Gary J. Kurtzman

Somatic gene therapy has been proposed as a means to achieve systemic delivery of therapeutic proteins. However, there is limited evidence that current methods of gene delivery can practically achieve this goal. In this study, we demonstrate that, following a single intramuscular administration of a recombinant adeno-associated virus (rAAV) vector containing the beta-galactosidase (AAV-lacZ) gene into adult BALB/c mice, protein expression was detected in myofibers for at least 32 weeks. A single intramuscular administration of an AAV vector containing a gene for human erythropoietin (AAV-Epo) into mice resulted in dose-dependent secretion of erythropoietin and corresponding increases in red blood cell production that persisted for up to 40 weeks. Primary human myotubes transduced in vitro with the AAV-Epo vector also showed dose-dependent production of Epo. These results demonstrate that rAAV vectors are able to transduce skeletal muscle and are capable of achieving sustained expression and systemic delivery of a therapeutic protein following a single intramuscular administration. Gene therapy using AAV vectors may provide a practical strategy for the treatment of inherited and acquired protein deficiencies.

The B19 parvovirus, which was serendipitously discovered in serum from blood-bank donors in 1975,1 has been associated with several human illnesses. B19 is the etiologic agent of the common childhood exanthem called fifth disease.2 Adults with fifth disease more commonly present with polyarthralgia than skin eruption, and B19 infection has frequently been detected in patients presenting with acute joint symptoms.3 , 4 In patients with underlying hemolysis, B19 infection is the cause of transient aplastic crisis, in which cessation of erythropoiesis leads to absolute reticulocytopenia and abrupt worsening of anemia.5 , 6 In utero infection with B19 has been associated with spontaneous abortion.7 8 9 Laboratory . . .

PARVOVIRUS B19 is the etiologic agent of transient aplastic crisis, an acute episode of bone marrow failure in persons with underlying hemolysis.1 , 2 In tissue-culture studies, parvovirus B19 infects3 and replicates4 in erythroid progenitor cells. The pathophysiologic basis of acute marrow failure in patients is viral tropism and cytotoxicity for erythroid progenitor cells.5 , 6 In experimental7 and natural8 infections, the infection is terminated with the development of specific antibodies, which neutralize the activity of the virus in vitro.9 In the immunocompromised host who is unable to produce neutralizing antibody,10 an infection with parvovirus B19 can persist and cause chronic bone marrow failure, . . .

B19 parvovirus has been shown to persist in some immunocompromised patients, and treatment with specific antibodies can lead to decreased quantities of circulating virus and hematologic improvement. A defective immune response to B19 parvovirus in these patients was shown by comparison of results using a capture RIA and immunoblotting. In normal individuals, examination of paired sera showed that the dominant humoral immune response during early convalescence was to the virus major capsid protein (58 kD) and during late convalescence to the minor capsid species (83 kD). In patients with persistent parvovirus infection, variable titers against intact particles were detected by RIA, but the sera from these patients had minimal or no IgG to capsid proteins determined by Western analysis. Competition experiments suggested that this discrepancy was not explicable on the basis of immune complex formation alone and that these patients may have a qualitative abnormality in antibody binding to virus. In neutralization experiments, in which erythroid colony formation in vitro was used as an assay of parvovirus activity, sera from patients with poor reactivity on immunoblotting were also inadequate in inhibiting viral infectivity. A cellular response to purified B19 parvovirus could not be demonstrated using proliferation assays and PBMC from individuals with serologic evidence of exposure to virus. These results suggest that production of neutralizing antibody to capsid protein plays a major role in limiting parvovirus infection in man.