Major: Interim Results of a Phase 1 Clinical Trial
Authors: MacKenzie Tippi C. , Frascoli Michela , Sper Renan , Lianoglou Billie Rachael , Gonzalez Velez Juan , Dvorak Christopher C. , Kharbanda Sandhya , Vichinsky Elliott 
Published: February 2021

Introduction: In utero hematopoietic cell transplantation (IUHCT) has the potential to treat patients with blood disorders by taking advantage of bidirectional maternal-fetal tolerance during pregnancy. Although engraftment has only been seen in immunodeficient patients in the past, recent preclinical studies indicate that a higher dose of maternal cells should be given intravascularly to maximize engraftment in other settings. We hypothesized that: 1) fetuses will tolerate a maternal IUHCT without conditioning and develop low engraftment with sustained tolerance to maternal alloantigens; and 2) tolerant patients could receive a postnatal booster maternal transplant with minimal immunoablative conditioning and thereby achieve a cure. To test the first of these hypotheses in a controlled setting, we have initiated a phase 1 clinical trial of IUHCT in fetuses with alpha thalassemia major (ATM), who already require a fetal intervention (in utero transfusions (IUT)) to survive the pregnancy. We present interim results from the first two patients.
Methods: After obtaining IND approval, we initiated the clinical trial in 2017 (NCT02986698). Fetuses with ATM between 18-26 weeks' gestation without other major anomalies were included. To date, we have evaluated 19 patients for possible inclusion: 15 patients terminated the pregnancy, 2 patients were outside the inclusion window and underwent IUT alone, and 2 have been transplanted and completed the 1 year follow up. For IUHCT, 300 ml of maternal bone marrow was harvested, CD34+ cells were selected and transplanted fresh into the fetus via the umbilical vein, along with a standard IUT. IUTs were repeated every 3 weeks until delivery. Maternal cell chimerism was detected using qRT-PCR for non-inherited maternal HLA-DR alleles on sorted cell populations in cord blood and after birth. Mixed lymphocyte reactions (MLRs) were performed by culturing CFSE-labeled host CD4 and CD8 responder cells with maternal (or third-party) derived antigen presenting cells (APC; direct pathway) or self APC/maternal lysate (indirect pathway).
Results: Both fetuses were diagnosed with severe hydrops on ultrasound and received an initial IUT prior to the IUHCT. Patient 1 was transplanted at 23 weeks with 1x 108CD34+ cells/kg + 1 x 107T cells/kg and Patient 2 was transplanted at 25 weeks with 5x 107CD34+ cells/kg + 5 x 106T cells/kg; both received 4 more IUTs until birth. Prenatal hydrops resolved in both and neonatal hospitalizations were 16 and 13 days, respectively. Maternal chimerism was detected in both patients, but levels were higher and persisted throughout the year only in patient 1 (Fig 1A). We detected 70-fold higher levels of erythrocyte precursors (Fig 1B) but normal levels of CD34+ cells in the cord blood of patients compared to healthy controls, highlighting a potential barrier to engraftment in a disease with prenatal hypoxia.
Patient 1 demonstrated sustained tolerance to maternal antigens by MLR throughout the year, whereas patient 2 showed reactivity against maternal antigens (Fig 1C). HLA antibodies were negative in both. Based on the finding of donor-specific tolerance in patient 1, we offered a booster transplant with maternal cells but the parents declined. Both patients are undergoing monthly transfusions and have had normal development and Bayley neurologic testing at 1 year. There were no unanticipated safety events in either mother/child pair.
Conclusions: These results demonstrate the safety of IUHCT in fetuses with ATM and the potential of fetal transplantation in the second trimester to sustain tolerance to maternal cells after birth. Low engraftment levels are predicted in the setting of host hyperproliferation and lack of space-making conditioning, confirming the need for a two-step strategy (IUHCT and postnatal booster transplant) for a definitive cure. The earlier transplantation in patient 1 enabled a higher dose of cells and may explain the improved outcome observed. Implementation of earlier screening in at-risk patient groups could allow earlier diagnosis and enrollment in the future. Although parents of fetuses with ATM are commonly counseled for pregnancy termination due to concern for poor outcomes, our results indicate that fetal therapy with IUTs can result in survival to term and excellent quality of life. This strategy of maternal IUHCT could be employed to treat fetuses with other hematologic disorders in the future.
Figure 1

Disclosures
MacKenzie: Acrigen: Membership on an entity's Board of Directors or advisory committees; Ultragenyx: Research Funding. Vichinsky:Global Blood Therapeutics: Consultancy; Agios: Research Funding; Pfizer: Research Funding.