Engineered nanoparticles enable deep proteomics studies at scale by leveraging tunable nano-bio interactions.

SignificanceDeep profiling of the plasma proteome at scale has been a challenge for traditional approaches. We achieve superior performance across the dimensions of precision, depth, and throughput using a panel of surface-functionalized superparamagnetic nanoparticles in comparison to conventional workflows for deep proteomics interrogation. Our automated workflow leverages competitive nanoparticle-protein binding equilibria that quantitatively compress the large dynamic range of proteomes to an accessible scale. Using machine learning, we dissect the contribution of individual physicochemical properties of nanoparticles to the composition of protein coronas. Our results suggest that nanoparticle functionalization can be tailored to protein sets. This work demonstrates the feasibility of deep, precise, unbiased plasma proteomics at a scale compatible with large-scale genomics enabling multiomic studies.

Rapid, deep and precise profiling of the plasma proteome with multi-nanoparticle protein corona.

Large-scale, unbiased proteomics studies are constrained by the complexity of the plasma proteome. Here we report a highly parallel protein quantitation platform integrating nanoparticle (NP) protein coronas with liquid chromatography-mass spectrometry for efficient proteomic profiling. A protein corona is a protein layer adsorbed onto NPs upon contact with biofluids. Varying the physicochemical properties of engineered NPs translates to distinct protein corona patterns enabling differential and reproducible interrogation of biological samples, including deep sampling of the plasma proteome. Spike experiments confirm a linear signal response. The median coefficient of variation was 22%. We screened 43 NPs and selected a panel of 5, which detect more than 2,000 proteins from 141 plasma samples using a 96-well automated workflow in a pilot non-small cell lung cancer classification study. Our streamlined workflow combines depth of coverage and throughput with precise quantification based on unique interactions between proteins and NPs engineered for deep and scalable quantitative proteomic studies.

Targeted binding of PLA microparticles with lipid-PEG-tethered ligands.

Solid core polymeric particles are an attractive delivery vehicle as they can efficiently encapsulate drugs of different physical and chemical characteristics. However, the effective targeting of such particles for therapeutic purposes has been somewhat elusive. Here, we report novel polymeric particles comprised of poly(lactic acid) (PLA) with incorporated poly(ethylene glycol)-lipids (PEG-lipids). Particles are characterized for morphology, surface charge, and composition with field-emission scanning electron microscopy (FESEM), zeta potential measurements, and proton nuclear magnetic resonance ((1)H NMR) spectroscopy, respectively. The surface densities of PEG lipids determined by (1)H NMR and particle size distributions are consistent with scaling theory for adsorption of chains onto a surface. We observe significant binding of liganded PEG-lipid tethers when the molecular weight is greater than the unliganded PEG-lipids for significant binding events. Importantly, the binding is not completely lost when the unliganded PEG molecular weight is greater than the liganded PEG-lipid tether. We observe a similar trend for the lower affinity ligand (thioctic acid), but the degree of binding is significantly lower than the high affinity ligand (biotin). This novel technique used to fabricate these liganded particles combined with the lipid bilayer binding studies provides a platform for systematic optimization of particle binding.

Preclinical development and clinical translation of a PSMA-targeted docetaxel nanoparticle with a differentiated pharmacological profile.

We describe the development and clinical translation of a targeted polymeric nanoparticle (TNP) containing the chemotherapeutic docetaxel (DTXL) for the treatment of patients with solid tumors. DTXL-TNP is targeted to prostate-specific membrane antigen, a clinically validated tumor antigen expressed on prostate cancer cells and on the neovasculature of most nonprostate solid tumors. DTXL-TNP was developed from a combinatorial library of more than 100 TNP formulations varying with respect to particle size, targeting ligand density, surface hydrophilicity, drug loading, and drug release properties. Pharmacokinetic and tissue distribution studies in rats showed that the NPs had a blood circulation half-life of about 20 hours and minimal liver accumulation. In tumor-bearing mice, DTXL-TNP exhibited markedly enhanced tumor accumulation at 12 hours and prolonged tumor growth suppression compared to a solvent-based DTXL formulation (sb-DTXL). In tumor-bearing mice, rats, and nonhuman primates, DTXL-TNP displayed pharmacokinetic characteristics consistent with prolonged circulation of NPs in the vascular compartment and controlled release of DTXL, with total DTXL plasma concentrations remaining at least 100-fold higher than sb-DTXL for more than 24 hours. Finally, initial clinical data in patients with advanced solid tumors indicated that DTXL-TNP displays a pharmacological profile differentiated from sb-DTXL, including pharmacokinetics characteristics consistent with preclinical data and cases of tumor shrinkage at doses below the sb-DTXL dose typically used in the clinic.

Patterning adjacent supported lipid bilayers of desired composition to investigate receptor-ligand binding under shear flow.

To achieve efficient targeting, carriers containing either drugs or imaging agents must have surface properties that promote binding to targets yet at the same time block rapid immune system clearance. Here we describe a versatile technique that allows simultaneous comparison of the effects of carrier surface composition on binding properties under identical flow conditions. Parallel lanes of supported lipid bilayers that mimic the surface of liposomal delivery vehicles are formed using the vesicle fusion method in microfluidic channels created via standard soft lithography techniques. Vesicle stock solutions are premixed and injected into lanes formed by a poly(dimethylsiloxane) (PDMS) stamp reversibly sealed to a glass slide to create adjacent lanes of distinct composition. After removing the stamp, an adsorbed layer of bovine serum albumin (BSA) is used to prevent bilayer spreading before assembling the patterned substrate into a flow chamber for binding studies. Advantages of this method include easy and rapid preparation of bilayers with desired compositions from an unlimited number of lipid types, choice of feature size, time-stable features, and low nonspecific binding. Feature sizes on the order of tens of microns allow multiple compositions to be analyzed in one field of view, thereby reducing the number of experiments, ensuring identical flow conditions, and enabling simultaneous incorporation of controls. We show that the presence of a long poly(ethylene glycol) (PEG) tether (MW 2000) between the lipid and ligand results in higher detachment resistances as compared to a short six-carbon spacer.