Radical-containing Nanoparticles Research Articles

Probing the inner structure and dynamics of pH-sensitive block copolymer nanoparticles with nitroxide radicals using scattering and EPR techniques

This research emphasizes the crucial role of electron paramagnetic resonance (EPR) spectroscopy in nanoparticle analysis, showcasing its distinctive ability to probe molecular-level details. We demonstrate the synthesis and self-assembly of a new class of pH-responsive amphiphilic diblock copolymers, specifically poly[N-(2-hydroxypropyl)-methacrylamide]-block-poly[2-(diisopropylamino)ethyl methacrylate] (PHPMA-b-PDPA), incorporating 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) radicals covalently bound to the hydrophilic PHPMA segment. The copolymer synthesis involved a three-step process, combining reversible addition − fragmentation chain transfer (RAFT) polymerization with carbodiimide (DCC) chemistry. TEMPO radical-containing nanoparticles (RNPs) were created using microfluidic (MF) nanoprecipitation, a technique essential for generating uniform nanoparticles with predictable biodistribution and cellular uptake. Adjusting MF protocol parameters allowed fine-tuning of RNP sizes. EPR spectroscopy confirmed the formation of core–shell RNPs, with features aligning closely with those obtained from scattering techniques like dynamic light scattering (DLS), static light scattering (SLS), small-angle X-ray scattering (SAXS), and cryo-transmission electron microscopy (cryo-TEM). EPR spectroscopy emerges as a potent tool for molecular-level analysis of colloidal polymer systems. This study highlights the EPR-spin label method’s efficacy in probing the internal structural features and dynamics of core–shell nanoparticles, especially their response to pH-triggered disassembly. The findings open new avenues for the use of pH-responsive TEMPO-labeled diblock copolymers in controlled drug delivery applications.

Nitroxide radical-containing redox nanoparticles protect neuroblastoma SH-SY5Y cells against 6-hydroxydopamine toxicity

Nitroxide radical-containing redox nanoparticles protect neuroblastoma SH-SY5Y cells against 6-hydroxydopamine toxicity

Management of tumor growth and angiogenesis in triple-negative breast cancer by using redox nanoparticles

In cancer, angiogenesis is a critical phenomenon of nascent blood vessel development to facilitate the oxygen and nutrient supply prerequisite for tumor progression. Therefore, targeting tumors at the angiogenesis step may be significant to prevent their advanced progression and metastasis. Although angiogenesis inhibitors can limit the further growth of tumors, complete eradication of tumors may not be possible by monotherapy alone. Therefore, a therapeutic regimen targeting both tumor growth and its vasculature is essential. Because reactive oxygen species (ROS) are fundamental to both angiogenesis and tumor growth, the use of antioxidants may be an effective dual approach to inhibit tumors. We previously confirmed that our original antioxidant nitroxide radical-containing nanoparticles (RNPs) such as pH-sensitive RNPN, and pH-insensitive RNPO, effectively attenuates the tumorigenic and metastasis potentials of triple-negative breast cancer. In this study, we further investigated the efficacy of RNPs to limit the tumor progression by inhibiting the ROS-regulated cancer angiogenesis in a triple-negative breast cancer model. Here, we confirmed that RNPs significantly inhibited in vitro angiogenesis, attributed to the downregulation of the ROS-regulated angiogenesis inducer, vascular endothelial growth factor (VEGF) in the breast cancer cell line (MDA-MB231) and human umbilical vein endothelial cells (HUVEC), which was consistent with decreased cellular ROS. TEMPOL, a low-molecular-weight (LMW) control antioxidant, exhibited anti-angiogenic effects accompanied by cytotoxicity to the endothelial cells. In an in vivo xenograft model for breast cancer, RNPs exerted significant anti-tumor effect due to the decreased expression of tumor VEGF, which prevented accumulation of the endothelial cells. It should be noted that such efficacy of RNPs was obtained with negligible off-target effects. On the other hand, TEMPOL, because of its size, exerted anti-angiogenesis effect accompanied with injuries to the kidneys, which corroborated with previous reports. Our findings imply that RNPs are more potential antioxidants than their LMW counterparts, such as TEMPOL, for the management of breast cancers.

Novel neuroprotection using antioxidant nanoparticles in a mouse model of head trauma.

