Transplantation of Hematopoietic Stem and Progenitor Cells from Cadaveric Organ Donors Leads to Long-Term Multilineage Engraftment in NSG Mice

Background

Infusion of Hematopoietic Stem and Progenitor Cells (HSPC) from cadaveric organ donors (CD) has been used anecdotally in the setting of allogeneic Hematopoietic Stem Cell transplants (HSCT) most commonly with the intention for induction of mixed hematopoietic chimerism and subsequent immunological tolerance. No systematic study has addressed the functional capacity of these cells for inducing long-term multi-lineage engraftment. Since the introduction of post-transplant Cyclophosphamide (PTCy) it has been possible to reliably overcome HLA barriers and achieve stable engraftment across extensively HLA-mismatched donor and recipient pairs. Banked HSPC from cadaveric donors may provide a new approach to expand the number of donors for HSCT. Usage of HSPC from bone marrow (BM) of banked cadaveric donors is viewed as no different than live donor aspirated marrow by the FDA but is not yet available on the registry. This emerging source could be of high importance to allow rapid on-demand access to transplants for patients who lack HSPC donors or induce stable mixed chimerism and immune tolerance in patients undergoing combined organ and BM transplantation. The objective of our studies was to investigate whether hCD34⁺HSC selected from cadaveric BM retain their capacity for long-term multi-lineage engraftment.

Methods and Results

We compared CD34⁺-selected HSPC obtained from CD and living donors (LD) regarding their in vitro and in vivo function and with particular focus on their ability to achieve long-term multi-lineage engraftment following transplantation into NSG mice. Bone marrow was recovered from vertebral bodies of CD and CD34⁺ selected. 5x10⁵ hCD34⁺ cells from 24 different CD were transplanted i.v. in to NSG or SGM3 mice (n=5 per donor) following 1Gy total body irradiation (TBI). Blood was drawn every two weeks, starting week 4 and human CD45⁺ chimerism was analyzed via flow cytometry. All donors engrafted successfully and robust long-term multi-lineage engraftment >16 weeks was seen in 19 out of 22 donors. To further investigate the long-term repopulating capacity, we performed secondary transplants. BM of 15 primary engrafted donors was harvested, hCD45⁺ were selected with immunobeads and transplanted into 1Gy irradiated NSG mice. Successful multi-lineage engraftment after secondary transfer was observed in 12 out of 15 donors in the peripheral blood (PB). Results of tissue engraftment are still outstanding for most experiments. So far they showed mean engraftment of human CD45⁺ cells of 6.16%+/-2.4% for donor 1 and 8.48%+/-0.74% for donor 2 in the BM. To compare CD and LD we set up two side-by-side transplantation experiments and intra-mouse competitive engraftment studies using HLA-mismatched donors. Using side-by-side experiments no significant difference in human CD45⁺ chimerism was detected in blood measured at weeks 6, 12 and 16 pos-transplant. The tissues showed a mean engraftment of human CD45⁺ cells of 87% +/- 9.6% vs 71% +/- 9.5% in spleen and 76% +/- 5.3% vs 78% +/- 4% (NS) in BM upon takedown 20 weeks post-transplant. For our competitive assays, we transplanted 2.5x10⁵ hCD34⁺ i.v. from LD and CD simultaneously into NSG mice (n=5 per donor pair) following 1Gy TBI. In total 8 CD and 3 LD were used, out of whom 4 CD reached higher engraftment than the LD.

Conclusion

Our data suggest that CD34⁺HSPC from CD retain their long-term multi-lineage engraftment capacity, reaching at least equipotent levels as HSPC from living donors. Based on our findings cadaveric bone marrow should be considered as HSPC source for HSCT.

Gu:Ossium Health: Current Employment, Current equity holder in private company, Current holder of stock options in a privately-held company. Johnstone:Ossium Health: Current Employment, Current equity holder in private company, Current holder of stock options in a privately-held company. Woods:Ossium Health: Current Employment, Current equity holder in private company, Current holder of stock options in a privately-held company. Mapara:Ossium Health: Consultancy, Research Funding.