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Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

ABSTRACT: Immune responses against vectors or encoded transgenes can impose limitations on gene therapy. We demonstrated that tetracycline-regulated high-capacity adenoviral vectors (HC-Ads) sustain regulated transgene expression in the brain even in the presence of systemic pre-existing immune responses against adenoviruses. In this study we assessed whether systemic pre-existing immune responses against the transgene products, i.e., beta-Gal or the tetracycline-dependent (TetON) regulatory transcription factors (rtTA2(S)M2 and the tTS(Kid)), affect transgene expression levels and the safety profile of HC-Ads in the brain. We pre-immunized mice with plasmids encoding the TetON switch expressing rtTA2(S)M2 and the tTS(Kid) or beta-Gal. HC-Ads expressing beta-Gal under the control of the TetON switch were then injected into the striatum. We assessed levels and distribution of beta-Gal expression, and evaluated local inflammation and neuropathological changes. We found that systemic immunity against beta-Gal, but not against the TetON switch, led to inflammation and reduction of transgene expression in the striatum. Therefore, the regulatory TetON switch appears to be safe to use, and capable of sustaining transgene expression in the brain even in the presence of an immune response against its components. Systemic immunity against the transgene had the effect of curtailing its expression, thereby affecting the efficacy and safety of gene delivery to the brain. This factor should be considered when developing gene therapies for neurological use.

SUBMITTER: Xiong W

PROVIDER: S-EPMC2703652 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

Xiong Weidong W Candolfi Marianela M Kroeger Kurt M KM Puntel Mariana M Mondkar Sonali S Larocque Daniel D Liu Chunyan C Curtin James F JF Palmer Donna D Ng Philip P Lowenstein Pedro R PR Castro Maria G MG

Molecular therapy : the journal of the American Society of Gene Therapy 20080108 2

Immune responses against vectors or encoded transgenes can impose limitations on gene therapy. We demonstrated that tetracycline-regulated high-capacity adenoviral vectors (HC-Ads) sustain regulated transgene expression in the brain even in the presence of systemic pre-existing immune responses against adenoviruses. In this study we assessed whether systemic pre-existing immune responses against the transgene products, i.e., beta-Gal or the tetracycline-dependent (TetON) regulatory transcription ...[more]

PMID: 18180781

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Project description:BackgroundTrypanosoma cruzi is the causative agent of Chagas disease. Chagas disease is an endemic infection that affects over 8 million people throughout Latin America and now has become a global challenge. The current pharmacological treatment of patients is unsuccessful in most cases, highly toxic, and no vaccines are available. The results of inadequate treatment could lead to heart failure resulting in death. Therefore, a vaccine that elicits neutralizing antibodies mediated by cell-mediated immune responses and protection against Chagas disease is necessary.Methodology/principal findingsThe "antigen capsid-incorporation" strategy is based upon the display of the T. cruzi epitope as an integral component of the adenovirus' capsid rather than an encoded transgene. This strategy is predicted to induce a robust humoral immune response to the presented antigen, similar to the response provoked by native Ad capsid proteins. The antigen chosen was T. cruzi gp83, a ligand that is used by T. cruzi to attach to host cells to initiate infection. The gp83 epitope, recognized by the neutralizing MAb 4A4, along with His6 were incorporated into the Ad serotype 5 (Ad5) vector to generate the vector Ad5-HVR1-gp83-18 (Ad5-gp83). This vector was evaluated by molecular and immunological analyses. Vectors were injected to elicit immune responses against gp83 in mouse models. Our findings indicate that mice immunized with the vector Ad5-gp83 and challenged with a lethal dose of T. cruzi trypomastigotes confer strong immunoprotection with significant reduction in parasitemia levels, increased survival rate and induction of neutralizing antibodies.Conclusions/significanceThis data demonstrates that immunization with adenovirus containing capsid-incorporated T. cruzi antigen elicits a significant anti-gp83-specific response in two different mouse models, and protection against T. cruzi infection by eliciting neutralizing antibodies mediated by cell-mediated immune responses, as evidenced by the production of several Ig isotypes. Taken together, these novel results show that the recombinant Ad5 presenting T. cruzi gp83 antigen is a useful candidate for the development of a vaccine against Chagas disease.

Project description:BackgroundGene-based therapy is a new paradigm for the treatment of Parkinson disease (PD) and offers considerable promise for precise targeting and flexibility to impact multiple pathobiological processes for which small molecule agents are not available. Some success has been achieved utilizing adeno-associated virus for this approach, but it is likely that the characteristics of this vector system will ultimately create barriers to progress in clinical therapy. Adenovirus (Ad) vector overcomes limitations in payload size and targeting. The cellular tropism of Ad serotype 5 (Ad5)-based vectors is regulated by the Ad attachment protein binding to its primary cellular receptor, the coxsackie and adenovirus receptor (CAR). Many clinically relevant tissues are refractory to Ad5 infection due to negligible CAR levels but can be targeted by tropism-modified, CAR-independent forms of Ad. Our objective was to evaluate the role of CAR protein in transduction of dopamine (DA) neurons in vivo.Methodology/principal findingsAd5 was delivered to the substantia nigra (SN) in wild type (wt) and CAR transgenic animals. Cellular tropism was assessed by immunohistochemistry (IHC) in the SN and striatal terminals. CAR expression was assessed by western blot and IHC. We found in wt animals, Ad5 results in robust transgene expression in astrocytes and other non-neuronal cells but poor infection of DA neurons. In contrast, in transgenic animals, Ad5 infects SNc neurons resulting in expression of transduced protein in their striatal terminals. Western blot showed low CAR expression in the ventral midbrain of wt animals compared to transgenic animals. Interestingly, hCAR protein localizes with markers of post-synaptic structures, suggesting synapses are the point of entry into dopaminergic neurons in transgenic animals.Conclusions/significanceThese findings demonstrate that CAR deficiency limits infection of wild type DA neurons by Ad5 and provide a rationale for the development of tropism-modified, CAR-independent Ad-vectors for use in gene therapy of human PD.

Dataset Information

Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

Publications

Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

Similar Datasets

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