Polymer nanoparticles (PNPs) that exhibit selective stimuli-responsive degradation and drug release at tumor sites are promising candidates in the development of smart nanomedicines. In this thesis, we demonstrate a microfluidic approach to manufacturing biological stimuli-responsive PNPs with flow-tunable physicochemical and pharmacological properties. The investigated PNPs contain cleavable disulfide linkages in two different locations (core and interface, DualM PNPs) exhibiting responsivity to elevated levels of glutathione (GSH), such as those found within cancerous cells.First, we conduct a mechanistic study on the microfluidic formation of DualM PNPs without encapsulated drug. We show that physicochemical properties, including size, morphology, and internal structure, of DualM PNPs are tunable with manufacturing flow rate. Microfluidic formation of DualM PNPs is explained by the interplay of shear-induced coalescence, shear-induced breakup, and intraparticle chain rearrangements. In addition, we demonstrate that rates of GSH-triggered changes in size and internal structure are linearly correlated with initial PNP sizes and internal structures, respectively.Next, we expand our study to focus on microfluidic control of pharmacological properties of DualM PNPs containing either an anticancer drug (paclitaxel, PAX-PNPs) or a fluorescent drug surrogate (DiI-PNPs). Microfluidic PAX-PNPs and DiI-PNPs show similar sizes and morphologies with their non-drug-loaded counterparts under the same flow conditions. We then show that pharmacological properties of DualM PNPs, including encapsulation efficiency, GSH-triggered release rate, cell uptake, cytotoxicity against MCF-7 (cancerous) and HaCaT (healthy), and relative difference in MCF-7 and HaCaT cytotoxicity, all increase linearly as flow-directed PNP size decreases, providing remarkably simple process-structure-property relationships. In addition, we show that microfluidic manufacturing improves encapsulation homogeneities within PNPs relative to bulk nanoprecipitation. These results highlight the potential of flow-directed shear iv processing in microfluidics for providing controlled manufacturing routes to biological stimuli-responsive nanomedicines optimized for specific therapeutic applications.Finally, we summarize various design strategies of biological stimuli-responsive PNPs. We show that the location and density of disulfide linkages within PNPs determines stimulus-triggered degradation mechanism and kinetics. In addition, we show various bottom-up approaches to tune PNP responsivities that involves chemical processing, including formulation chemistry and intramolecular forces. Along with the top-down microfluidic approach that we demonstrate experimentally, this chapter provides a more comprehensive understanding of process-structure-property relations opening up vast possibilities for manufacturing smarter nanomedicines.

We demonstrate microfluidic manufacturing of glutathione (GSH)-responsive polymer nanoparticles (PNPs) with controlled in vitro pharmacological properties for selective drug delivery. This work leverages previous fundamental work on microfluidic control of the physicochemical properties of GSH-responsive PNPs containing cleavable disulfide groups in two different locations (core and interface, DualM PNPs). In this paper, we employ a two-phase gas–liquid microfluidic reactor for the flow-directed manufacturing of paclitaxel-loaded or DiI-loaded DualM PNPs (PAX-PNPs or DiI-PNPs, where DiI is a fluorescent drug surrogate dye). We find that both PAX-PNPs and DiI-PNPs exhibit similar flow-tunable sizes, morphologies, and internal structures to those previously described for empty DualM PNPs. Fluorescent imaging of DiI-PNP formulations shows that microfluidic manufacturing greatly improves the homogeneity of drug dispersion within the PNP population compared to standard bulk microprecipitation. Encapsulation of PAX in DualM PNPs significantly increases its selectivity to cancerous cells, with various PAX-PNP formulations showing higher cytotoxicity against cancerous MCF-7 cells than against non-cancerous HaCaT cells, in contrast to free PAX, which showed similar cytotoxicity in the two cell lines. In addition, the characterization of DualM PNP formulations formed at various microfluidic flow rates reveals that critical figures of merit for drug delivery functionincluding encapsulation efficiencies, GSH-triggered release rates, rates of cell uptake, cytotoxicities, and selectivity to cancerous cellsexhibit microfluidic flow tunability that mirrors trends in PNP size. These results highlight the potential of two-phase microfluidic manufacturing for controlling both structure and drug delivery function of biological stimuli-responsive nanomedicines toward improved therapeutic outcomes.