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Despite the widespread use of platinum(Pt)-drugs for cancer-therapy, their development has stalled, as no new Pt-drugs have been approved in over a decade. Packaging small molecule drugs into nanoparticles is a way to enhance their therapeutic efficacy. However, an important consideration in designing Pt-nanoparticles, and the focus of this study, is the rational choice of oxidation-state of Pt. To date, there has been no direct comparison of relative merits of the choice of Pt oxidation-state in the same nanoparticle-system that would allow its optimal design. To address this lacuna, we designed an recombinant asymmetric triblock polypeptide (ATBP) that self-assembles into rod-shaped micelles and chelates Pt(II) or enable covalent conjugation of Pt(IV) with similar morphology and stability. Both ATBP-Pt(II) and ATBP-Pt(IV) nanoparticles enhanced the half-life of Pt by ~45-fold but ATBP-Pt(IV) had superior tumor regression efficacy compared to ATBP-Pt(II) and cisplatin. These results suggest loading Pt(IV) into genetically engineered nanoparticles may yield a new generation of more effective platinum-drug nanoformulations.

Biomolecular condensates composed of proteins and RNA, known as ribonucleoprotein granules, are one approach by which cells regulate post-transcriptional gene expression. The formation of ribonucleoprotein granules typically involves the liquid-liquid phase separation of intrinsically disordered proteins with a target mRNA, sequestering the mRNA into the condensate. This sequestration regulates gene expression by inhibiting translation or facilitating RNA processing, such as splicing. Here, we designed a recombinant fusion of the human Pumilio2 homology domain (Pum2) RNA-binding protein and a synthetic intrinsically disordered protein that exhibits liquid-liquid phase separation to create synthetic ribonucleoprotein granules that extrinsically regulate the expression of a target gene. We show that this fusion protein selectively binds an RNA transcript of interest that contains a Pum2-binding RNA sequence at its 3’-end and sequesters the mRNA within a biomolecular condensate. Sequestration of a target mRNA within the condensate largely reduces the translation of the target RNA in protocells and in E. coli compared to cells that do not sequester the target mRNA in a synthetic condensate. We demonstrate that the viability of E. coli that harbor a cytotoxic protein can be rescued by sequestering the mRNA encoding the cytotoxic protein within a synthetic condensate. Finally, we use RNA-seq to determine that the target RNA of interest is preferentially sequestered in synthetic ribonucleoprotein granules. This approach enables modulation of cell function via the formation of a synthetic biomolecular condensate that spatiotemporally regulates the expression of a target protein.

Combining surface-initiated, TdT (terminal deoxynucleotidyl transferase) catalyzed enzymatic polymerization (SI-TcEP) with precisely engineered DNA origami nanostructures (DONs) presents an innovative pathway for the generation of stable, polynucleotide brush-functionalized origami nanostructures. We demonstrate that SI-TcEP can site-specifically pattern DONs with brushes containing both natural and non-natural nucleotides. The brush functionalization can be precisely controlled in terms of the location of initiation sites on the origami core and the brush height and composition. Coarse-grained simulations predict the conformation of the brush-functionalized DONs that agree well with the experimentally observed morphologies. We find that polynucleotide brush-functionalization increases the nuclease resistance of DONs significantly, and that this stability can be spatially programmed through the site-specific growth of polynucleotide brushes. The ability to site-specifically decorate DONs with brushes of natural and non-natural nucleotides provides access to a large range of functionalized DON architectures that would allow for further supramolecular assembly, and for potential applications in smart nanoscale delivery systems.

Interaction of nanoparticles with biological systems is a key factor influencing their efficacy as a drug delivery vehicle. The inconsistency in defining the optimal design parameters across different nanoparticle types suggests that information gained from one model system need not apply to other systems. Therefore, selection of a versatile model system is critical for such studies. Cubosomes are one of the potential drug delivery vehicles due to their biocompatibility, stability, ability to carry hydrophobic, hydrophilic, and amphiphilic drugs, and ease of surface modification. Here we report the importance of surface architecture of cubosomes by comparing their cellular uptake mechanism with poly-ε-lysine (PεL)-coated cubosomes. Uncoated cubosomes entered cells by an energy-independent, cholesterol-dependent mechanism, whereas PεL-coated cubosomes relied on energy-dependent mechanisms to enter the endosomes. As endosomal entrapment was evaded by uncoated cubosomes, they can be preferably used for cytosolic delivery of therapeutic agents.

