We have continued to develop AAV vectors for gene transfer and we are actively engaged in evaluating their use in several gene therapy applications including gene transfer to the lung, CNS, eye, and salivary gland. In addition to distributing these vectors to labs throughout the world, we have continued to collaborate with a number of researchers in the field and in several publications describe the use of AAV based vectors to express genes in cancer application (Hida et al. 2010), the inner ear of guinea pigs for the treatment of hereditary deafness and balance disorders (Sheffield et al. 2011), primary salivary epithelial cells to repair damage to the gland (Gao et al. 2011), or produce therapeutic proteins (Nguyen et al 2010). We also are working with both intramural and extramural researches to test the effect of local expression AAV based vectors expressing AQP1 for the treatment of radiation induced xerostomia (Zheng et al 2011). In addition through funding received from the office of the scientific director as part of the salivary gland biology initiative, we are collaborating with researchers in the Laboratory of Cell and Developmental Biology, NIDCR at identifying AAV and adenovirus vectors for gene transfer to the developing salivary gland. Vector discovery and characterization In 2003 we reported the development of a microarray based high throughput screening technique for identifying gene expression patterns that correlated with a specific phenotype (Di Pasquale et al. 2003). This approach was termed comparative gene analysis (CGA). Since our initial publication we have continued to refine the bioinformatics aspect of this approach and recently applied it to several diverse systems. Through funding receive as part of an NIAID biodefense grant we have worked with extramural researchers to identify genes that correlate with Ebola virus entry. Unlike AAV, the entry pathway of enveloped viruses is very complex and may take several routes. We have recently reported a component of Ebola entry involves macropinocytosis and specifically the activation of RhoC and Axl, a largely unstudied regulator of this pathway (Quinn et al. 2009, Brindley et al. 2011). We have continued this project and have recently reported the identification of a new class of receptor molecules for Ebola (TIM1) (Kondratowicz 2011). TIM-1 is also recognized at the receptor for hepatitis A virus and its expression was shown to be critical for Ebola virus entry. TIM-1 was not expressed in all permissive cell lines, suggesting that additional, currently unidentified cellular proteins may also enhance filovirus entry. Sjogrens syndrome Sjogrens syndrome is an autoimmune disease, characterized by lymphoid cell infiltration into the salivary and lacrimal glands, and affects 0.5% of the population in the United States of which 90% are women. The consequence of chronic immune cell activation in these exocrine glands is diminished secretory function, which leads to symptoms of dry mouth and dry eyes. Currently several mouse models of this disease are available and we are reviewing and evaluating them as an alternative to the NOD mouse. We recently reported on the development of a novel mouse model based on immunization with peptides from a common antigen associated with Sjogrens syndrome Ro60, and identified several cytokine changes key to the development of disease (Yin et al. 2011). TH17 cell populations that secrete predominantly interleukin (IL)-17A have been shown to play an important role in an increasing number of autoimmune diseases, including SS. Using gene transfer technology to test the role of IL-17 in mice, C57BL/6J mice expressing IL17 locally in the salivary gland developed a SS-like disease profile, including the appearance of lymphocytic foci, increased cytokine levels, changes in antinuclear antibody profiles, and temporal loss of saliva flow (Nguyen et al. 2010a). In contrast injection of an IL-17 antagonist, IL17R:Fc, exhibited a rapid temporal, yet persistent, decrease in the levels of serum IL17 as well as the overall numbers of CD4+IL17+T cells present in their spleens accompanied by a decrease in lymphocytic infiltrations of their salivary glands, normalization of their antinuclear antibodies repertoire, and increased saliva secretion (Nguyen et al. 2010b). In addition to cytokines, adhesion molecules play a critical role in the initiation of inflammation and inhibitors of this pathway have been developed for the treatment of other autoimmune diseases. In parallel with ongoing studies in the NIDCR Sjogrens syndrome clinic we have investigated the potential of inhibitors of LFA1 activation (soluble ICAM) to block the onset of the Sjogrens syndrome phenotype in NOD mice (Roescher et al 2011). Early treatment with ICAM-1/Fc resulted in decreased average number of inflammatory foci without changes in T and B cell composition. In contrast, late treated mice did not show any change in focus scores, but immunohistochemical staining showed an increase in the overall number of CD4+ and CD8+ T cells. Moreover, early treated mice showed decreased IgM within the SGs, whereas late treated mice had increased IgM levels, and on average higher IgG and IgA. This adverse response to inhibition of LFA activation mirrors those observed in the clinic upon treatment with raptiva, an anti-LFA1 molecule. These findings suggest that expression of LFA1 inhibitors and our previous work on TNFa support the used of current animal models in preclinical studies of Sjogrens syndrome. Furthermore, the results of this study support a continued close working relationship with the Sjgrens clinic to evaluate potential therapies. In summary, the future directions for the AAV Biology Section will be to continue examination and development of gene transfer vectors for use in treating disease as well as refine our tools for studying interactions necessary for cellular transduction.

Project Start
Project End
Budget Start
Budget End
Support Year
12
Fiscal Year
2011
Total Cost
$1,832,712
Indirect Cost
Name
National Institute of Dental & Craniofacial Research
Department
Type
DUNS #
City
State
Country
Zip Code
Corden, A; Handelman, B; Yin, H et al. (2017) Neutralizing antibodies against adeno-associated viruses in Sjögren's patients: implications for gene therapy. Gene Ther 24:241-244
Alevizos, I; Zheng, C; Cotrim, A P et al. (2017) Immune reactivity after adenoviral-mediated aquaporin-1 cDNA transfer to human parotid glands. Oral Dis 23:337-346
Zeng, Mei; Szymczak, Mitchell; Ahuja, Malini et al. (2017) Restoration of CFTR Activity in Ducts Rescues Acinar Cell Function and Reduces Inflammation in Pancreatic and Salivary Glands of Mice. Gastroenterology 153:1148-1159
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Alevizos, I; Zheng, C; Cotrim, A P et al. (2017) Late responses to adenoviral-mediated transfer of the aquaporin-1 gene for radiation-induced salivary hypofunction. Gene Ther 24:176-186
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Weller, Melodie L; Gardener, Matthew R; Bogus, Zoe C et al. (2016) Hepatitis Delta Virus Detected in Salivary Glands of Sjögren's Syndrome Patients and Recapitulates a Sjögren's Syndrome-Like Phenotype in Vivo. Pathog Immun 1:12-40
Chiorini, John A (2016) And one to bind them all. Oral Dis :
Lai, Zhennan; Yin, Hongen; Cabrera-Pérez, Javier et al. (2016) Aquaporin gene therapy corrects Sjögren's syndrome phenotype in mice. Proc Natl Acad Sci U S A 113:5694-9

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