Biology subjects

Cheema, Y.

Publications and source records attributed to Cheema, Y..

4 recordsLinked to original sources

bioRxiv2025

Barrier Function of the Extracellular Matrix in AAV Gene Therapy

PurposeThe extracellular matrix (ECM) is a major component of the tissue microenvironment which may pose a barrier to the distribution of AAV in target organs, preventing delivery of therapeutic cargo. We sought to address this potential barrier to AAV gene therapy by furthering our understanding of AAV-ECM interactions. We hypothesized that both the AAV serotype and ECM composition will impact AAV transport and gene delivery. MethodsAAV2, AAV6, and AAV8 viral vectors were fluorescently labeled to allow for visualization of their diffusion through the ECM. Lung, liver, and small intestinal submucosal dECM hydrogels were formulated as models of the ECM with tissue-specific biomolecular content. We then characterized AAV and nanoparticle diffusion within decellularized ECM using fluorescent video microscopy and multiple particle tracking. Additionally, we evaluated AAV transduction in dECM-incorporated 2D and 3D spheroid tissue culture models. ResultsAll AAV displayed reduced diffusivity through ECM as compared to similarly sized nanoparticles. AAV2 diffusion was least affected by the presence of ECM across tissue types as compared to AAV6 and AAV8. AAV transduction in dECM incorporated in vitro models was significantly reduced in both a 2D and 3D setting. ConclusionsThese results suggest binding of AAV to the ECM may decrease their therapeutic effect in target tissues throughout the body. The barrier function of the ECM should be considered in development of AAV for gene therapy applications.

bioengineering↗

bioRxiv2025

AAV-mediated MUC5AC siRNA delivery to prevent mucociliary dysfunction in asthma

The main structural components of mucus produced in the lung are mucin 5B (MUC5B) and mucin 5AC (MUC5AC) where a relatively higher expression of MUC5B is typical in health. In the lungs of individuals with asthma, there is a shift from MUC5B to MUC5AC as the predominantly secreted mucin which has been shown to impair mucociliary clearance (MCC) and increase mucus plug formation in the airways. Given its role in asthmatic lung disease, MUC5AC represents a potential therapeutic target where a gene delivery approach could be leveraged to modulate its expression. For these purposes, we explored adeno-associated virus serotype 6 (AAV6), as a lung-tropic viral gene vector to target airway epithelial cells and reduce MUC5AC expression via siRNA delivery. We confirmed that AAV6 was able to transduce epithelial cells in the airways of healthy mice with high transgene expression in mucus-secreting goblet cells. Using multiple particle tracking analysis, we observed that AAV6 was capable of penetrating both normal and MUC5AC-enriched mucus barriers. Successful transduction with AAV6 was also achieved in IL-13 stimulated human airway epithelial (HAE) cells differentiated at air-liquid interface (ALI). AAV6 expressing MUC5AC-targeting siRNA was evaluated as a prophylactic treatment in HAE cell cultures before IL-13 challenge. IL-13 stimulated HAE cultures treated with AAV6-MUC5AC siRNA had significantly reduced MUC5AC mRNA and protein expression compared to untreated controls. Mucociliary transport in IL-13 stimulated HAE cultures was also maintained and comparable to healthy controls following AAV6-MUC5AC siRNA treatment. Together, these findings support that AAV6 may be used as an inhaled gene therapy to suppress MUC5AC overexpression and restore normal airway clearance function in asthma.

cell biology↗

bioRxiv2023

Synthetic mucus barrier arrays as a nanoparticle formulation screening platform

A mucus gel layer lines the luminal surface of tissues throughout the body to protect them from infectious agents and particulates. As a result, nanoparticle drug delivery systems delivered to these sites may become trapped in mucus and subsequently cleared before they can reach target cells. As such, optimizing the properties of nanoparticle delivery vehicles, such as their surface chemistry and size, is essential to improving their penetration through the mucus barrier. In previous work, we developed a mucin-based hydrogel that has viscoelastic properties like that of native mucus which can be further tailored to mimic specific mucosal tissues and disease states. Using this biomimetic hydrogel system, a 3D-printed array containing synthetic mucus barriers was created that is compatible with a 96-well plate enabling its use as a high-throughput screening platform for nanoparticle drug delivery applications. To validate this system, we evaluated several established design parameters to determine their impact on nanoparticle penetration through synthetic mucus barriers. Consistent with the literature, we found nanoparticles of smaller size and coated with a protective PEG layer more efficiently penetrated through synthetic mucus barriers. In addition, we evaluated a mucolytic (tris (2-carboxyethyl) phosphine, TCEP) for use as a permeation enhancer for mucosal drug delivery. In comparison to N-acetyl cysteine (NAC), we found TCEP significantly improved nanoparticle penetration through a disease-like synthetic mucus barrier. Overall, our results establish a new high-throughput screening approach using synthetic mucus barrier arrays to identify promising nanoparticle formulation strategies for drug delivery to mucosal tissues.

bioengineering↗

bioRxiv2021

Shaping nanoparticle diffusion through biological barriers to drug delivery

Nanoparticle drug delivery systems encounter many biological barriers, such as the extracellular matrix and mucus gels, that they must bypass to gain access to target cells. A design parameter that has recently gained attention is nanoparticle shape, as it has been shown elongated rod-shaped nanoparticles achieve higher diffusion rates through biological gels. However, the optimal dimensions of rod-shaped nanoparticles to enhance this effect has yet to be established. To systematically approach this, rod-shaped nanoparticles were synthesized by mechanically stretching 100 nm, 200 nm, and 500 nm spherical nanoparticles. Transmission electron microscopy confirmed this procedure yields a significant fraction of elongated rods and remaining spheres could be removed by centrifugation. Fluorescent microscopy and multiple particle tracking analysis was then used to characterize rod-shaped and spherical nanoparticle diffusion in MaxGel(R), a model extracellular matrix hydrogel. When dispersed in MaxGel, we found rod-shaped nanoparticles exhibited the greatest enhancement in diffusion rate when their length far exceeds the average hydrogel network size. These results further establish the importance of shape as a design criterion to improve nanoparticle-based drug delivery systems.

bioengineering↗