EVALUATION OF EXPRESSION OF TRANSFERRED GENES IN DIFFERENTIATING MYELOID CELLS - EXPRESSION OF HUMAN GLUCOCEREBROSIDASE IN MURINE MACROPHAGES

The retroviral vector LGSN, in which the human glucocerebrosidase (GC) cDNA is driven by the Moloney murine leukemia virus (MoMLV) long term inal repeat (LTR), was tested for expression in the murine myelomonocy tic leukemia cell line M1 before and after induction of differentiatio n with interleukin-6 (IL-6). Southern analysis of the seven transduced clones selected for neomycin resistance in Geneticin (G-418 sulfate) demonstrated one to eight copies of intact provirus with rearrangement s in only two clones. Absolute levels of human GC RNA and protein incr eased with increased copy numbers of provirus in the clones. Upon indu ction with IL-6 of the seven transduced clones to the macrophage pheno type, there was no significant change, overall, in RNA levels but some increase in human GC protein levels could be detected. Although this was the average trend, considerable clonal variation in RNA and protei n levels was observed upon induction. Transduction of the M1 cells did not interfere with the ability of the cells to differentiate from bla sts to macrophages as seen by the appearance of membrane receptors for the constant region of immunoglobulins (FcgammaRI) and, lysozyme prod uction in the differentiated M1 cells. Thus, the M1 cell line can be u sed for testing retroviral vector expression in myeloid lineages at ea rly and late stages of differentiation. This rapid in vitro testing of potential retroviral vectors will be beneficial for gene therapy of d isorders that affect differentiated macrophages such as Gaucher's dise ase.