Free radicals and reactive oxygen species are related to deteriorating pathological conditions after head trauma because of their secondary effects. 2,2,6,6-Tetramethylpiperidine-1-oxyl (TEMPO) scavenges free radicals; however, this molecule is also toxic. Here, we have evaluated the neuroprotective effect of antioxidant nanoparticles, which consisted of a novel core-shell type nanoparticle containing 4-amino-TEMPO, that is, redox-active nitroxide radical-containing nanoparticles (RNPs). Institute of Cancer Research mice were subjected to a head-impact procedure, randomly divided into four groups and intravenously (3 mg/kg) administered phosphate-buffered saline, TEMPO, micelle (a self-assembling block copolymer micelle without a TEMPO moiety), or RNP through the tail vein immediately thereafter and intraperitoneally at days 1, 3, and 5 after traumatic brain injury (TBI). The RNP distribution was detected by rhodamine labeling. Cognitive behavior was assessed using the neurological severity score and a rotarod test at days 1, 3, and 7 following TBI, and contusion volume was measured at day 7 after TBI. Free radical-scavenging capacity was analyzed by electron paramagnetic resonance on day 1 after TBI, and immunostaining was used to observe mobilization of microglia (Iba-1) and rescued neuronal cells (NeuN). Redox-active nitroxide radical-containing nanoparticle was detected in the microvessels around the injured area in the brain. Cognitive behavior assessment was significantly better, and contusion volume was significantly smaller in the RNP group compared with the other groups. Superoxide anion scavenging capacity was significantly higher in the RNP group, and neuronal loss was significantly suppressed around the injured area at day 7 after TBI. Furthermore, in the RNP group, neurodegenerative microglia production was suppressed at days 3 and 7 after TBI, whereas neuroprotective microglia production was higher at day 7 after TBI. The RNP administration after TBI improved cognitive behavior and reduced contusion volume by improving reactive oxygen species scavenging capacity. Therefore, RNP may have a neuroprotective effect after TBI. Therapeutic test.

Nitroxide radical-containing nanoparticles attenuate tumorigenic potential of triple negative breast cancer

The critical importance of reactive oxygen species (ROS) as oncogene activators and essential secondary messengers in cancer cell survival have been widely reported. Since oxidative stress has been implicated as being pivotal in various cancers, antioxidant therapy seems an apt strategy to abrogate ROS-mediated cellular processes to attenuate cancers. We therefore synthesized ROS scavenging nitroxide radical-containing nanoparticles (RNPs); pH insensitive RNPO and pH sensitive RNPN, to impede the proliferative and metastatic characteristics of the triple negative breast cancer cell line, MDA-MB-231, both in vitro and in vivo. RNPs significantly curtailed the proliferative and clonogenic potential of MDA-MB-231 and MCF-7 cell lines. Inhibition of ROS-mediated migratory and invasive characteristics of MDA-MB-231, via down regulation of NF-κB and MMP-2, was also confirmed. Furthermore, a significant anti-tumor and anti-metastatic potential of RNPs was observed in an MDA-MB-231 mouse xenograft model. Such tumoricidal effects of RNPs were attained with negligible adverse effects, compared to conventional low molecular weight antioxidants, TEMPOL. Thus, the tumoricidal effects of RNPs are suggestive of insights on precedence of nanoparticle-based therapeutics over current low molecular weight antioxidants to curtail ROS-induced tumorigenesis of various cancers.

OP-41 - Nitroxide radical-containing nanoparticles impairs the tumorigenic potential of triple negative breast cancer

Persistent elevated intracellular reactive oxygen species (ROS), requisite for cancerous phenotypes, has been known to affect cancers in a positive feedback manner, by playing critical roles in survival, proliferation and metastasis processes. Hence, scavenging of ROS to abrogate cell survival signaling pathways may therefore offer a potential therapeutic regime to mitigate the progression of various cancers. We therefore synthesized ROS scavenging nitroxide radical-containing nanoparticles (RNPs); pH insensitive RNPO and pH sensitive RNPN, to impede the proliferative and metastatic characteristics of the triple negative breast cancer cell line, MDA-MB-231, both in vitro and in vivo. RNPs significantly inhibited proliferation, colony forming potential, migration and invasion of MDA-MB-231 cells, accompanied by decrease in ROS level. Furthermore, anti-tumor and anti-metastatic potential of RNPs was observed in an MDA-MB-231 xenograft mouse model via downregulation of ROS-sensitive transcriptional factor, NF-κB and migration regulator, MMP-2 with negligible adverse effect compared to low molecular weight TEMPOL and paclitaxel. Conclusively, our findings provide insights into the use of nanoparticle over low molecular weight antioxidants in the redox-based cancer therapeutics.

