Complete allogeneic hematopoietic chimerism achieved by a combined strategy of in utero hematopoietic stem cell transplantation and postnatal donor lymphocyte infusion

In utero hematopoietic stem cell transplantation (IUHSCTx) can achieve mixed hematopoietic chimerism and donorspecific tolerance without cytoreductive conditioning or immunosuppression. The primary limitation to the clinical application of IUHSCTx has been minimal donor cell engraftment, well below therapeutic levels for most target diseases. Donor lymphocyte infusion (DLI) has been used in postnatal circumstances of mixed chimerism as targeted immunotherapy to achieve a graft-versus-hematopoietic effect and to increase levels of donor cell engraftment. In this report we demonstrate in the murine model that a combined approach of IUHSCTx followed by postnatal DLI can convert low-level, mixed hematopoietic chimerism to complete donor chimerism across full major histocompatibility complex barriers with minimal risk for graftversus-host disease (GVHD)

Ionizable lipid nanoparticles for in utero mRNA delivery.

Clinical advances enable the prenatal diagnosis of genetic diseases that are candidates for gene and enzyme therapies such as messenger RNA (mRNA)-mediated protein replacement. Prenatal mRNA therapies can treat disease before the onset of irreversible pathology with high therapeutic efficacy and safety due to the small fetal size, immature immune system, and abundance of progenitor cells. However, the development of nonviral platforms for prenatal delivery is nascent. We developed a library of ionizable lipid nanoparticles (LNPs) for in utero mRNA delivery to mouse fetuses. We screened LNPs for luciferase mRNA delivery and identified formulations that accumulate within fetal livers, lungs, and intestines with higher efficiency and safety compared to benchmark delivery systems, DLin-MC3-DMA and jetPEI. We demonstrate that LNPs can deliver mRNAs to induce hepatic production of therapeutic secreted proteins. These LNPs may provide a platform for in utero mRNA delivery for protein replacement and gene editing

The Pediatric Cell Atlas:Defining the Growth Phase of Human Development at Single-Cell Resolution

Single-cell gene expression analyses of mammalian tissues have uncovered profound stage-specific molecular regulatory phenomena that have changed the understanding of unique cell types and signaling pathways critical for lineage determination, morphogenesis, and growth. We discuss here the case for a Pediatric Cell Atlas as part of the Human Cell Atlas consortium to provide single-cell profiles and spatial characterization of gene expression across human tissues and organs. Such data will complement adult and developmentally focused HCA projects to provide a rich cytogenomic framework for understanding not only pediatric health and disease but also environmental and genetic impacts across the human lifespan

In utero hematopoietic cell transplantation: Induction of donor specific immune tolerance and postnatal transplants

In utero hematopoietic cell transplantation (IUHCT) is a nonmyeloablative nonimmunosuppressive transplant approach that allows for donor cell engraftment across immunologic barriers. Successful engraftment is associated with donor-specific tolerance. IUHCT has the potential to treat a large number of congenital hematologic, immunologic and genetic diseases either by achieving high enough engraftment levels following a single IUHCT or by inducing donor specific tolerance to allow for nontoxic same-donor postnatal transplants. This review evaluates donor specific tolerance induction achieved by IUHCT. Specifically it addresses the need to achieve threshold levels of donor cell engraftment following IUHCT to consistently obtain immunologic tolerance. The mechanisms of tolerance induction including partial deletion of donor reactive host T cells by direct and indirect antigen presentation and the role of regulatory T cells in maintaining tolerance are reviewed. Finally, this review highlights the promising clinical potential of in utero tolerance induction to provide a platform on which postnatal cellular and organ transplants can be performed without myeloablative or immunosuppressive conditioning

Complete allogeneic hematopoietic chimerism achieved by in utero hematopoietic cell transplantation and cotransplantation of LLME-treated, MHC-sensitized donor lymphocytes

Objective. In utero hematopoietic cell transplantation (IUHCT) typically achieves low-level mixed hematopoietic chimerism. However, the goal of IUHCT is to achieve therapeutic levels of chimerism. We hypothesized that prenatal adoptive immunotherapy might achieve high-level donor chimerism after IUHCT. Materials and Methods. BALB/CE15 fetal mice were transplanted with a mixture of C57BL/6 (B6) T-cell-depleted bone marrow (TCD BM) cells and splenocytes from B6 mice presensitized to BALB/C alloantigen. The splenocytes were preincubated in L-leucyl-L-leucine methyl ester (LLME), to minimize graft vs host disease (GVHD). Recipients were followed after birth for donor cell chimerism and GVHD. Results. Full donor hematopoietic chimerism following a single prenatal transplant was achieved in seven transplanted animals. Fully chimeric animals were healthy, without evidence of GVHD, and maintained their engraftment for the duration of the study (48 weeks). However, the addition of presensitized LLME-treated cells decreased survival until weaning relative to TCD BM alone, suggesting that some animals were lost to acute GVHD. Surviving chimeric animals demonstrated increased frequencies of T-regulatory cell populations in their spleen and BM, suggesting that they had successfully suppressed GVHD, allowing survival. Conclusions. This study represents proof in principle that prenatal immunotherapeutic strategies may achieve complete hematopoietic engraftment across full MHC barriers when combined with IUHCT. However, strategies with greater hematopoietic specificity must be developed prior to consideration of clinical application. © 2004 International Society for Experimental Hematology. Published by Elsevier Inc

High-level allogeneic chimerism achieved by prenatal tolerance induction and postnatal nonmyeloablative bone marrow transplantation

Clinical application of allogeneic bone marrow transplantation (BMT) has been limited by toxicity related to cytoreductive conditioning and immune response. In utero hematopoietic stem cell transplantation (IUHSCT) is a nonablative approach that achieves mixed chimerism and donor-specific tolerance but has been limited by minimal engraftment. We hypothesized that mixed chimerism achieved by IUHSCT could be enhanced after birth by nonmyeloablative total body irradiation (TBI) followed by same-donor BMT. To test this hypothesis, mixed chimerism was created by IUHSCT in a major histocompatibility complex-mismatched strain combination. After birth, chimeric animals received nonmyeloablative TBI followed by transplantation of donor congenic bone marrow cells. Our results show that: (1) low-level chimerism after IUHSCT can be enhanced to high-level chimerism by this strategy; (2) enhancement of chimerism is dependent on dose of TBI; (3) the mechanism of TBI enhancement is via a transient competitive advantage for nonirradiated hematopoietic stem cells; (4) engraftment observed in the tolerant, fully allogeneic IUHSC transplant recipient is equivalent to a congenic recipient; and (5) host-reactive donor lymphocytes are deleted with no evidence of graft-versus-host disease. This study supports the concept of prenatal tolerance induction to facilitate nonmyeloablative postnatal strategies for cellular therapy. If clinically applicable, such an approach could dramatically expand the application of IUHSCT. © 2002 by The American Society of Hematology