Abstract 5086: De-differentiation of CD45+ hematopoietic cells to CD45- stromal cells in vitro

Abstract

Abstract Bone marrow (BM) consists of both hematopoietic cells and non-hematopoietic stromal cells, which support hematopoiesis. Recent reports have suggested that some non-hematopoietic BM cells, such as multipotent adult progenitor cells, can produce hematopoietic stem and/or progenitor cells. It has been speculated that non-hematopoietic BM stromal cells may generate myeloid malignancy following conversion of non-hematopoietic cells to hematopoietic cells. To study the conversion of non-hematopoietic cells (CD45-) to hematopoietic cells (CD45+), we used the NUP98-HOXD13 (NHD13) mouse model of myelodysplastic syndrome (MDS). NHD13 mice develop MDS which mimics the human condition in terms of peripheral blood (PB) cytopenias, ineffective hematopoiesis, and transformation to acute myeloid leukemia (AML). Two cell lines (251 and 63B) established from NHD13 mice with AML grow as a combination of adherent and suspension cells. Upon sub-culture of the suspension cells, the suspension cells become adherent. Although the suspension cells are almost entirely (>95%) CD45+, the adherent cells are only 50% CD45+, suggesting that a substantial fraction of the CD45+ cells became CD45-. In conjunction with acquisition of CD45, the cell lines acquire Mac-1, CD16/32 and F4/80 positivity, which are lost upon loss of CD45. In order to rule out the possibility that a small fraction of CD45- cells in the suspension phase (99.85%) population of CD45+ cells. After 14 days of sub-culture, FACS revealed that 46% of the cells were now CD45-, demonstrating conversion of CD45+ cells to CD45- cells. To further explore this finding, CD45+ and CD45- populations were purified and expression of the NHD13 transgene (driven by the pan-hematopoietic Vav1 promoter) was determined by Q-PCR to be 100-fold higher in the CD45+ fraction compared to the CD45- fraction. These results demonstrate that the cell lines can cycle between non-hematopoietic and hematopoietic cells. To determine if the cell lines have retained malignant potential, we transplanted irradiated recipients via intravenous injection. One of 3 recipients transplanted with 1×10^6 cells showed low level engraftment [1.7% of peripheral blood (PB) cells] at week 24 post-transplant. Remarkably, this mouse was leukemic 2 weeks later, with splenomegaly, anemia, and monocytosis (8.57 K/uL), and donor engraftment of 17-53% in the PB, BM, and spleen. To improve engraftment, we used intra-femoral injection, which resulted in consistent engraftment at 6 weeks in the PB. This experiment is ongoing; at 9 weeks one recipient has developed lymphopenia and anemia, suggesting the possibility of malignant transformation. Taken together, these findings suggest that malignant cells can cycle between stromal and hematopoietic cells, and support the hypothesis that non-hematopoietic BM cells may be an important part of the malignant clone in patients with MDS and AML. Citation Format: Yang Jo Chung, Suntae Kim, Peter Aplan. De-differentiation of CD45+ hematopoietic cells to CD45- stromal cells in vitro. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5086. doi:10.1158/1538-7445.AM2015-5086