P-315 - Cytodiagnosis of hypersialylated metastatic cancers by phenylboronic acid-installed PEGylated gold nanoparticle and their management by nitroxide radical-containing nanoparticles

Correlation between metastatic incidence and aberrantly high surface expression of reactive oxygen species (ROS) regulated sialic acid (hypersialylation) on tumors accentuates the sialyation signaling pathways as venerable targets for cytodiagnosis and redox therapeutics of hypersialyated metastatic cancers. Hence, in this line, we herein report on the diagnosis of hypersialylated tumors by surface-enhanced Raman spectroscopy (SERS) by phenylboronic acid-installed PEGylated gold nanoparticles coupled with Toluidine blue O (T/BA-GNPs) and redox cancer therapeutics by our original ROS scavenging polymeric micelle, nitroxide-radical containing nanoparticles (RNPs). In this study, we confirmed specific high SERS signal to noise ratio from metastatic cancer cell lines (MDA-MB-231 and Colon-26) and tumors in contrast to non-metastatic cancer cells (MCF-7) and healthy tissues. Furthermore, we also confirmed reduction in sialic acid-associated SERS signals from MDA-MB-231 surface upon treatment with RNPs, which was mediated by attenuation of ROS sensitive NF-κB-sialyltransferase signaling cascade. The aforementioned findings thus establish T/BA-GNPs based SERS as a potential cytodiagnostic system and RNPs as a promising antioxidant nanotherapeutics for hypersialylated metastatic tumors.

Abstract WMP119: Neurovascular Unit Protection Using Nitroxide Radicals-containing Nanoparticles (TEMPO-RNP) for Cerebral Ischemia-reperfusion Injury in Mice

Purpose: Reperfusion injury after thrombolytic therapy or mechanical thrombectomy would adverse neurological outcome. Generation of reactive oxygen species (ROS) due to reperfusion is relevant for aggravation. The development of drug therapy for scavenging free radical is expected to lead to better neurological prognosis. We developed a new core-shell type nanoparticle, RNP (radical containing nanoparticle), which consists of micelle containing 4-amino-TEMPO. The purpose of this study is to evaluate an efficacy of intra-arterial injection of RNP after reperfusion using tMCAO mice model. Methods: C57BL/6J mice underwent tMCAO for 60 minutes and received RNP by intra-arterial injection from common carotid artery. We evaluated infarction size, neurological scale, and Evans blue extravasation at 24h after reperfusion. RNP distribution was detected by rhodamine labeling. We further examined immunofluorescense of CD31, Occludin, TUNEL, dihydroethydium, and 8-hydroxy-deoxyguanosine (8-OHdG). Multiple free radical scavenging capacity of ischemic hemisphere was analyzed by Electron Paramagnetic Resonance (EPR). Results: RNPs were detected in endothelial cells and around neuronal cells in the ischemic lesion. The infarction size, neurological scale, Evans blue extravasation of the mice treated by RNP were significantly lower than the mice treated by PBS , control group. Intra-arterial RNP treatment preserved endothelium and expression of occludin in the ischemic brain. Further, apoptosis of neuronal cells and production of superoxide anions and 8OHdG were suppressed. By using EPR, multiple ROS scavenging capacity (•OH, •ROO, •O 2 - ) of the ischemic brain treated by RNP were higher. Conclusions: These results suggest that intra-arterial injection of RNP have efficacy of neurovascular unit protection and reduce infarction volume by improving multiple ROS scavenging capacity.

Novel Synthesized Radical-Containing Nanoparticles Limit Infarct Size Following Ischemia and Reperfusion in Canine Hearts.

