INTRAPERITONEAL INJECTION OF GENETICALLY-MODIFIED, HUMAN MESOTHELIAL CELLS FOR SYSTEMIC GENE-THERAPY

An ideal cell type for ex vivo gene therapy should be easy to biopsy, propagate, and genetically engineer in culture, should be transplantab le using simple procedures, and should express therapeutic proteins at useful levels, The mesothelial cell appears to satisfy these criteria , Several thousand proliferative mesothelial cells were present in typ ical specimens of nonpathologic human peritoneal fluid obtained by nee dle aspiration, These divided rapidly in a specialized medium to yield pure cultures of similar to 10(7) cells within 2 weeks, The replicati ve lifespan of mesothelial cells cultured from adults was similar to 4 2-52 population doublings, permitting expansion and cryopreservation o f a lifetime supply of autologous cells from one fluid sample. Cells t ransduced with a human growth hormone (hGH) adenoviral vector secreted 100-300 mu g of hGH/10(6) cells per day for at least 6 weeks in cultu re when maintained at quiescence. Intraperitoneal injection of transdu ced cells into athymic mice resulted in rapid systemic delivery of hGH , with peak plasma levels of 0.1-1 mu g/ml declining over 3 weeks to < 1 ng/ml, Mice receiving a second injection of engineered cells display ed the same plasma hGH levers and duration as naive mice, Cells labele d with a beta-galactosidase vector were identifiable by in situ enzyma tic staining as clusters attached to peritoneal surfaces at multiple s ites for at least 19 days after injection, Cells serially passaged thr ough about three-quarters of their lifespan before transduction and in jection mere as effective at hGH delivery as earlier-passage cells, Th ese results indicate the clinical potential for ex vivo gene therapy u sing mesothelial cells.