Article

Cytotoxic T lymphocyte responses to transgene product, not adeno-associated viral capsid protein, limit transgene expression in mice

Department of Immunotherapy Research, Genzyme, Framingham, MA 01701, USA.
Human gene therapy (Impact Factor: 3.62). 11/2008; 20(1):11-20. DOI: 10.1089/hum.2008.055
Source: PubMed

ABSTRACT The use of adeno-associated viral (AAV) vectors for gene replacement therapy is currently being explored in several clinical indications. However, reports have suggested that input capsid proteins from AAV-2 vector particles may result in the stimulation of cytotoxic T lymphocyte (CTL) responses that can result in a loss of transduced cells. To explore the impact of anti-AAV CTLs on AAV-mediated transgene expression, both immunocompetent C57BL=6 mice and B cell-deficient muMT mice were immunized against the AAV2 capsid protein (Cap) and were injected intravenously with an AAV-2 vector encoding alpha-galactosidase (alpha-Gal). C57BL=6 mice, which developed both CTL and neutralizing antibody responses against Cap, failed to show any detectable alpha-Gal expression. In contrast, serum alpha-Gal levels comparable to those of naive mice were observed in muMT mice despite the presence of robust CTL activity against Cap, indicating that preexisting Cap-specific CTLs did not have any effect on the magnitude and duration of transgene expression. The same strategy was used to assess the impact of CTLs against the alpha-Gal transgene product on AAV-mediated gene delivery and persistence of transgene expression. Preimmunization of muMT mice with an Ad=alpha-Gal vector induced a robust CTL response to alpha-Gal. When these mice were injected with AAV2=alpha-Gal vector, initial levels of alpha-Gal expression were reduced by more than 1 log and became undetectable by 2 weeks postinjection. Overall, our results indicate that CTLs against the transgene product as opposed to AAV capsid protein are more likely to interfere with AAV transgene expression.

Download full-text

Full-text

Available from: Johanne M Kaplan, Aug 31, 2015
2 Followers
 · 
96 Views
 · 
44 Downloads
  • Source
    • "Transduction efficiency can be also be increased by mutating surface exposed tyrosine residues on the capsid, which is thought to reduce proteasomal degradation, increasing trafficking to the nucleus (Zhong et al., 2008; Markusic et al., 2010). Though a variety of mechanisms are involved in these studies, they, along with other studies in animals, are united by a common theme: in current murine models, functional CD8+ T cell infiltrates in AAV transduced tissues are primary directed against the transgene product rather than the capsid, while an antibody response is often observed to both potential immunogens (Siders et al., 2009). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Gene replacement therapy by in vivo delivery of adeno-associated virus (AAV) is attractive as a potential treatment for a variety of genetic disorders. However, while AAV has been used successfully in many models, other experiments in clinical trials and in animal models have been hampered by undesired responses from the immune system. Recent studies of AAV immunology have focused on the elimination of transgene-expressing cells by the adaptive immune system, yet the innate immune system also has a critical role, both in the initial response to the vector and in prompting a deleterious adaptive immune response. Responses to AAV vectors are primarily mediated by the TLR9-MyD88 pathway, which induces the production of pro-inflammatory cytokines by activating the NF-κB pathways and inducing type I IFN production; self-complementary AAV vectors enhance these inflammatory processes. Additionally, the alternative NF-κB pathway influences transgene expression in cells transduced by AAV. This review highlights these recent discoveries regarding innate immune responses to AAV and discusses strategies to ablate these potentially detrimental signaling pathways.
    Frontiers in Microbiology 09/2011; 2:194. DOI:10.3389/fmicb.2011.00194 · 3.94 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatic adeno-associated virus (AAV)-serotype 2-mediated gene transfer results in sustained transgene expression in experimental animals but not in human subjects. We hypothesized that loss of transgene expression in humans might be caused by immune memory mechanisms that become reactivated upon AAV vector transfer. Here, we tested the effect of immunological memory to AAV capsid on AAV-mediated gene transfer in a mouse model. Upon hepatic transfer of an AAV2 vector expressing human factor IX (hF.IX), mice immunized with adenovirus (Ad) vectors expressing AAV8 capsid before AAV2 transfer developed less circulating hF.IX and showed a gradual loss of hF.IX gene copies in liver cells as compared to control animals. This was not observed in mice immunized with an Ad vectors expressing AAV2 capsid before transfer of rAAV8-hF.IX vectors. The lower hF.IX expression was primarily linked to AAV-binding antibodies that lacked AAV-neutralizing activity in vitro rather than to AAV capsid-specific CD8(+) T cells.
    Molecular Therapy 05/2009; 17(7):1215-24. DOI:10.1038/mt.2009.79 · 6.43 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Viral vectors are potent gene delivery platforms used for the treatment of genetic and acquired diseases. However, just as viruses have evolved to infect cells efficiently, the immune system has evolved to fight off what it perceives as invading pathogens. Therefore, innate immunity and antigen-specific adaptive immune responses against vector-derived antigens reduce the efficacy and stability of in vivo gene transfer. In addition, a number of vectors are derived from parent viruses that humans encounter through natural infection, resulting in preexisting antibodies and possibly in memory responses against vector antigens. Similarly, antibody and T-cell responses may be directed against therapeutic gene products that often differ from the endogenous nonfunctional or absent protein that is being replaced. As details and mechanisms of such immune reactions are uncovered, novel strategies are being developed, and vectors are being specifically engineered to avoid, suppress or manipulate the response, ideally resulting in sustained expression and immune tolerance to the transgene product. This review provides a summary of our current knowledge of the interactions between the immune system adeno-associated virus, adenoviral and lentiviral vectors, and their transgene products.
    Gene therapy 02/2010; 17(2):294. DOI:10.1038/gt.2009.168 · 4.20 Impact Factor
Show more