(480a) Merging ‘Micro' with ‘Nano': On-Chip High-Throughput Synthesis of Polymeric Nanoparticles for Cancer Therapy

Polymeric nanoparticles (NPs) comprised of poly(lactide-co-glycolide)-b-polyethyleneglycol (PLGA-PEG) block copolymers are promising drug delivery vehicles with the advantages of controlled drug release, enhanced stability, and ability to carry thousands of drug molecules per NP [1]. Such NPs can be targeted to differentially deliver drugs to the site of interest in the body, translating to improvement in the therapeutic index of drugs [2]. However, physicochemical properties of NPs such as size, charge, surface chemistry, payload, and targeting ligand density significantly affect the biodistribution of the drug, and the delicate balance between each these properties needs to be experimentally determined and precisely engineered for ultimate in vivo success [3]. Here we report the development of a microfluidic system that enables reproducible, high-throughput preparation of a library of distinct homogeneous, PLGA-PEG NPs. We demonstrate the ability to synthesize and in vitro screen tens of different NP formulations per day by integrating our microfluidic technology with high-throughput flow cytometry (FACS). This makes an optimal system for the translation of nanovehicles to in vivo applications. To the best of our knowledge, this is the first time a microfluidic system is developed for the synthesis and screening of polymeric nanovehicles for drug development applications.

Our microfluidic system, made of PDMS, is composed of a mixing unit and a NP assembly unit. The mixing unit consists on a multi-inlet 2-layer mixer where different precursors such as PLGA-PEG of different MW and charge, PLGA-PEG-folate (targeting agent), drugs, and solvents are mixed at different ratios into a homogenous solution. In the ?assembly' unit, the precursor solution is rapidly mixed with an anti-solvent (i.e. water) using 3D hydrodynamic flow focusing where NPs self-assemble after complete mixing [4]. With this system we can make, in a single step, NPs of different size, charge, surface chemistry, targeting agent density, and drug loading, by simply varying the flow ratios of the streams with different precursors entering the mixing unit.

For the NPs to be effective, it is essential for them to specifically bind to cancerous cells, undergo endocytosis, and release the drug inside the cell [5]. In vitro screening of NPs with varying properties including surface charge, size, targeting ligand density, PEG densities, and chemical composition has the potential to yield valuable insights into the interactions of the NPs at the cellular level. With this aim, we used high-throughput FACS to screen for the uptake of different NP formulations by macrophage cell-lines and cancerous cell-lines as a proof-of-concept. Preliminary data portrays a delicate balance between PEG and folate surface densities for NPs to successfully enter cancer cells while avoiding macrophages. In addition, we demonstrate the capability of our system to combinatorially synthesize and screen NPs against different cell lines.

Our experiments demonstrate a simple yet powerful technology to optimize polymeric nanoparticles in a high-throughput manner by integrating a microfluidic system with high-throughput flow cytometry. This technology forms a robust platform for investigating the effects of different physicochemical properties of NPs on cellular level interactions. This is a clear example where the advantages of microfluidics and nanotechnology are harnessed in a single system.

References 1.?Formulation of functionalized PLGA-PEG nanoparticles for in vivo targeted drug delivery,? J. Cheng et al, Biomaterials, 28(5), 869 (2007) 2.?Targeted nanoparticles for cancer therapy,? F. Gu et al, Nano Today, 2(3), 14 (2007) 3.?Precise engineering of targeted nanoparticles by using self-assembled biointegrated block copolymers,? F. Gu et al, PNAS, 105 (7), 2586 (2008) 4.?Microfluidic platform for controlled synthesis of polymeric nanoparticles,? R. Karnik et al, Nano Letters, 8 (9), 2906 (2008). 5.?Opsonization, biodistribution, and pharmacokinetics of polymeric nanoparticles,? D. E. Owens et al, International Journal of Pharmaceutics, 307 (1), 93 (2006)