GTMB 7 - Gene Therapy & Molecular Biology

Gene Therapy and Molecular Biology Vol 7, page 1831986). Prenatal diagnosis is usually performed bydetection of the CFTR mutation in placental tissue, fetalskin cells or blood after chorionic villus biopsy,amniocentesis or cordocentesis respectively.Submucosal gland development has been studied inthe rhesus monkey fetus (Plopper et al, 1986) and althoughnot characterized in the human fetal airways, submucosalgland progenitors have been identified in the human adultlung (Engelhardt et al, 1995). Gene transfer to a humanfetal lung xenograft model in SCID mice was efficientlyachieved using adenoviral vectors (Peault et al, 1994) andlong-term expression in the surface epithelial andsubmucosal gland cells was observed up to 4 weeks and 9months after administration of adeno-associated andlentiviral vectors respectively (Lim et al, 2002, 2003).The early disease manifestation and poor results fromgene therapy treatment of adults with CF has led toresearch on in utero gene therapy for this disease in animalmodels. Despite the multiorgan manifestation of CF, firstapproaches are directed towards gene delivery to the fetalairways, which has been achieved by intra-amnioticapplication and, in larger animals by intratracheal injection(see chapter IV). Other genetic diseases which couldbenefit from progress achieved in pulmonary genedelivery are α1-antitrypsin deficiency (Stecenko andBrigham 2003) and surfactant protein B deficiency (Coleet al, 2003).B. Metabolic disordersInherited inborn errors of metabolism can affect anumber of metabolic pathways. For example the ureacycle disorders are caused by defects in genes encodingenzymes or membrane transporters in ureagenesis. Theirprevalence is approximately 1:30,000 births and ornithinetranscarbamylase (OTC) deficiency is one of the mostsevere of these conditions (Summar and Tuchman, 2001).OTC deficiency is transmitted as a partially dominant X-linked trait. In patients with partial OTC deficiency, suchas hemizygous males and heterozygous females, the firstclinical episode is delayed for months or years with lesssevere hyperammonemia. However, patients withcomplete OTC deficiency present with life-threateninghyperammonemia within one week of birth and despitemedical therapy to reduce the ammonia levels, 50% of thechildren are dead by the age of 4, and of those surviving,the mean IQ is less than 50 (Maestri et al, 1999). Since theurea cycle is principally sited in the liver, gene therapydirected towards hepatocytes has the potential to correctthe metabolic abnormality. Indeed the success of orthopticliver transplantation in long-term treatment of thiscondition supports the concept (Lee and Goss, 2001).Adenoviral vectors have been shown to transientlycorrect OTC deficiency in the sparse fur murine modelafter neonatal and adult treatment (Stratford-Perricaudet etal, 1990; Ye et al, 1996). In a phase I human clinical trialin patients with partial OTC deficiency, adenoviral vectorsexpressing the human OTC–cDNA were administered.There was evidence of dose-related toxicity to theadenovirus and the last patient treated suffered a systemicinflammatory response syndrome that lead to his death(Raper et al, 2002).Because of its early onset, severity and presentdifficulties in postnatal gene therapy, OTC deficiency is aninteresting candidate for in utero gene application targetedto the fetal liver (see chapter IV). Prenatal diagnosis forOTC deficiency by detection of the genetic mutation infetal DNA is available in families with a known congenitalabnormality. In non-informative families, deficiency ofOTC enzyme can be detected in the fetal liver after liverbiopsy (Holzgreve and Golbus, 1986). Other seriousgenetic diseases that would primarily require hepatocytedirected gene transfer are amino acid disorders (e.g.phenylketonuria, tyrosinaemia), carbohydrate disorders(e.g. galactosaemia) and fatty acid oxidation disorders(e.g. long-chain acyl-CoA dehydrogenase deficiency)(Preece and Green 2002).C. Storage disordersThe lysosomal storage disorders (LSDs) are a groupof congenital deficiencies of one or more lysosomalenzymes. In mucopolysaccharidosis type VII (MPS typeVII) a deficiency of β-glucuronidase activity leads toaccumulation of undegraded glycosaminoglycans inlysosomes. Clinically, patients develop hepatosplenomegaly,mental and growth retardation, hearing and visiondefects, skeletal deformities and die of cardiac failure.Many of the LSDs present already during fetal life withhydrops fetalis and prenatal diagnosis can be performed bydetection of β-glucuronidase deficiency in chorionic villior fetal blood (Geipel et al, 2002). Although individuallyrare, as a group they occur in approximately 1 in 7500 livebirths and are one of the more prevalent groups ofinherited diseases in humans (Wraith, 2002). Bone marrowtransplantation and enzyme replacement therapy are beingdeveloped for many of the mucopolysaccharidoses.However, the short half-life of lysosomal enzymes in thecirculation means that patients need biweekly parenteraladministration which increases the risk of an immuneresponse to the infused enzyme. In addition, systemicallyadministered enzyme is unable to cross the blood-brainbarrier and can therefore not be used to treat centralnervous system disease manifestation.The LSDs are considered to be good candidates forgene therapy and the liver may be the ideal site for genetransfer. Newly synthesized lysosomal enzymes aresecreted into the systemic circulation and are recapturedby distant cells. Based on the observed enzyme levels inpatients with mild late-onset disease, the amount ofenzyme needed to correct the deficiency may only be 1-10% of normal levels (Cheng and Smith, 2003). Genetransfer to naturally occurring animal models of MPS typeVII has been investigated using adeno-associated virus(Daly et al, 1999), adenovirus (Kamata et al, 2003) andlentivirus (McCray Jr et al, 2001). Intravenousadministration of retroviral vectors containing canine β-glucuronidase to neonatal MPS type VII dogs preventedsome bone and joint abnormalities, corneal clouding andheart valve defects that commonly occur in this animal183