Browsing by Author "Nilewski, Lizanne G."

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    Antioxidant Carbon Nanoparticles Inhibit Fibroblast-Like Synoviocyte Invasiveness and Reduce Disease Severity in a Rat Model of Rheumatoid Arthritis
    (MDPI, 2020) Tanner, Mark R.; Huq, Redwan; Sikkema, William K.A.; Nilewski, Lizanne G.; Yosef, Nejla; Schmitt, Cody; Flores-Suarez, Carlos P.; Raugh, Arielle; Laragione, Teresina; Gulko, Pércio S.; Tour, James M.; Beeton, Christine; The NanoCarbon Center
    Reactive oxygen species have been involved in the pathogenesis of rheumatoid arthritis (RA). Our goal was to determine the effects of selectively scavenging superoxide (O2•−) and hydroxyl radicals with antioxidant nanoparticles, called poly(ethylene glycol)-functionalized hydrophilic carbon clusters (PEG-HCCs), on the pathogenic functions of fibroblast-like synoviocytes (FLS) from patients with rheumatoid arthritis (RA) and on the progression of an animal model of RA. We used human FLS from patients with RA to determine PEG-HCC internalization and effects on FLS cytotoxicity, invasiveness, proliferation, and production of proteases. We used the pristane-induced arthritis (PIA) rat model of RA to assess the benefits of PEG-HCCs on reducing disease severity. PEG-HCCs were internalized by RA-FLS, reduced their intracellular O2•−, and reduced multiple measures of their pathogenicity in vitro, including proliferation and invasion. In PIA, PEG-HCCs caused a 65% reduction in disease severity, as measured by a standardized scoring system of paw inflammation and caused a significant reduction in bone and tissue damage, and circulating rheumatoid factor. PEG-HCCs did not induce lymphopenia during PIA. Our study demonstrated a role for O2•− and hydroxyl radicals in the pathogenesis of a rat model of RA and showed efficacy of PEG-HCCs in treating a rat model of RA.
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    Carbon nanomaterials and their small molecule analogues for biomedical applications
    (2017-09-26) Nilewski, Lizanne G.; Tour, James M
    Carbon nanomaterials possess unique structural features that give them interesting properties and reactivity for applications in medicine, electronics, materials, catalysis, and more. Herein, the synthesis of PEG-HCCs, PEG-GQDs, and small molecule analogues of these functionalized carbon nanomaterials, as well as their biomedical applications as drug delivery vehicles and antioxidants, are described. PEGylated hydrophilic carbon clusters (PEG-HCCs) are non-toxic water-soluble carbon nanomaterials synthesized by the oxidation of single walled carbon nanotubes (SWCNTs). It is shown that PEG-HCCs can be non-covalently loaded with drugs and covalently modified with various targeting peptides to deliver drugs selectively to glioblastoma cells in vitro and tumors in vivo. It was also found that sensitive imaging of tumors over normal tissue could be achieved by loading the peptide-targeted PEG-HCCs with fluorescent dyes instead of drugs. PEG-HCCs are also powerful antioxidants that are capable of quenching superoxide and hydroxyl radicals. The application of PEG-HCCs as immunomodulators for the treatment of multiple sclerosis (MS) and other T cell-mediated autoimmune disorders like rheumatoid arthritis, was explored. PEG-HCCs were observed to selectively target T cells over other immune cells, and were found to modulate T cell activity by inhibiting proliferation, and they significantly reduced the severity of symptoms in a rat model of MS. In addition to PEG-HCCs, another class of antioxidant carbon nanomaterials, coal-derived graphene quantum dots (GODs) and PEG-GQDs, was developed. These highly oxidized redox active materials were characterized and tested with respect to superoxide and hydroxyl radical activity. PEG-GQDs were also studied in a rat model of traumatic brain injury. It was found that GQDs and PEG-GQDs possess similar characteristics, reactivity, and antioxidant abilities to the HCCs both in cell-free systems and in vivo. Furthermore, perylene diimides (PDIs) and naphthalene diimides (NDIs) were synthesized to mimic the structure and antioxidant activity of the HCCs and GQDs. The PDIs in particular were found to mimic both HCCs and the enzyme SOD by catalyzing the dismutation of superoxide into O2 and H2O2. PDIs were studied with respect to their antioxidant properties, and modified PDIs were also synthesized to modulate their redox properties. Finally, PDIs were studied as antioxidants in in vitro T cell experiments, where it was found they have similar behavior to the PEG-HCCs as potential immunomodulators. Lastly, another class of biologically active targeted molecules was studied for the treatment of cancer. Light-activated nanomachines based on the previous work of the Tour lab were selectively targeted to cancer cells using short peptides, similar to the targeting of PEG-HCCs to tumors. These peptide-targeted nanomachines were shown to associate with their intended target cancer cells over control cells, and once bound to the membranes of target cells, the nanomachines were activated using UV light. This induced MHz rotation of the “motor” moiety and mechanically disrupted the cell membrane leading to cell death. Overall, the PEG-HCCs, PEG-GQDs, PDI/NDI small molecules, and nanomachines were studied in cell-free systems, in vitro, and in vivo with promising results that warrant continued investigation of their mechanism and applications.
