'Supercharged cells' for delivery of recombinant human iduronate-2-sulfatase

Expression of iduronate-2-sulfatase (IDS) from three different promoters in four retroviral vectors was studied in peripheral blood lymphocytes from patients with Hunter syndrome (PBL(MPS)), i.e., the LTR in vectors L2SN and L2, avian β-actin promoter in LB2, and the CMV early promoter in LNC2. PBL(MPS) were exposed to packaging cell supenatant resulting in transduction frequencies ranging 10- gold from 5 to 49%. Surprisingly, IDS activities were equally high in all transduced lymphocyte populations: 515 U/mg/h in PBL(MPS)-L2SN, 734 in PBL(MPS)-LB2, 352 in PBL(MPS)-L2, and 389 in PBL(MPS)-LNC2 compared to controls (<10 in PBL(MPS)-LXSN or PBL(MPS)). The half-life of endocytosed IDS in PBL(MPS) was 1.9 days. However, the level of lymphocyte IDS activity from proviral expression was found to be only a fraction of the total, a large portion being derived from reuptake of enzyme from murine packaging cells, i.e., a 'second source' of enzyme. Therefore, measurement of transgene lysosomal enzyme soon after exposure of target cells to vector supernatant may yield a gross overestimate of long-term transgene expression by transduced cells. Nevertheless, patient fibroblasts cocultured with transduced PBL(MPS) had reduced ³⁵SO₄-GAG accumulation, levels similar to those of normal fibroblasts. These studies revealed a broadly applicable phenomenon: cells can be charged with a lysosomal enzyme to levels much higher than those found in nature. By 'supercharging' cells with a lysosomal protein (or other molecule bearing the mannose-6- phosphate ligand), such cells may be exploited as vehicles for systemic delivery of therapeutic or diagnostic agents. (C) Academic Press.