Although nitroxyl radicals such as 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) scavenge free radicals, their short half-life and considerable side effects such as systemic hypotension and bradycardia have limited their clinical application. Since a radical-containing nanoparticle (RNP) delivers nitroxyl radicals with a prolonged half-life specific to ischemic hearts, we investigated whether RNPs reduce infarct size without the occurrence of substantial side effects and whether nitric oxide (NO) contributes to the cardioprotective effects of RNPs. The left anterior descending coronary arteries of dogs were occluded for 90min, followed by reperfusion for 6h. Either RNPs, micelles (not containing TEMPO) (control), or 4-hydroxy-TEMPO (TEMPOL) was injected into a systemic vein for 5min before reperfusion. We evaluated the infarct size, myocardial apoptosis, plasma NO levels in coronary venous blood, and the RNP spectra using an electron paramagnetic resonance assay. RNPs reduced infarct size compared with the control group and TEMPOL group (19.5±3.3 vs. 42.2±3.7 vs. 30.2±3.4%). RNPs also reduced myocardial apoptosis compared with the control and TEMPOL group. Coronary venous NO levels increased in the RNP group. In conclusion, the administration of 2,2,6,6-tetramethylpiperidine-1-oxyl as a RNP exerted cardioprotective effects against ischemia and reperfusion injury in canine hearts without exerting unfavorable hemodynamic effects. RNPs may represent a promising new therapy for patients with acute myocardial infarction.

Neurovascular Unit Protection From Cerebral Ischemia-Reperfusion Injury by Radical-Containing Nanoparticles in Mice.

Reperfusion therapy by mechanical thrombectomy is used to treat acute ischemic stroke. However, reactive oxygen species generation after reperfusion therapy causes cerebral ischemia-reperfusion injury, which aggravates cerebral infarction. There is limited evidence for clinical efficacy in stroke for antioxidants. Here, we developed a novel core-shell type nanoparticle containing 4-amino-4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (nitroxide radical-containing nanoparticles [RNPs]) and investigated its ability to scavenge reactive oxygen species and confer neuroprotection. C57BL/6J mice underwent transient middle cerebral artery occlusion and then received RNPs (9 mg/kg) through the common carotid artery. Infarction size, neurological scale, and blood-brain barrier damage were visualized by Evans blue extravasation 24 hours after reperfusion. RNP distribution was detected by rhodamine labeling. Blood-brain barrier damage, neuronal apoptosis, and oxidative neuronal cell damage were evaluated in ischemic brains. Multiple free radical-scavenging capacities were analyzed by an electron paramagnetic resonance-based method. RNPs were detected in endothelial cells and around neuronal cells in the ischemic lesion. Infarction size, neurological scale, and Evans blue extravasation were significantly lower after RNP treatment. RNP treatment preserved the endothelium and endothelial tight junctions in the ischemic brain; neuronal apoptosis, O2- production, and gene oxidation were significantly suppressed. Reactive oxygen species scavenging capacities against OH, ROO, and O2- improved by RNP treatment. An intra-arterial RNP injection after cerebral ischemia-reperfusion injury reduced blood-brain barrier damage and infarction volume by improving multiple reactive oxygen species scavenging capacities. Therefore, RNPs can provide neurovascular unit protection.

Nitroxide radical-containing nanoparticles as potential candidates for overcoming drug resistance in epidermoid cancers

Multidrug resistance in cancer cells contributes to the failure of conventional chemotherapy in more than 90% of cancer patients (metastatic). This is attributed to reactive oxygen species (ROS)-regulated drug efflux proteins, P-glycoprotein (P-gp) and multidrug resistance-associated protein 1 (MRP1). In this study, we focused on overcoming multidrug resistance with a therapeutic application of ROS-scavenging nitroxide radical-containing nanoparticles, RNPN (pH-sensitive) and RNPO (pH-insensitive), in combination with the conventional chemotherapeutic drug, doxorubicin (Dox), in drug-resistant epidermoid cancer cell lines, KB-C2 (P-gp expressing) and KB/MRP (MRP1 expressing). We confirmed that the combination treatment with RNPs increased Dox uptake in multidrug-resistant cancer cells, which further enhanced cell cytotoxicity. The abrogation of the crucial ROS signaling was confirmed with RNP treatment, which deterred ROS-regulated drug efflux protein (P-gp and MRP1) expression, resulting in the sensitization of resistant cells to Dox. These results establish ROS-scavenging RNPs as potential therapeutic candidates to overcome drug resistance in multidrug-resistant cancers.

