A Conditional Fibroblast Growth Factor Receptor-1 Derivative Stimulates Extensive Self Renewal of Genetically Modified Primary Hematopoietic Cells.

Fibroblast growth factor receptor-1 (FGFR-1) signaling has been implicated in self renewal and proliferation of hematopoietic stem cells. We have studied the effect of a conditional FGFR-1 in genetically modified primary mouse bone marrow cells. The activity of the conditional FGFR-1 is dependent on the presence of a drug called a chemical inducer of dimerization (CID). For these studies we used a bicistronic vector encoding green fluorescence protein (GFP) as a marker and a fusion protein made up of the intracellular portion of FGFR-1 and the CID binding domain. To test the construct in vitro we plated transduced mouse bone marrow cells in 10% serum with or without the CID, AP20187 (100 nM). While the cells without CID died within two weeks, in the culture with CID the transduced, GFP marked cells proliferated. These hematopoietic cells have proliferated in vitro for over 30 weeks. This makes FGFr-1 only the second CID dependent receptor, other than mpl, that is able to support long-term self renewal of primary hematopoietic cells. By day 28 of culture cells had expanded from 10⁶ to 1.2x10¹² cells. The differentiation potential of these cells was tested by transplanting 8 million cells into each of 5 lethally irradiated (1050 cGy) congenic recipients. Four mice are alive 76 days post transplantation, with mean frequencies of GFP positive red cells, platelets, granulocytes, monocytes, B cells and T cells of 55%, 5%, 85%, 98%, 42% and 2%, respectively. These findings indicate that cells expanded under the influence of FGFR-1 signaling retain both lymphoid and myeloid repopulating ability. Additionally, CID-mediated activation of FGFR-1 induced a dramatic rise in genetically modfied red cells, granulocytes and monocytes in vivo. These findings point to a clear difference between CID induced activation of FGFR-1 compared to our previous studies using the thrombopoietin receptor, where effects on neutrophils and monocytes are modest and variable. The CID controlled activation of FGFR-1 may have utility for control of genetically modified hematopoietic cells.