Biology subjects

Bazaz, S.

Publications and source records attributed to Bazaz, S..

3 recordsLinked to original sources

bioRxiv2026

Evaluation of Tetraspanins in Extracellular Vesicle Bioengineering

Extracellular vesicles (EVs) are nano-scale structures produced by cells that transport biological substances for intercellular communication. The tetraspanins CD9, CD81, and CD63 are crucial to EV biogenesis and function. This study uses CRISPR-Cas9 system to knock out (KO) CD9, CD63, and CD81 in HEK293T cells. The goal is to investigate the role of these tetraspanins in EV bioengineering with the hypothesis that repressing endogenous production may increase the availability of exogenously introduced tetraspanin-fusion constructs and increase engineered EV production. Firstly, it is observed that individually knocking out a tetraspanin does not significantly affect EV formation. However, when all three tetraspanins are simultaneously knocked out, there is a marked decrease in EV production, as measured by nanoparticle tracking analysis (NTA). Secondly, upon reintroduction of the corresponding tetraspanins fused to firefly ThermoLuc (Tluc) or neon green (mNG) into the PanKO-, CD9KO, CD63KO-, and CD81KO-cells, the engineered EVs display a significant increase in production by 50% to 70% compared to transduction of wild-type (WT) cells, as measured by luminometer and imaging flow cytometry. These findings emphasize the potential of tetraspanin KO in the bioengineering of EVs, paving the way for new therapeutic applications by enhancing production and potentially modifying their cargo.

bioengineering↗

bioRxiv2025

Peptide nanoparticles for systemic mRNA delivery in rodents and non-human primates

The therapeutic potential of mRNA is vast, and yet translating this potential into effective treatments requires overcoming significant challenges of achieving safe and efficient delivery. This process is hampered by biological barriers that limit cellular uptake, degrade exposed mRNA, and thus necessitate effective endosomal escape to reach the cytoplasm. To address these challenges, we developed a hPep peptide-based nanoparticle (PNP) system that encapsulates mRNA, forming stable and biocompatible particles, which are rapidly taken up by the cells and enable efficient mRNA delivery across various cell culture models. Following systemic administration in mice, lead hPep3/mRNA PNPs achieve broad mRNA expression across multiple tissues, including the lungs, liver, and spleen, and enable effective mRNA delivery to the central nervous system upon local administration. Furthermore, we established a high-yield ([≥]70%) microfluidics-based protocol to scale up the production of well-defined, sterile hPep3/mRNA PNP formulations (approximately 70 nm, PDI around 0.170). Most importantly, in a proof-of-concept study in nonhuman primates (NHPs), we demonstrate that hPep PNPs loaded with human erythropoietin (hEPO) mRNA induce dose-dependent expression of hEPO protein in monkey serum, reaching up to 10 ng/ml at 1.0 mg/kg dose, following both single and repeated administration, while remaining systemically well-tolerated. These findings underscore the potential of hPep peptide-based nanoparticles as a versatile platform for mRNA delivery across multiple tissues, highlighting their promise in advancing the development of mRNA therapeutics.

pharmacology and toxicology↗

bioRxiv2022

SARS-CoV-2 and HSV-1 Induce Amyloid Aggregation in Human CSF

The corona virus (SARS-CoV-2) pandemic and the resulting long-term neurological complications in patients, known as long COVID, have renewed the interest in the correlation between viral infections and neurodegenerative brain disorders. While many viruses can reach the central nervous system (CNS) causing acute or chronic infections (such as herpes simplex virus 1, HSV-1), the lack of a clear mechanistic link between viruses and protein aggregation into amyloids, a characteristic of several neurodegenerative diseases, has rendered such a connection elusive. Recently, we showed that viruses can induce aggregation of purified amyloidogenic proteins via the direct physicochemical mechanism of heterogenous nucleation (HEN). In the current study, we show that the incubation of HSV-1 and SARS-CoV-2 with human cerebrospinal fluid (CSF) leads to the amyloid aggregation of several proteins known to be involved in neurodegenerative diseases, such as: APLP1 (amyloid beta precursor like protein 1), ApoE, clusterin, 2-macroglobulin, PGK-1 (phosphoglycerate kinase 1), ceruloplasmin, nucleolin, 14-3-3, transthyretin and vitronectin. Importantly, UV-inactivation of SARS-CoV-2 does not affect its ability to induce amyloid aggregation, as amyloid formation is dependent on viral surface catalysis via HEN and not its ability to replicate. Additionally, viral amyloid induction led to a dramatic drop in the soluble protein concentration in the CSF. Our results show that viruses can physically induce amyloid aggregation of proteins in human CSF and result in soluble protein depletion, and thus providing a potential mechanism that may account for the association between persistent and latent/reactivating brain infections and neurodegenerative diseases. Significance StatementViruses have generally been excluded from the etiology of amyloid pathologies based on the assumption that amyloid formation requires a proteinaceous conformational template (a prion) to form. Here we show that neuroinvasive viruses induce amyloid aggregation of a plethora of proteins in human CSF even after UV inactivation. Our work illustrates that viruses can induce amyloid aggregation of endogenous human proteins in their native environment by acting as physical catalysts of amyloid nucleation and phase transition. Demonstrating this direct mechanistic link, which is independent of templating, can help better understand the link between viruses and neurodegenerative disorders, especially in the post-COVID-19 era.

microbiology↗