The recognition that nucleic acids can be used as polymeric materials led to the blossoming of the field of DNA nanotechnology, with a broad range of applications in biotechnology, biosensors, diagnostics, and drug delivery. These applications require efficient methods to synthesize and chemically modify high molecular weight DNA. Here, we discuss terminal deoxynucleotidyl transferase (TdT)-catalyzed enzymatic polymerization (TcEP) as an alternative to conventional enzymatic and solid-phase DNA synthesis. We describe biochemical requirements for TcEP and provide step-by-step protocols to carry out TcEP in solution and from surfaces.

RNAi is emerging as a promising technology for treatment of various diseases due to its ability to silence specific target genes. To date, a number of nanoparticle based formulations have been reported for the delivery of small interfering RNA (siRNA), with continuous modifications in the nanoparticle design for enhancing their efficiency. While majority of the design aspects are focused on avoiding or overcoming endosomal entrapment, limited studies are available that address the role of interaction of nanoparticles with the RNA induced silencing complex (RISC) machinery, which is a crucial aspect deciding the outcome. Here, we systematically probed the effect of steric hindrance of nanoparticles on RISC interaction, by modulating two parameters, nanoparticle size and hardness. An assay was developed for quantifying the extent of RISC interaction of different nanoparticles in vitro, which was then correlated with their gene knockdown efficiency. The results suggest that the soft and small nanoparticles were most efficacious in knocking down polo-like-kinase 1 (PLK1) siRNA, a gene overexpressed in a variety of cancer types.

Despite the promising photophysical properties of fluorescent graphene quantum dots (GQDs), their cellular toxicity needs to be addressed before their full potential could be completely realized in biomedicine. A simple method for mitigating the toxicity of GQDs by embedding them in PEG matrix is reported here. The enhanced biocompatibility of polymer modified, P-GQDs, is attributed to reduced reactive oxygen species generation, as measured by an intracellular ROS assay. We also demonstrate the enhanced loading and efficient intracellular delivery of therapeutics by P-GQDs.

We report the preparation and characterization of monoolein cubosomes that can be easily surface modified through adsorption of a single layer of cationic poly-epsilon-lysine. Poly-epsilon-lysine coated cubosomes show remarkable stability in serum solution, are nontoxic and, are readily internalized by HeLa cells. The poly-epsilon-lysine coating provides chemical handles for further bioconjugation of the cubosome surface. We also demonstrate that the initial release rate of a hydrophilic drug, Naproxen sodium, from the cubosomes is retarded with just a single layer of polymer. Interestingly, cubosomes loaded with Naproxen sodium, recently shown to have anticancer properties, cause more apoptosis in HeLa cells when compared to free unencapsulated drug.

This article describes a new and unified approach to computerized hand diagnosis and rehabilitation. The system uses a workstation to automate diagnosis data collection and analysis and to assess the rehabilitation progress. A new diagnosis glove was developed and tested. This device measures grasping forces applied to 16 regions of the hand. A physician using this system can also utilize modern diagnosis devices such as electronic dynamometer, pinchmeter, and goniometer. Additionally, three VR rehabilitation exercises were created using WorldToolKit graphics library and run on the same workstation. These exercises were modeled after standard hand rehabilitation procedures and involve manipulation of virtual objects and transparent real-time data gathering. Grasping forces were modeled and fed back using the Rutgers Master worn on the patient's hand. An Oracle database was used to store, analyze, and integrate the patient's diagnosis and rehabilitation data. The system is presently undergoing clinical trials.

A virtual model of a knee joint with muscles, ligaments and bones has been developed. This model includes realistic 3-D surface deformation and tissue stiffnesses. Tissue and bone deformation (palpation) produces real time force feedback to the user hand wearing a DataGlove and Rutgers Master. Collision detection algorithms determine when and where the virtual hand palpates the surface model. This user interaction with the muscles and bones of a human knee model may be used as a training and planning tool for knee surgery.