PRECLINICAL STUDIES OF LYMPHOCYTE GENE-THERAPY FOR MILD HUNTER SYNDROME (MUCOPOLYSACCHARIDOSIS TYPE-II)

To explore the feasibility of ex vivo lymphocyte gene therapy for mild Hunter syndrome (mucopolysaccharidosis type II), we evaluated retrovi rus-mediated gene transfer of the iduronate-2-sulfatase (IDS) coding s equence into peripheral blood lymphocytes from enzyme-deficient indivi duals (PBL(MPS)). Moloney murine leukemia virus-derived retroviral vec tors were constructed by inserting the IDS cDNA under transcriptional regulation of the long terminal repeat (LTR) (in vector L2SN) or the c ytomegalovirus (CMV) early promoter (vector LNC2). High-titer virus-pr oducer cells were generated using amphotropic PA317 packaging cells. A fter 3 days of in vitro stimulation of T lymphocytes with anti-CD3 ant ibody and interleukin-2 (IL-2), PBL(MPS) were transduced once on each of the next 3 days. Seven to 21 days later, cultured PBL(MPS) were eva luated for gene transfer and IDS specific activity. Heterogeneous popu lations of L2SN-transduced PBL(MPS) had high levels of IDS enzyme acti vity (456 U/mg per hr +/- SD 292) despite a gene transfer efficiency o f 5% or less. Owing to overexpression of IDS in that percentage of PBL (MPS) successfully transduced, IDS activity was increased above the de ficiency found in patients with Hunter syndrome (<20 U/mg per hr) to a level comparable with that of normal individuals (mean activity of un cultured normal leukocytes 807 U/mg per hr; SD 252). Reduced (SO4)-S-3 5-glucosaminoglycan (GAG) accumulation was observed in PBL(MPS) that h ad been transduced with L2SN, or when PBL(MPS) were grown in medium th at had been ''conditioned'' by growth of L2SN-transduced cells. This l atter result indicated that metabolic cross-correction occurred by mea ns of intercellular enzyme transfer. These studies of retrovirus-media ted expression and metabolic correction, finding near-normal levels of IDS in cultured PBL(MPS) and metabolic correction, demonstrate the po tential for treatment of mild, nonneuropathic Hunter syndrome by means of ex vivo lymphocyte gene therapy.