Risdiplam in Type 1 Spinal Muscular Atrophy.

BACKGROUND: Type 1 spinal muscular atrophy is a rare, progressive neuromuscular disease that is caused by low levels of functional survival of motor neuron (SMN) protein. Risdiplam is an orally administered, small molecule that modifies SMN2 pre-messenger RNA splicing and increases levels of functional SMN protein. METHODS: We report the results of part 1 of a two-part, phase 2-3, open-label study of risdiplam in infants 1 to 7 months of age who had type 1 spinal muscular atrophy, which is characterized by the infant not attaining the ability to sit without support. Primary outcomes were safety, pharmacokinetics, pharmacodynamics (including the blood SMN protein concentration), and the selection of the risdiplam dose for part 2 of the study. Exploratory outcomes included the ability to sit without support for at least 5 seconds. RESULTS: A total of 21 infants were enrolled. Four infants were in a low-dose cohort and were treated with a final dose at month 12 of 0.08 mg of risdiplam per kilogram of body weight per day, and 17 were in a high-dose cohort and were treated with a final dose at month 12 of 0.2 mg per kilogram per day. The baseline median SMN protein concentrations in blood were 1.31 ng per milliliter in the low-dose cohort and 2.54 ng per milliliter in the high-dose cohort; at 12 months, the median values increased to 3.05 ng per milliliter and 5.66 ng per milliliter, respectively, which represented a median of 3.0 times and 1.9 times the baseline values in the low-dose and high-dose cohorts, respectively. Serious adverse events included pneumonia, respiratory tract infection, and acute respiratory failure. At the time of this publication, 4 infants had died of respiratory complications. Seven infants in the high-dose cohort and no infants in the low-dose cohort were able to sit without support for at least 5 seconds. The higher dose of risdiplam (0.2 mg per kilogram per day) was selected for part 2 of the study. CONCLUSIONS: In infants with type 1 spinal muscular atrophy, treatment with oral risdiplam led to an increased expression of functional SMN protein in the blood. (Funded by F. Hoffmann-La Roche; ClinicalTrials.gov number, NCT02913482.).

Pharmaceutical Properties of a Tinted Formulation of a Biguanide Antiseptic Agent, Olanexidine Gluconate.

Olanexidine gluconate-containing preoperative antiseptic (OLG-C) is colorless, which makes it difficult to determine its area of application. To overcome this drawback, we realized a stable orange-tinted antiseptic (OLG-T) by adding new additives to OLG-C and investigated its pharmaceutical properties compared with OLG-C and povidone iodine (PVP-I). We evaluated the influence of the additives on the antimicrobial activity and adhesiveness of medical adhesives to OLG-T-applied skin by in vitro time-kill/ex vivo micropig skin assays and a peel test using excised micropig skin, respectively. In the in vitro time-kill assay, the bactericidal/fungicidal activity of OLG-T and OLG-C were equivalent. In the ex vivo micropig skin assay, their fast-acting and persistent bactericidal activities against vancomycin-resistant Enterococcus faecalis were higher than that of PVP-I. In the peel test, the adhesion force of the incise drape and the amount of stripped corneocytes on the peeled drape were comparable between OLG-T- and OLG-C-applied skin, but both were less than those of PVP-I-applied skin. The drapes for OLG-T- and OLG-C-applied skin had moderate adhesion force, and the drape-related injuries were expected to be weak. These results suggest that OLG-T performs no worse than OLG-C in terms of its antimicrobial activity and medical adhesive compatibility. Therefore, we expect OLG-T to lead to more convenient preoperative skin preparation and further contribute to lowering surgical site infection rates.

Combination therapy with onasemnogene and risdiplam in spinal muscular atrophy type 1.

INTRODUCTION/AIMS: There are currently three medications approved for spinal muscular atrophy (SMA), but the use of these medications in combination has not been well described. METHODS: This is a retrospective report of four cases of SMA treated with dual onasemnogene and risdiplam therapy at our institution. RESULTS: Following onasemnogene therapy, all four patients experienced a perceived plateau of therapeutic benefit, at which time daily risdiplam was started. Transient fatigue and weakness was seen in two patients following risdiplam initiation, but this resolved within 1 mo. One patient was hospitalized with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and post-viral pneumonia, weeks following risdiplam initiation. No other adverse effects related to onasemnogene and risdiplam combination therapy were identified and all patients experienced objective and subjective improvement. DISCUSSION: Combination therapy with onasemnogene and risdiplam in patients with SMA appears to be well-tolerated. Further large prospective trials are needed to determine whether dual therapy is more efficacious than monotherapy, and to identify rare adverse events that may occur with the use of combination therapy.