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    Efficacy of Novel Carbon Nanoparticle Antioxidant Therapy in a Severe Model of Reversible Middle Cerebral Artery Stroke in Acutely Hyperglycemic Rats
    (Frontiers, 2018) Fabian, Roderic H.; Derry, Paul J.; Rea, Harriett Charmaine; Dalmeida, William V.; Nilewski, Lizanne G.; Sikkema, William K.A.; Mandava, Pitchaiah; Tsai, Ah-Lim; Mendoza, Kimberly; Berka, Vladimir; Tour, James M.; Kent, Thomas A.
    INTRODUCTION: While oxidative stress can be measured during transient cerebral ischemia, antioxidant therapies for ischemic stroke have been clinically unsuccessful. Many antioxidants are limited in their range and/or capacity for quenching radicals and can generate toxic intermediates overwhelming depleted endogenous protection. We developed a new antioxidant class, 40 nm × 2 nm carbon nanoparticles, hydrophilic carbon clusters, conjugated to poly(ethylene glycol) termed PEG-HCCs. These particles are high-capacity superoxide dismutase mimics, are effective against hydroxyl radical, and restore the balance between nitric oxide and superoxide in the vasculature. Here, we report the effects of PEG-HCCs administered during reperfusion after transient middle cerebral artery occlusion (tMCAO) by suture in the rat under hyperglycemic conditions. Hyperglycemia occurs in one-third of stroke patients and worsens clinical outcome. In animal models, this worsening occurs largely by accelerating elaboration of reactive oxygen species (ROS) during reperfusion. METHODS: PEG-HCCs were studied for their protective ability against hydrogen peroxide in b.End3 brain endothelial cell line and E17 primary cortical neuron cultures. In vivo, hyperglycemia was induced by streptozotocin injection 2 days before tMCAO. 58 Male Sprague-Dawley rats were analyzed. They were injected IV with PBS or PEG-HCCs (4 mg/kg 2×) at the time of recanalization after either 90- or 120-min occlusion. Rats were survived for up to 3 days, and infarct volume characteristics and neurological functional outcome (modified Bederson Score) were assessed. RESULTS: PEG-HCCs were protective against hydrogen peroxide in both culture models. In vivo improvement was found after PEG-HCCs with 90-min ischemia with reduction in infarct size (42%), hemisphere swelling (46%), hemorrhage score (53%), and improvement in Bederson score (70%) (p = 0.068-0.001). Early high mortality in the 2-h in the PBS control group precluded detailed analysis, but a trend was found in improvement in all factors, e.g., reduction in infarct volume (48%; p = 0.034) and a 56% improvement in Bederson score (p = 0.055) with PEG-HCCs. CONCLUSION: This nano-antioxidant showed some improvement in several outcome measures in a severe model of tMCAO when administered at a clinically relevant time point. Long-term studies and additional models are required to assess potential for clinical use, especially for patients hyperglycemic at the time of their stroke, as these patients have the worst outcomes.
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    Molecular machines open cell membranes
    (Springer Nature, 2017) García-López, Víctor; Chen, Fang; Nilewski, Lizanne G.; Duret, Guillaume; Aliyan, Amir; Kolomeisky, Anatoly B.; Robinson, Jacob T.; Wang, Gufeng; Pal, Robert; Tour, James M.
    Beyond the more common chemical delivery strategies, several physical techniques are used to open the lipid bilayers of cellular membranes[1]. These include using electric[2] and magnetic[3] fields, temperature[4], ultrasound[5] or light[6] to introduce compounds into cells, to release molecular species from cells or to selectively induce programmed cell death (apoptosis) or uncontrolled cell death (necrosis). More recently, molecular motors and switches that can change their conformation in a controlled manner in response to external stimuli have been used to produce mechanical actions on tissue for biomedical applications[7,8,9]. Here we show that molecular machines can drill through cellular bilayers using their molecular-scale actuation, specifically nanomechanical action. Upon physical adsorption of the molecular motors onto lipid bilayers and subsequent activation of the motors using ultraviolet light, holes are drilled in the cell membranes. We designed molecular motors and complementary experimental protocols that use nanomechanical action to induce the diffusion of chemical species out of synthetic vesicles, to enhance the diffusion of traceable molecular machines into and within live cells, to induce necrosis and to introduce chemical species into live cells. We also show that, by using molecular machines that bear short peptide addends, nanomechanical action can selectively target specific cell-surface recognition sites. Beyond the in vitro applications demonstrated here, we expect that molecular machines could also be used in vivo, especially as their design progresses to allow two-photon, near-infrared and radio-frequency activation[10]
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    Perylene Diimide as a Precise Graphene-like Superoxide Dismutase Mimetic
    (American Chemical Society, 2017) Jalilov, Almaz S.; Nilewski, Lizanne G.; Berka, Vladimir; Zhang, Chenhao; Yakovenko, Andrey A.; Wu, Gang; Kent, Thomas A.; Tsai, Ah-Lim; Tour, James M.; The NanoCarbon Center