Synthesis and Solution Properties of PCL-b-PHPMA Diblock Copolymers Containing Stable Nitroxyl Radicals

This work focuses on the synthesis and the aqueous solution properties of novel amphiphilic PCL-b-PHPMA diblock copolymers possessing 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) stable radicals covalently conjugated to the hydrophobic poly(e-caprolactone) (PCL) block. A new synthetic approach (a four-step pathway) combining ring-opening polymerization (ROP), carbodiimide chemistry (DCC method), a reversible addition–fragmentation chain transfer (RAFT) polymerization technique, and finally chemical oxidation was employed to successfully produce a series of TEMPO-containing PCL-b-PHPMA diblock copolymers for the first time. EPR spectroscopy was applied to verify successful oxidation of the synthesized diblock copolymers and to investigate the dynamics of the polymer chains before and after micellization. The diblock copolymers self-assembled in PBS solution into spherical radical-containing nanoparticles (RNPs), which were characterized by 1H NMR spectroscopy, dynamic (DLS), static (SLS) light scattering, ...

Radical-containing nanoparticles with ph and redox dual sensitive properties

Radical-containing nanoparticles with ph and redox dual sensitive properties

Silica-installed redox nanoparticles for novel oral nanotherapeutics – improvement in intestinal delivery with anti-inflammatory effects

Silica nanoparticles were synthesized via a sol–gel method in which tetraethyl orthosilicate was hydrolyzed by the alkaline core of the nitroxide radical-containing nanoparticle (RNP). The silica nanoparticles were successively captured in the RNP core to obtain silica/RNP nanocomposite (siRNP). Alternatively, siRNP was prepared using commercially available silica nanoparticles. The amount of elemental Si present in the siRNPs was controlled from 3 wt% to 12 wt%. Notably, the obtained siRNPs were stable in acidic media, whereas the starting RNP disintegrated immediately. Crosslinking of the RNP by the entrapped silica might improve stability of the siRNPs under such acidic conditions. Rebamipide was found to be stably encapsulated in the cores of the prepared siRNPs even under acidic conditions, probably due to the both basic environment of the cores and absorption tendencies of the entrapped silica. Under neutral to alkaline conditions, release of the rebamipide is accelerated, which is probably due to the repulsion between the anionic silica surface and the anionic rebamipide. Rebamipide-loaded siRNPs (rebamipide@siRNP) were orally administered to mice, and the plasma level of rebamipide was checked at predetermined time intervals, showing a significantly higher uptake of rebamipide in the plasma when compared to orally-administered free rebamipide. Because siRNP possesses nitroxide radicals in the core, it is confirmed that dextran sodium sulfate-induced colon inflammation was effectively suppressed by the oral administration of rebamipide@siRNP in mice.

Abstract P2-09-22: Synthesis of nitroxyl radical-containing nanoparticles with pH-sensitive behavior and their application for cancer therapy

Abstract Extensive research has documented that redox dysregulation is a causative involvement in tumor progression, and reactive oxygen species (ROS) are involved in cancer initiation and progression. ROS are mutagenic and may promote cancer. Early studies observed that the increased ROS production was in response to oncogenic activity. It is now widely accepted that redox chemotherapeutics represents a novel class of promising anticancer agents. Small molecule nitroxides are stable free radicals which are able to scavenge radicals and act as radical chain reaction terminators. Nitroxyl radical, 2, 2, 6, 6-tetramethyl-piperidine-1-oxyl (TEMPO), is capable of reacting with ROS and shows chemotherapeutic efficacy in various cancer cell lines and xenograft models. However, low-molecular-weight TEMPO derivatives are hard to utilize in the clinic, owing to a preferential renal clearance and side effects such as antihypertensive effect. To address these issues, we have designed and developed nitroxyl radical-containing nanoparticles with pH-sensitive behavior using the first generation PAMAM (G1 PAMAM) partially modified with 3,4,5,6-tetrahydrophthalic anhydride (THPA) and PROXYL. The synthesis of the modified PAMAM was carried out by a two-step reaction. Firstly, part of the primary amine end group of PAMAM dendrimers reacted with anhydride THPA to produce a THPA-modified PAMAM. The b-carboxylic acid amides between G1 PAMAM and THPA are acid-sensitive. Therefore, the THPA-modified PAMAM are stable and negatively charged at physiological pH, thus preventing blood serum aggregation. However, the THPA-modified PAMAM degrades at acidic pH, converting its surface to positive charge which is benefit for cellular uptake. Secondly, the THPA-modified PAMAM reacted subsequently with 3-carboxy-PROXYL whose carboxyl group was activated by N-hydroxysuccinimide. The EPR signal of the product was analyzed by EPR measurement and confirmed the PROXYL group covalently conjugated onto THPA-modified PAMAM. The obtained THPA-PROXYL-modified PAMAM was denoted as TP-PAMAM. Experimental results showed that TP-PAMAM was stable at pH 7.4, and hydrolyzed at pH 6.0. The rate of hydrolysis was increased with pH decreasing. In vitro studies indicated that cell proliferation of MDA-MB-231 was greatly inhibited by TP-PAMAM compared to low-molecular-weight TEMPO. This may due to the fact that cellular uptake is easy for nanoparticles with pH-sensitive behavior. Therefore, the nitroxyl radical conjugated onto the pH-sensitive nanoparticles scavenges ROS more effectively. As a result, these new nitroxyl radical-containing nanoparticles have a significantly high therapeutic efficacy in the application for cancer therapy. Hence, engineered nitroxyl radical-containing nanoparticles with pH-sensitive behavior may enable therapeutic selectivity and to overcome drug resistance and metastatic cancer. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-09-22.