Selecting disease-modifying medications in 5q spinal muscular atrophy.

Spinal muscular atrophy (SMA) is an inherited lower motor neuron disease. SMA occurs secondary to alterations in the survival motor neuron 1 gene (SMN1), which is the main driver of SMN protein production. The severity of the disease is determined by the number of copies of the SMN2 gene, which is a homolog to SMN1 but not as efficient in protein production. Three medications have recently been approved for the treatment of SMA. Nusinersen is an intrathecal antisense oligonucleotide that alters SMN2 pre-mRNA, onasemnogene abeparvovec-xioi is an intravenous SMN1 gene replacement therapy, and risdiplam is an oral small molecule splicing modifier of SMN2. No head-to-head studies have been conducted comparing these medications, so selection of one of these medications for an individual with SMA can be challenging. In this article we outline the efficacy, safety, and other pertinent factors to consider when selecting a therapy for an individual with SMA. The age of the individual and comorbidities, such as liver or kidney disease, help guide treatment choices. All three of these medications are efficacious, and early initiation is critical for obtaining the best outcomes.

Patch testing with a textile dye mix with and without Disperse Orange 3.

BACKGROUND: The textile dye mix (TDM) 6.6% pet. contains Disperse Blue (DB) 35, Disperse Yellow 3, Disperse Orange (DO) 1 and 3, Disperse Red 1 and 17, and DB 106 and 124. The most frequent allergen in TDM-positive patients is DO 3. Around 85% of p-phenylenediamine (PPD)-allergic dermatitis patients have shown positive patch test reactions to DO 3. There has been a discussion to exclude DO 3 from TDM 6.6% because of frequent, strong reactions to TDM 6.6% and PPD. OBJECTIVES: To study if DO 3 can be omitted from a TDM. METHODS: Patch tests were performed on 2250 dermatitis patients with TDM 6.6%, TDM 5.6% pet., TDM 7.0% pet., and PPD 1.0% pet.; 122 patients were also patch tested with DO 3 1.0% pet. RESULTS: Among the 2250 patients patch tested, contact allergy prevalence to TDM 6.6% was 2.4%, to TDM 5.6% 1.8%, and to TDM 7.0% 2.0%. Of the 54 TDM 6.6%-positive patients, 55.6% reacted to PPD; as much as 42.2% of PPD-allergic women and 50% of PPD-allergic men reacted to TDM 6.6%. Of the 17 DO 3-positive patients, 94.1% showed a positive reaction to PPD. CONCLUSION: Results indicate that DO 3 can probably be omitted from TDM, but patch testing with TDM 6.6%, TDM 7.0%, DO 3 1.0%, and PPD 1.0% simultaneously is needed to finally decide whether it is possible or not.

A phase 1 healthy male volunteer single escalating dose study of the pharmacokinetics and pharmacodynamics of risdiplam (RG7916, RO7034067), a SMN2 splicing modifier.

AIMS: Risdiplam (RG7916, RO7034067) is an orally administered, centrally and peripherally distributed, survival of motor neuron 2 (SMN2) mRNA splicing modifier for the treatment of spinal muscular atrophy (SMA). The objectives of this entry-into-human study were to assess the safety, tolerability, pharmacokinetics (PK) and pharmacodynamics of risdiplam, and the effect of the strong CYP3A inhibitor itraconazole on the PK of risdiplam in healthy male volunteers. METHODS: Part 1 had a randomized, double-blind, adaptive design with 25 subjects receiving single ascending oral doses of risdiplam (ranging from 0.6-18.0 mg, n = 18) or placebo (n = 7). A Bayesian framework was applied to estimate risdiplam's effect on SMN2 mRNA. The effect of multiple doses of itraconazole on the PK of risdiplam was also assessed using a two-period cross-over design (n = 8). RESULTS: Risdiplam in the fasted or fed state was well tolerated. Risdiplam exhibited linear PK over the dose range with a multi-phasic decline with a mean terminal half-life of 40-69 h. Food had no relevant effect, and itraconazole had only a minor effect on plasma PK indicating a low fraction of risdiplam metabolized by CYP3A. The highest tested dose of 18.0 mg risdiplam led to approximately 41% (95% confidence interval 27-55%) of the estimated maximum increase in SMN2 mRNA. CONCLUSIONS: Risdiplam was well tolerated and proof of mechanism was demonstrated by the intended shift in SMN2 splicing towards full-length SMN2 mRNA. Based on these data, Phase 2/3 studies of risdiplam in patients with SMA are now ongoing.

