617. A Synthetic Promoter Drives Macrophage Expression of ApoE and Ameliorates Atherosclerosis

Abstract

Macrophages, derived from hematopoietic stem cells (HSC), play a key role in atherosclerosis. To treat atherosclerosis, we plan to alter expression of relevant genes in macrophages by ex vivo transduction of HSC with lentiviral vectors carrying a macrophage-specific promoter. Myeloid promoter cis-elements were randomly ligated and the resultant DNA inserted into a reporter vector upstream of a myeloid basal promoter. Luciferase activity of some synthetic promoters was 10-200-fold over that of the CSF1R macrophage promoter in Thp-1 and other macrophage cell lines, but quite low in non-macrophage cells compared with the CMV promoter. Mouse bone marrow was collected, transduced ex vivo with lentiviral vectors expressing GFP driven either by a synthetic promoter (SP-GFP) or CMV promoter (CMV-GFP), and transplanted into lethally irradiated recipients. At various time points (1.5, 4, 8, 11, 15 months) post-transplant, peripheral blood analyzed by FACS and GFP expression was quite stable over time. SP-GFP expression was detected in most CD11b+ leukocytes, but only in a small fraction of CD11b- cells and not at all in erythrocytes, while CMV-GFP expression was observed ubiquitously. To test the promoters in macrophage gene therapy for atherosclerosis, we transduced apoE-/- BM cells with a lentiviral vector encoding human apoE (apoE) and transplanted them into apoE-/- mice. Comparable levels of apoE were measured in culture media of peritoneal macrophages from either SP-apoE or CMV-apoE recipient mice. However, serum apoE of SP-apoE recipients was only one fourth of that of CMV-apoE, implying that SP activity was restricted to the monocyte/macrophage lineage. Macrophage expression of apoE from 10 to 26 weeks of age significantly reduced atherosclerotic lesions in recipient mice. In conclusion, the combination of the synthetic macrophage promoter, lentiviral vectors, and bone marrow stem cell transplantation ameliorates atherogenesis and leads to a novel treatment of this disease.