An Orally Administered Redox Nanoparticle That Accumulates in the Colonic Mucosa and Reduces Colitis in Mice

Drugs used to treat patients with ulcerative colitis are not always effective because of nonspecific distribution, metabolism in the gastrointestinal tract, and side effects. We designed a nitroxide radical-containing nanoparticle (RNP(O)) that accumulates specifically in the colon to suppress inflammation and reduce the undesirable side effects of nitroxide radicals. RNP(O) was synthesized by assembly of an amphiphilic block copolymer that contains stable nitroxide radicals in an ether-linked hydrophobic side chain. Biodistribution of RNP(O) in mice was determined from radioisotope and electron spin resonance measurements. The effects of RNP(O) were determined in mice with dextran sodium sulfate (DSS)-induced colitis and compared with those of low-molecular-weight drugs (4-hydroxyl-2,2,6,6-tetramethylpiperidine-1-oxyl [TEMPOL] or mesalamine). RNP(O), with a diameter of 40 nm and a shell of poly(ethylene glycol), had a significantly greater level of accumulation in the colonic mucosa than low-molecular-weight TEMPOL or polystyrene latex particles. RNP(O) was not absorbed into the bloodstream through the intestinal wall, despite its long-term retention in the colon, which prevented its distribution to other parts of the body. Mice with DSS-induced colitis had significantly lower disease activity index and less inflammation following 7 days of oral administration of RNP(O) compared with mice with DSS-induced colitis or mice given low-molecular-weight TEMPOL or mesalamine. We designed an orally administered RNP(O) that accumulates specifically in the colons of mice with colitis and is more effective in reducing inflammation than low-molecular-weight TEMPOL or mesalamine. RNP(O) might be developed for treatment of patients with ulcerative colitis.

Abstract 2900: Redox nanoparticle enhances effect of anticancer chemotherapy and suppresses cardio-toxicity

Abstract Utilizing the self-assembled core-shell-type polymeric micelle technique, high-performance nanoparticles possessing stable nitroxide radicals in the core and reactive groups on the periphery were prepared (Nitroxide Radical-containing Nanoparticles; RNP). The average diameter of the nanoparticles was ca. 40 nm, and the nanoparticles emitted intense electron paramagnetic resonance (EPR) signals. Owing to the confinement of the nitroxide groups in the core of the nanoparticles, it reduces its toxicity, prolongs its blood circulation time. Under the low pH environment in vivo, it was confirmed that RNP disintegrated to expose the nitroxide radicals outside of the particle. It is sharp contrast against low-molecular-weight TEMPOL, which was reduced and excreted rapidly from the blood stream. Recently, we have reported that RNP is effectively worked to cerebral ischemia reperfusion injury. The RNP disintegrates in penumbra area due to the acidosis of ischemic region, followed by the scavenging reactive oxygen species (ROS) generated in this area. This strategy could be applied to other ischemic injuries such as renal and myocardial infarction. Quite recently, we have found that scavenging of ROS in tumor region by RNP improves efficiency of anticancer drug, doxorubicin (DOX). For example, RNP administration prior to the DOX-injection remarkably improved its efficiency. When RNP was administered via mice tail veil, it accumulated in tumor due to the enhancement permeation and retention (EPR) effect. The accumulated RNP scavenged excessive generated ROS in tumor, which suppressed activation of transcription factors such as NF-κB. Such suppression of inflammation related transcription factors might increase effectiveness of anticancer drug such as doxorubicin (DOX). The RNP pre-administration also reduced its side effect. Actually, 25mg/kg intravenous administrations of RNP before and after the DOX-injection (20mg/kg) reduced significantly the lactate dehydrogenase (LDH) and the creatine phosphokinase (CPK) activity. Thus, the nanoparticle anti-oxidant treatment is promising as a new adjuvant for anti-cancer chemotherapy without severe side effect by anticancer drug. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2900. doi:1538-7445.AM2012-2900

