Myelodysplastic/Myeloproliferative Neoplasm in Crebbp+/− Mice Is a Transplantable Disease in Which Multiple Hematopoietic Cell Populations Are Involved, As Well As the Microenvironment

Abstract 1705

Myelodysplastic/myeloproliferative neoplasms (MDS/MPNs) are myeloid malignancies that display features of both MDS and MPN, but cannot be properly assigned to either MDS or MPN. It is currently not known whether it originates from the hematopoietic stem cell (HSC) compartment (like MDS), from a more committed myeloid progenitor population, or a combination thereof. Fifteen to 40% of MDS/MPN patients develop acute myeloid leukemia (AML); whether the transformation occurs in a particular cell population is also unknown. We previously demonstrated that mice heterozygous for the CREB binding protein gene (Crebbp) develop MDS/MPN at 9–12 months of age and ∼40% of them progress to develop a hematologic malignancy. Thus, Crebbp^+/− mice are an excellent model to address the before mentioned questions, which is important for the development of better strategies to treat MDS/MPN.

For this purpose, we harvested and combined bone marrow from 1.5-year old Crebbp^+/− mice (10 donors per experiment, thereby ensuring that the marrow of ∼4 donors harbored malignant hematopoietic cells) and transplanted it into lethally irradiated, wild-type recipients. Groups of mice either received unfractionated whole bone marrow (WBM) or populations purified by fluorescence-activated cell sorting. Naive Crebbp^+/− mice had demonstrated functional and/or quantitative abnormalities in long-term and short-term HSCs, common myeloid progenitors (CMPs) and granulocyte/macrophage progenitors (GMPs) and we therefore focused on these populations. All transplant recipients also received unfractionated wild-type “helper cells” to increase survival. Mice were closely monitored and those suspected of having developed a hematopoietic disease were sacrificed and their hematopoietic system analyzed. Four independent experiments were performed and data were combined for analysis. Among the 18 recipients who received Crebbp^+/− WBM, 8 recipients (44%) developed an early-onset AML with myelofibrosis, 2–7 months after the transplant, which was not preceded by MDS. The other 10 recipients (56%) developed MDS/MPN, 12–18 months after the transplant. These mice displayed ineffective hematopoiesis, evidenced by a normocellular bone marrow, significant leukopenia, and trilineage dysplasia. One of these 10 Crebbp^+/− WBM recipients that developed MDS/MPN subsequently progressed to AML. In contrast, none of the 15 recipients of Crebbp^+/− HSCs (defined as Lin⁻Sca-1⁺c-Kit⁺⁺ (LSK) cells, including long-term and short-term progenitors, as well as lymphoid-restricted progenitors) developed early-onset AML. Instead, 1 developed MDS/MPN while the remainder developed MDS by 11–17 months after the transplant, with one of them progressing to a disease resembling human mature T-cell leukemia. Transplantation of Crebbp^+/− CMPs and GMPs also failed to cause early-onset AML and, as expected, gave rise to extremely low long-term reconstitution. Thus, these mice were mostly reconstituted by the co-transplanted wild-type “helper cells”. However, unexpectedly, 9 out of 24 (38%) showed <10% dysplastic cells in 1 or more lineages, while 4 (17%) developed overt MDS, i.e. >10% dysplastic cells and 2 (8%) developed MPD or AML with myelofibrosis. Control mice, i.e., recipients of wild-type BM cells remained healthy for the duration of the experiments.

The results of these transplantation experiments show that in this mouse model, MDS/MPN is transplantable. However, it requires transplantation of WBM, since the transplantation of LSK cells resulted in MDS, suggesting that the microenvironment may play a crucial role in the etiology of MDS/MPN. This notion is in concordance with our previous study, demonstrating that Crebbp^+/− mice transplanted with wild-type cells developed MPD that originated from the transplanted wild-type cells. This notion is further supported by the outcome of the CMP and GMP transplantation experiments, suggesting that abnormal myeloid progenitors are also important factors for MDS/MPN disease development. Moreover, malignant transformation seems to occur in a non-LSK cell that is more differentiated than CMPs and GMPs. Alternatively, malignant transformation requires all hematopoietic and non-hematopoietic cells to be present, again suggesting that MDS/MPN is a complex disease where both the hematopoietic compartment and its bone marrow microenvironment are affected.