MGTA-456, a First-in-Class Cell Therapy Produced from a Single Cord Blood Unit, Enables a Reduced Intensity Conditioning Regimen and Enhances Speed and Level of Human Microglia Engraftment in the Brains of NSG Mice

Background. Allogeneic bone marrow transplant (BMT) is a promising, curative approach for patients with inherited metabolic disorders (IMDs), a class of pediatric diseases characterized by a single enzyme deficiency. The goal of transplant is to provide cells that produce functional enzymes otherwise deficient in these patients, and thereby prevent or ameliorate neurological complications associated with selected IMDs. Donor-derived microglial cells are protective, limiting neurological disease progression. For IMD patients who do not have an HLA matched, non-carrier related donor, cord blood (CB) is the preferred HSPC source given its rapid availability and superior clinical outcomes compared to other graft sources. CB, however, is associated with delayed hematopoietic recovery and relatively poor engraftment due to the limited numbers of hematopoietic stem cells (HSCs) in a CB unit, delaying enzyme/protein reconstitution and cross-correction of non-hematopoietic cells.

An aryl hydrocarbon receptor antagonist (AHRa)-based culture has been shown to expand CB CD34+ and CD34+CD90+ cells 330-fold and 100-fold, respectively, leading to rapid hematopoietic recovery after infusion of the clinical product, MGTA-456 (Wagner et al., Cell Stem Cell 2016 and Orchard et al., ASH 2018). As microglia are thought to be derived from HSCs, we hypothesized that MGTA-456 might lead to faster and greater microglial engraftment and potentially enable reduced intensity conditioning. Here, we assessed human hematopoietic and brain engraftment in NSG mice after transplant with MGTA-456 and showed that microglia engrafted faster with MGTA-456, less conditioning was needed, and that, mechanistically, these cells are derived from the CD34+CD90+ cell fraction.

Methods. CB CD34+ cells were expanded in growth factor-supplemented media with or without an AHRa for 10 days. NSG mice were transplanted with unmanipulated CB CD34+ cells or the expanded product after 200 cGy total body irradiation or busulfan (BU) dosed at 20 or 40 mg/kg ip. Microglial engraftment was measured by flow cytometry of homogenized brains, quantitating the number of CD45+CD11b+Iba1+ cells, and by immunohistochemistry of brain sections.

Results. Relative to naïve, unmanipulated CB CD34+ cells, transplant of MGTA-456 into sublethally irradiated mice led to an 8-fold increase in hematopoietic engraftment and a 10-fold increase in microglial engraftment in the brain (p<0.0001, n=15 mice), with histology consistent with engrafting microglia. As high dose BU enables enhanced microglia engraftment relative to irradiation by crossing the blood brain barrier and clearing host microglia (Capotondo et al., PNAS 2013), we evaluated the effectiveness of MGTA-456 after BU conditioning at 20 or 40 mg/kg. Transplant of MGTA-456 led to a 37-fold increase in engraftment relative to mice transplanted with unmanipulated CB CD34+ cells (p<0.001, n=8). Notably, transplant of MGTA-456 into mice conditioned with low-dose BU (20 mg/kg) led to a 15-fold increase in engraftment relative to high-dose BU (40 mg/kg)-conditioned animals transplanted with unmanipulated CB CD34+ cells (p<0.001, n=8). To evaluate speed of microglial engraftment, we evaluated brains weekly to 16 weeks after transplant. A 28-fold increase in microglial engraftment was demonstrated as early as 2 weeks post-transplant with MGTA-456 (p<0.0001, n=8). Number of engrafting hematopoietic cells in the periphery correlated with number of engrafting microglia in the brain (p<0.0001). Lastly, subpopulations of MGTA-456 were evaluated to determine the source of microglial engraftment. Only CD34+CD90+ cells, but not CD34+CD90- or CD34- cells, led to brain engraftment, consistent with the subpopulation of cells that result in hematopoietic engraftment following transplant of unexpanded cells (Radtke et al., Sci Trans Med 2017 and Goncalves et al., Blood 2017 130:659).

Conclusions. These studies demonstrate that microglial engraftment is faster and greater in recipients of MGTA-456 even after lower dose BU conditioning, that microglial engraftment correlates with peripheral blood recovery, and that microglia cells are derived from CD34+CD90+ cells. These results suggest that lower dose BU may improve safety without jeopardizing efficacy in IMD recipients of MGTA-456. A Phase 2 clinical trial is ongoing to evaluate transplant of MGTA-456 in patients with select IMDs.

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