Nitroxide Radicals and Nanoparticles: a Partnership for Nanomedicine Radical Delivery

This article aims to provide a research update on nitroxide radical compounds for application of anti-oxidative stress therapy. Nitroxide compounds such as 2,2,6,6-tetramethylpyperidine-1-oxyl (TEMPO) can catalytically react with reactive oxygen species (ROS) and are anticipated as new anti-oxidant therapies for several diseases. However, low-molecular-weight nitroxide compounds pose several problems such as nonspecific dispersion in normal tissues, preferential renal clearance and rapid reduction of the nitroxide radical to the corresponding hydroxylamine. Nitroxide radical compounds are also known to show dose-related antihypertensive action accompanied by reflex tachycardia, increased skin temperature, and seizures. The author has recently designed novel nanoparticles, which possess nitroxide radicals in the core for novel bioimaging and nanotherapy. Nitroxide radical-containing nanoparticles (RNP) shows high safety, long blood circulation, magnetic resonance imaging and ESR imaging sensitive character and efficient therapeutic effects to several diseases such as cerebral and renal ischemia reperfusions, ulcerative colitis and Alzheimer's disease models. RNPs are, thus, promising as new nanotherapeutic materials.

Novel redox nanomedicine improves gene expression of polyion complex vector

Gene therapy has generated worldwide attention as a new medical technology. While non-viral gene vectors are promising candidates as gene carriers, they have several issues such as toxicity and low transfection efficiency. We have hypothesized that the generation of reactive oxygen species (ROS) affects gene expression in polyplex supported gene delivery systems. The effect of ROS on the gene expression of polyplex was evaluated using a nitroxide radical-containing nanoparticle (RNP) as an ROS scavenger. When polyethyleneimine (PEI)/pGL3 or PEI alone was added to the HeLa cells, ROS levels increased significantly. In contrast, when (PEI)/pGL3 or PEI was added with RNP, the ROS levels were suppressed. The luciferase expression was increased by the treatment with RNP in a dose-dependent manner and the cellular uptake of pDNA was also increased. Inflammatory cytokines play an important role in ROS generation in vivo. In particular, tumor necrosis factor (TNF)-α caused intracellular ROS generation in HeLa cells and decreased gene expression. RNP treatment suppressed ROS production even in the presence of TNF-α and increased gene expression. This anti-inflammatory property of RNP suggests that it may be used as an effective adjuvant for non-viral gene delivery systems.

The use of nitroxide radical-containing nanoparticles coupled with piperine to protect neuroblastoma SH-SY5Y cells from Aβ-induced oxidative stress

The antioxidant effect and potential mechanism of nitroxide radical-containing nanoparticles (RNPs) coupled with piperine (PI) were investigated in human neuroblastoma SH-SY5Y cells. The effects of RNP/PI on SH-SY5Y cell lines was determined by WST assay for cell viability, nitroblue tetrazolium and deoxyribose assay for reactive oxygen species generation, ELISA assay for reactive oxygen species products and apoptotic cell death, and biochemical techniques for catalase and glutathione peroxidase activity. The RNP/PI significantly reduced the reactive oxygen species level and reactive oxygen species products compared with those of cells treated with RNPs alone. The RNP/PI treatment enhanced catalase and glutathione peroxidase activity. The combination of RNP/PI has been found to have an augmented antioxidant effect on an Alzheimer’s model in vitro. The mechanism of the protective effect of this combination therapy was correlated in this study with its ability to reduce the generation of reactive oxygen species and prevent apoptosis via scavenging enzyme action pathways.