Humanα1-Antitrypsin Gene Transfer toIn VivoMouse Hepatocytes

Abstract

The in vivo gene transfer to mouse hepatocytes of pTG 7101, a plasmid containing the full-length gene encoding human alpha 1-antitrypsin (alpha 1-AT) DNA, has been studied by iv administration of recombinant DNA (100 ng/mouse) encapsulated in large and small liposomes. Our results from immunohistochemical liver sections and cytophotometric analysis of hepatocyte chromophore absorbance indicate that human alpha 1-AT was expressed in liver parenchymal cells from mice treated (48 hr before) with DNA encapsulated in small liposomes, and this effect remained for at least 2 weeks. In contrast, the efficiency was greatly limited when large liposomes were used as a vehicle for gene transfer. Additional experiments were performed to study using an ELISA procedure the presence in mouse plasma of human alpha 1-AT from mice treated with encapsulated plasmid in small liposomes or small empty liposomes plus free DNA. According to the immunohistochemical data, the results indicate that detectable alpha 1-AT can only be observed in plasma from mice treated with encapsulated plasmid in small liposomes.

Long-term therapeutic levels of human alpha-1 antitrypsin in plasma after hydrodynamic injection of nonviral DNA

“…To confirm this observation, we performed additional dose-response experiments. The results support our early suggestions that genomic DNA offers advantages to mediate in vivo hAAT gene transfer, 6,8,19,23 and that the final efficacy is limited by the DNA carrier. 21, 22 Now we demonstrate that naked nonviral DNA containing genomic hAAT can mediate long-term therapeutic levels of hAAT protein in mouse plasma.…”

“…Since human AAT deficiency may be a good candidate for clinical and experimental gene therapy strategies, a broad variety of systems have been developed to transfer genes into the liver, resulting in detectable serum levels of hAAT in animal models. [3][4][5][6][7][8] Adenoviral vectors, 9,10 and more recently recombinant adeno-associated viral vectors, 11 have shown the ability to mediate therapeutic serum concentrations of hAAT. However, their usefulness in gene therapy could be limited by the pathogenic risks involved.…”

Increased receptor-mediated gene delivery to the liver by protamine-enhanced-asialofetuin-lipoplexes

“…It is already known that delivery to parenchymal cells in vivo is favored by small liposomes. 6 For most vectors, high efficiency of transfection correlates with a large excess of cationic charges, [38][39][40] however, this large excess in vivo facilitates non-specific interactions with many undesired elements such as extracellular matrix and negatively charged serum components. Taking this into account, the first aim of this work was to optimize the size and the zeta potential of AF-lipoplexes for maximum uptake by the liver after systemic administration.…”

“…Although some of the virus-mediated gene transfer systems have been found to be quite effective, their usefulness is limited, given that they induce an immune response, leading to the rapid rejection of transduced cells. To overcome this problem, artificial, non-viral gene delivery systems are being developed, including small liposomes, [6][7][8] particle bombardment, 9 gene gun, 10 electroporation, 11 chylomicron remnants 12 and cationic polymers. 13 Successful transfer and expression in the liver can also be achieved by systemic administration of naked plasmid using a hydrodynamic-based procedure.…”

Progress with Recombinant Adeno-Associated Virus Vectors for Gene Therapy of Alpha-1 Antitrypsin Deficiency

“…[6][7][8][9][10][11][12][13] In this review we will focus on the development of gene therapy for AAT-associated lung disease through elevation of human AAT serum levels (hAAT = M-AAT). In the final section we will discuss potential options for treating AATassociated liver disease in addition to the lung disease.…”