The nitric oxide donor JS-K sensitizes U87 glioma cells to repetitive irradiation.

As a potent radiosensitizer nitric oxide (NO) may be a putative adjuvant in the treatment of malignant gliomas which are known for their radio- and chemoresistance. The NO donor prodrug JS-K (O2-(2.4-dinitrophenyl) 1-[(4-ethoxycarbonyl) piperazin-1-yl] diazen-1-ium-1,2-diolate) allows cell-type specific intracellular NO release via enzymatic activation by glutathione-S-transferases overexpressed in glioblastoma multiforme. The cytotoxic and radiosensitizing efficacy of JS-K was assessed in U87 glioma cells in vitro focusing on cell proliferation, induction of DNA damage, and cell death. In vivo efficacy of JS-K and repetitive irradiation were investigated in an orthotopic U87 xenograft model in mice. For the first time, we could show that JS-K acts as a potent cytotoxic and radiosensitizing agent in U87 cells in vitro. This dose- and time-dependent effect is due to an enhanced induction of DNA double-strand breaks leading to mitotic catastrophe as the dominant form of cell death. However, this potent cytotoxic and radiosensitizing effect could not be confirmed in an intracranial U87 xenograft model, possibly due to insufficient delivery into the brain. Although NO donor treatment was well tolerated, neither a retardation of tumor growth nor an extended survival could be observed after JS-K and/or radiotherapy.

Evaluation of genetic toxicity of 6-diazo-5-oxo-l-norleucine (DON).

DON (6-diazo-5-oxo-l-norleucine), a glutamine antagonist, was demonstrated to exhibit analgesic, antibacterial, antiviral and anticancer properties. The study was performed to characterize its in vitro and in vivo genetic toxicity potential. DON was tested in the bacterial reverse mutation assay (Ames test) using Salmonella typhimurium tester strains (TA98, TA100, TA1535 and TA1537) and Escherichia coli tester strain (WP2 uvrA) with and without S9 and also with reductive S9. In addition, DON was tested for the chromosome aberrations in Chinese hamster ovary (CHO) cells with or without S9 to evaluate the clastogenic potential. Furthermore, DON was also evaluated for its in vivo clastogenic activity by detecting micronuclei in polychromatic erythrocyte (PCE) cells in bone marrow collected from the male mice dosed intravenously with 500, 100, 10, 1 and 0.1 mg/kg at 24 and 48-h post-dose. The Ames mutagenicity assay showed no positive mutagenic responses. However, the in vitro chromosome aberration assay demonstrated dose dependent statistically positive increase in structural aberrations at 4 and 20-h exposure without S9 and also at 4-h exposure with S9. The in vivo micronucleus assay also revealed a statistically positive response for micronucleus formation at 500, 100 and 10 mg/kg at 24 and 48-h post-dose. Thus, DON appears to be negative in the Ames test but positive in the in vitro chromosome aberration assay and in the in vivo micronucleus assay. In conclusion, the results indicate DON is a genotoxic compound with a plausible epigenetic mechanism.

Overexpression of FOXO1 ameliorates the podocyte epithelial-mesenchymal transition induced by high glucose in vitro and in vivo.

Accumulating evidence has suggested that the epithelial-mesenchymal transition (EMT) is a pathway that potentially leads to podocyte depletion and proteinuria in diabetic nephropathy (DN). Therefore, this study was designed to investigate the protective effects of forkhead transcription factor O1 (FOXO1) on podocyte EMT, under high-glucose (HG) conditions in vitro and under diabetic conditions in vivo. The results showed that HG-induced podocyte EMT was associated with FOXO1 inactivation, which was accompanied by activation of the transforming growth factor (TGF)-ß1/SMAD3/integrin-linked kinase (ILK) pathway. Accordingly, constitutive FOXO1 activation suppressed the TGF-ß1/Smad3/ILK pathway and partially reversed EMT, similar to the effects observed after treatment with SIS3 or QLT0267, which are selective inhibitors of TGF-ß1-dependent SMAD3 phosphorylation and ILK, respectively. In addition, lentiviral-mediated FOXO1 overexpression in the kidneys of diabetic mice considerably increased FOXO1 expression and activation, while decreasing proteinuria and renal pathological injury. These data suggested that forced FOXO1 activation inhibited HG-induced podocyte EMT and ameliorated proteinuria and renal injury in diabetic mice. Our findings further highlighted that FOXO1 played a protective role against diabetes in mice and may potentially be used as a novel therapeutic target for treating diabetic nephropathy.