    Here we show that the active portion of a graphitic nanoparticle can be mimicked by a perylene diimide (PDI) to explain the otherwise elusive biological and electrocatalytic activity of the nanoparticle construct. Development of molecular analogues that mimic the antioxidant properties of oxidized graphenes, in this case the poly(ethylene glycolated) hydrophilic carbon clusters (PEG–HCCs), will afford important insights into the highly efficient activity of PEG–HCCs and their graphitic analogues. PEGylated perylene diimides (PEGn–PDI) serve as well-defined molecular analogues of PEG–HCCs and oxidized graphenes in general, and their antioxidant and superoxide dismutase-like (SOD-like) properties were studied. PEGn–PDIs have two reversible reduction peaks, which are more positive than the oxidation peak of superoxide (O2•–). This is similar to the reduction peak of the HCCs. Thus, as with PEG–HCCs, PEGn–PDIs are also strong single-electron oxidants of O2•–. Furthermore, reduced PEGn–PDI, PEGn–PDI•–, in the presence of protons, was shown to reduce O2•– to H2O2 to complete the catalytic cycle in this SOD analogue. The kinetics of the conversion of O2•– to O2 and H2O2 by PEG8–PDI was measured using freeze-trap EPR experiments to provide a turnover number of 133 s–1; the similarity in kinetics further supports that PEG8–PDI is a true SOD mimetic. Finally, PDIs can be used as catalysts in the electrochemical oxygen reduction reaction in water, which proceeds by a two-electron process with the production of H2O2, mimicking graphene oxide nanoparticles that are otherwise difficult to study spectroscopically.
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    Preferential uptake of antioxidant carbon nanoparticles by T lymphocytes for immunomodulation
    (Springer Nature, 2016) Huq, Redwan; Samuel, Errol L.G.; Sikkema, William K.A.; Nilewski, Lizanne G.; Lee, Thomas; Tanner, Mark R.; Khan, Fatima S.; Porter, Paul C.; Tajhya, Rajeev B.; Patel, Rutvik S.; Inoue, Taeko; Pautler, Robia G.; Corry, David B.; Tour, James M.; Beeton, Christine; The NanoCarbon Center
    Autoimmune diseases mediated by a type of white blood cell—T lymphocytes—are currently treated using mainly broad-spectrum immunosuppressants that can lead to adverse side effects. Antioxidants represent an alternative approach for therapy of autoimmune disorders; however, dietary antioxidants are insufficient to play this role. Antioxidant carbon nanoparticles scavenge reactive oxygen species (ROS) with higher efficacy than dietary and endogenous antioxidants. Furthermore, the affinity of carbon nanoparticles for specific cell types represents an emerging tactic for cell-targeted therapy. Here, we report that nontoxic poly(ethylene glycol)-functionalized hydrophilic carbon clusters (PEG-HCCs), known scavengers of the ROS superoxide (O2•−) and hydroxyl radical, are preferentially internalized by T lymphocytes over other splenic immune cells. We use this selectivity to inhibit T cell activation without affecting major functions of macrophages, antigen-presenting cells that are crucial for T cell activation. We also demonstrate the in vivo effectiveness of PEG-HCCs in reducing T lymphocyte-mediated inflammation in delayed-type hypersensitivity and in experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis. Our results suggest the preferential targeting of PEG-HCCs to T lymphocytes as a novel approach for T lymphocyte immunomodulation in autoimmune diseases without affecting other immune cells.
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    Total Synthesis of Viridicatumtoxin B and Analogues Thereof: Strategy Evolution, Structural Revision, and Biological Evaluation
    (American Chemical Society, 2014) Nicolaou, K.C.; Hale, Christopher R.H.; Nilewski, Christian; Ioannidou, Heraklidia A.; ElMarrouni, Abdelatif; Nilewski, Lizanne G.; Beabout, Kathryn; Wang, Tim T.; Shamoo, Yousif
    The details of the total synthesis of viridicatumtoxin B (1) are described. Initial synthetic strategies toward this intriguing tetracycline antibiotic resulted in the development of key alkylation and Lewis acid-mediated spirocyclization reactions to form the hindered EF spirojunction, as well as Michael-Dieckmann reactions to set the A and C rings. The use of an aromatic A-ring substrate, however, was found to be unsuitable for the introduction of the requisite hydroxyl groups at carbons 4a and 12a. Applying these previous tactics, we developed stepwise approaches to oxidize carbons 12a and 4a based on enol- and enolate-based oxidations, respectively, the latter of which was accomplished after systematic investigations that revealed critical reactivity patterns. The herein described synthetic strategy resulted in the total synthesis of viridicatumtoxin B (1), which, in turn, formed the basis for the revision of its originally assigned structure. The developed chemistry facilitated the synthesis of a series of viridicatumtoxin analogues, which were evaluated against Gram-positive and Gram-negative bacterial strains, including drug-resistant pathogens, revealing the first structure-activity relationships within this structural type.