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Defective genes account for ∼80% of the total of more than 7,000 diseases known to date. Gene therapy brings the promise of a one-time treatment option that will fix the errors in patient genetic coding. Recombinant viruses are highly efficient vehicles for in vivo gene delivery. Adeno-associated virus (AAV) vectors offer unique advantages, such as tissue tropism, specificity in transduction, eliciting of a relatively low immune responses, no incorporation into the host chromosome, and long-lasting delivered gene expression, making them the most popular viral gene delivery system in clinical trials, with three AAV-based gene therapy drugs already approved by the US Food and Drug Administration (FDA) or European Medicines Agency (EMA). Despite the success of AAV vectors, their usage in particular scenarios is still limited due to remaining challenges, such as poor transduction efficiency in certain tissues, low organ specificity, pre-existing humoral immunity to AAV capsids, and vector dose-dependent toxicity in patients. In the present review, we address the different approaches to improve AAV vectors for gene therapy with a focus on AAV capsid selection and engineering, strategies to overcome anti-AAV immune response, and vector genome design, ending with a glimpse at vector production methods and the current state of recombinant AAV (rAAV) at the clinical level.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous cell population with high immunosuppressive activity that proliferates in infections, inflammation, and tumor microenvironments. In tumors, MDSC exert immunosuppression mainly by producing reactive oxygen species (ROS), a process triggered by the NADPH oxidase 2 (NOX2) activity. NOX2 is functionally coupled with the Hv1 proton channel in certain immune cells to support sustained free-radical production. However, a functional expression of the Hv1 channel in MDSC has not yet been reported. Here, we demonstrate that mouse MDSC express functional Hv1 proton channel by immunofluorescence microscopy, flow cytometry, and Western blot, besides performing a biophysical characterization of its macroscopic currents via patch-clamp technique. Our results show that the immunosuppression by MDSC is conditional to their ability to decrease the proton concentration elevated by the NOX2 activity, rendering Hv1 a potential drug target for cancer treatment.

SignificanceImmunosuppression by myeloid-derived suppressor cells (MDSC), especially near tumor surfaces, involves the extracellular production of reactive oxygen species (ROS). ROS generation in MDSC occurs during the oxidation of NADPH to NADP+, which NOX2 catalyzes. ROS react with the T cell receptor complex, abolishing the antigen presentation, which blocks the immune system elimination of the tumor cells. Extrusion of protons from MDSC by voltage-gated proton channel (Hv1) sustains ROS production. Here, we demonstrate the expression of Hv1 in mouse MDSC. In this way, Hv1 present in MDSC becomes a potential cancer therapeutic target since its inhibition seems to diminish immunosuppression activity in the tumoral microenvironment, allowing cancer cells to be attacked by the immune system.