IADR Abstract Archives
Aquaporin-1 Gene Therapy Corrects Sjögren’s Syndrome Fluid Secretion Defects in Human Acinar Cells
Objectives: Sjögren’s syndrome (SS) is a chronic autoimmune disease characterized by salivary and lacrimal dysfunction with main complaints of dry mouth and dry eyes. Reduced salivation reflects underlying fluid secretion defects attributable to reduced membrane permeability of acinar cells. We hypothesize that correcting fluid secretion defects through increased water permeability is a plausible strategy to treat salivary hypofunction in SS. In several animal models, including SS, the correction of membrane permeability using gene transfer of human aquaporin 1 (hAQP1) effectively restored fluid secretion. Moreover, hAQP1 gene transfer improved salivation and dry mouth in patients with radiation-induced xerostomia. To address this unmet clinical need, we aimed to test AAV2-hAQP1 gene transfer to restore fluid secretion in human minor salivary glands (MSG) from SS patients.
Methods: Patients received comprehensive evaluations, including lower lip MSG biopsies in the NIDCR Sjögren’s Syndrome Clinic. Sixteen patients were screened: 9 met American College of Rheumatology 2016 criteria. MSG biopsies were divided into RNAlater stored or immediately micro-dissected into lobule preparations in basal media. Lobule preparations were cultured in 24-well transwell plates at the air-media interface. After 2 hours of equilibration, MSG pieces were transduced with 6.25E+11 particles of AAV2-hAQP1 and 1.35E+11 AAV2-mCherry. After 24 hours of transduction, volume change response to 2uM carbachol was measured. Finally, the pieces were recovered and fixed in 4% paraformaldehyde for immunofluorescence or preserved in RNAlater for RNA sequencing.
Results: We found that in both the SS (p = 0.02) and the non-SS (p <0.01), MSG exhibited correction of their fluid secretion deficits up to 80% of normal. Using immunofluorescence, we confirm AAV2-hAQP1 transduction in the acinar tissue. RNA-sequencing is planned to investigate predictive biomarkers of response to gene transfer.
Conclusions: Our data suggest that, in the context of SS pathophysiology, patient’s salivary hypofunction could be treated using AAV2-hAQP1.
Methods: Patients received comprehensive evaluations, including lower lip MSG biopsies in the NIDCR Sjögren’s Syndrome Clinic. Sixteen patients were screened: 9 met American College of Rheumatology 2016 criteria. MSG biopsies were divided into RNAlater stored or immediately micro-dissected into lobule preparations in basal media. Lobule preparations were cultured in 24-well transwell plates at the air-media interface. After 2 hours of equilibration, MSG pieces were transduced with 6.25E+11 particles of AAV2-hAQP1 and 1.35E+11 AAV2-mCherry. After 24 hours of transduction, volume change response to 2uM carbachol was measured. Finally, the pieces were recovered and fixed in 4% paraformaldehyde for immunofluorescence or preserved in RNAlater for RNA sequencing.
Results: We found that in both the SS (p = 0.02) and the non-SS (p <0.01), MSG exhibited correction of their fluid secretion deficits up to 80% of normal. Using immunofluorescence, we confirm AAV2-hAQP1 transduction in the acinar tissue. RNA-sequencing is planned to investigate predictive biomarkers of response to gene transfer.
Conclusions: Our data suggest that, in the context of SS pathophysiology, patient’s salivary hypofunction could be treated using AAV2-hAQP1.