Microfluidics-assisted Aprotinin-based nano-carrier for Valproic acid delivery

Nanomedicine strategies leveraging bio-derived nano-carriers offer promising avenues for precision chemotherapy. Here, we introduce a microfluidics-assisted nanoconjugate platform that repurposes Aprotinin (Apr); a naturally occurring protein, as an enhanced anti-cancer nano-carrier for Valproic Acid (VPA), a histone deacetylase inhibitor with anticancer properties. The resulting Aprotinin-Valproic Acid (Apr-VPA) nanoconjugates synthesized via a flow-controlled microfluidic process were characterized by various physioco-chemical methods confirming the stable binding. When tested on in vitro studies in 2D cell culture followed by custom designed 3D tumor spheroid models, the Apr-VPA nanoconjugates demonstrated dose-dependent inhibition of spheroid growth and viability, supporting the nanoconjugate's therapeutic potential. Preliminary in vitro comparisons with healthy endothelial cells suggest selective cytotoxicity, warranting further investigation. Interestingly, the designed nanoconjugates also exhibited pH-responsive release behavior under acidic conditions, indicating potential for envirnment-triggered drug release. Thus, it was expected that a novel high technology-controlled synthesis and stimuli sensitive release system, increasing the bioavailability of drug when conjugated (Apr-VPA) as compared to free VPA may perhaps be tranlated to the clinics when validated in vivo. Therefore, plasma profile of VPA and Apr-VPA nanoconjugates by LC-MS/MS and serum enzymatic markers were assessed in wild-type balb/c mice, depicting significantly enhanced availability of drug when conjugated with a carrier. These findings highlight the feasibility of using aprotinin as a scalable protein-based nanocarrier for tumor-preferential therapeutic response in tested cancer models and support its future development in advanced nanomedicine applications.