BBB Seminar: James C. Mulloy

James C. Mulloy
Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, OH, USA

Complex biological processes are best analyzed under native conditions in a host organism. After decades of experimentation and exhaustive searches for representative model organisms, the scientific field has currently settled on the laboratory mouse as the top surrogate species for human studies. The versatility, genetic accessibility and cost-efficiency of this species have propelled it to dominance in scientific research. Nevertheless, innumerable documented differences exist between mouse and human cells with regard to mechanisms of cell transformation, gene regulation, development, and various normal physiological processes. For this reason, investigators have pursued avenues of research that would permit experimentation on human cells under conditions that closely mimic the native in vivo situation. The use of immunodeficient mice to study human hematopoietic function has been pursued for decades. The advent of the NOD/SCID (NS) mouse was a key development that greatly improved the consistency and ease of xenograft experiments. However, this strain is hampered by several traits, including susceptibility to endogenous spontaneous lymphomas beginning as early as 5-6 months of age, which greatly limits long-term studies. Residual innate immune function from NK cells also limits engraftment of human hematopoietic stem cells (HSCs). Furthermore, established grafts decline over time, are markedly biased to the B cell lineage, and are characterized by a small myeloid component and a complete lack of T cell production. Numerous attempts to modify the NS mouse have been made in an effort to boost the potential of human xenografts. Currently the most successful strain modification has been the genetic inactivation of interleukin-2 receptor gamma (IL2RG). Two such strains exist, one that expresses a truncated IL2RG lacking the cytoplasmic domain (NOG) and a second with a full gene deletion (NSG). In both cases, these mice have a further reduced capacity for innate immunity as a result of diminished macrophage and NK activity. Importantly, both strains have a block in lymphoid cytokine signaling, which greatly suppresses the endogenous lymphoma development that plagues NS mice. NOG and NSG are reported to have a much longer lifespan. As a result, studies of long-term hematopoiesis that were not previously possible can now be performed in the xenograft setting. Both NSG and NOG are capable of supporting robust, long-term, B cell-dominated grafts that over time also include significant T cell populations. In light of these advances, NSG and NOG mice are currently the best available strains for xenograft studies of normal human hematopoiesis and leukemia. While NOG and NSG are similar, it has been proposed that the extracellular portion of IL2RG in NOG may retain some limited function and allow signaling through heterodimerization with a subset of its receptor partners. Indeed, a recent study has found slight advantages for NSG over NOG mice in their role as hosts for CD34+ cells, particularly at limiting cell doses of CD34+ cells.

We aim to develop and provide unique mouse strains for use in human xenograft experiments. We have recently generated a new strain of NS mouse, the NSGS, by cross breeding the NSG strain with the NSS (NS mice transgenic for the human cytokines SCF, GM-CSF and IL-3) strain. This new strain promotes increased human myelopoiesis relative to the NSG mouse, and unexpectedly, promotes more rapid human T-cell development. This strain also shows improvements in the function of the engrafted human immune system and provides a new model of immune-mediated anemia. Our goal is to obtain a mouse with all the advantages of the NSG with an increased myeloid potential that could promote better acute myeloid leukemia (AML) xenografts as well as improved human hematopoiesis, in order to study malignant and normal human hematopoiesis. We have recently reported that when AML cell lines and patient samples were injected into matched cohorts of NSGS and NSG mice, the NSGS cohort produced more robust grafts more frequently. Some patient samples that failed to engraft NSG mice yielded significant grafts in NSGS mice, demonstrating a strict dependence of a subset of these samples on myeloid cytokine signals and revealing an increased sensitivity for human AML SCID-repopulating cells (SRC) in NSGS. We also showed that a pre-leukemia cell model that we use in the lab, whereby we express the leukemia oncogene AML1-ETO in normal human CD34+ cells and establish long-term cytokine-dependent cultures, showed significantly improved engraftment in NSGS relative to NSG. This gives us hope that other types of pre-malignant cells, including myelodysplastic syndrome (MDS) and myeloproliferative neoplasm (MPN) samples, will additionally demonstrate improved engraftment in NSGS or in improved models that are developing.

Chair: Emmet Mc Cormack, Institute of Medicine