Encapsulation of N-Diazeniumdiolates within Liposomes for Enhanced Nitric Oxide Donor Stability and Delivery.

The rapid decomposition of nitric oxide (NO) donors in aqueous environments remains a limitation for applications requiring extended NO release. Herein, we report the synthesis of dipalmitoylphosphatidylcholine-based liposomes capable of extended NO release using low molecular weight NO donors and a reverse-phase evaporation technique. The encapsulation of the NO donors within the liposomes enabled both prolonged NO release and enhanced storage compared to free NO donors alone. The NO-releasing liposomes also demonstrated enhanced efficacy against human pancreatic cancer cells. These NO-release vehicles represent attractive anticancer therapeutics due to their potential to store the majority of their NO payload until reaching cancerous tissue at which time the lower pH inherent to such environments will trigger an avalanche of NO.

Effect of a Pluronic(®) P123 formulation on the nitric oxide-generating drug JS-K.

PURPOSE: O(2)-(2,4-dinitrophenyl)1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate] or JS-K is a nitric oxide-producing prodrug of the arylated diazeniumdiolate class with promising anti-tumor activity. JS-K has challenging solubility and stability properties. We aimed to characterize and compare Pluronic(®) P123-formulated JS-K (P123/JS-K) with free JS-K. METHODS: We determined micelle size, shape, and critical micelle concentration of Pluronic(®) P123. Efficacy was evaluated in vitro using HL-60 and U937 cells and in vivo in a xenograft in NOD/SCID IL2Rγ (null) mice using HL-60 cells. We compared JS-K and P123/JS-K stability in different media. We also compared plasma protein binding of JS-K and P123/JS-K. We determined the binding and Stern Volmer constants, and thermodynamic parameters. RESULTS: Spherical P123/JS-K micelles were smaller than blank P123. P123/JS-K formulation was more stable in buffered saline, whole blood, plasma and RPMI media as compared to free JS-K. P123 affected the protein binding properties of JS-K. In vitro it was as efficacious as JS-K alone when tested in HL-60 and U937 cells and in vivo greater tumor regression was observed for P123/JS-K treated NOD/SCID IL2Rγ (null) mice when compared to free JS-K-treated NOD/SCID IL2Rγ (null) mice. CONCLUSIONS: Pluronic(®) P123 solubilizes, stabilizes and affects the protein binding characteristics of JS-K. P123/JS-K showed more in vivo anti-tumor activity than free JS-K.

Lateral drug diffusion in human nails.

The main objective of the current work is to demonstrate the process of passive lateral diffusion in the human nail plate and its effect on the passive transungual permeation of antifungal drug ciclopirox olamine (CPO). A water soluble dye, methyl red sodium salt (MR) was used to visualize the process of lateral diffusion using a novel suspended nail experiment. The decline in concentration of CPO correlates with that of concentration of MR from the proximal to the distal end of the nail in suspended nail study. Three toenails each were trimmed to 5 mm × 5 mm (25 mm(2)), 7 mm × 7 mm (49 mm(2)), and 9 mm × 9 mm (81 mm(2)) to study the extent and effect of lateral diffusion of the CPO on its in vitro transungual permeation. The permeation flux of CPO decreased as the surface area of the toenail increased. There was a positive correlation between the concentrations of CPO and MR in the area of application and in the peripheral area of the toenails of the three surface areas, confirming the findings in the suspended nail experiment. Profound lateral diffusion of CPO was demonstrated and shown to reduce the in vitro passive transungual drug permeation and prolong the lag-time in human toenails. The study data implies that during passive in vitro transungual permeation experiments, the peripheral nail around the area of drug application has to be kept to a minimum, in order to get reliable data which mimics the in vivo situation.

T-cell potential of human adult and cord blood hemopoietic stem cells expanded with the use of aryl hydrocarbon receptor antagonists.

BACKGROUND AIMS: Expansion of hemopoietic stem cells (HSCs) in vitro is a potential strategy for improving transplant outcomes, but expansion methods tend to promote differentiation and loss of stem cell potential. Aryl hydrocarbon receptor antagonists (AhRAs) have recently been shown to protect HSC stemness during expansion; however, little is known of the T-cell regenerative capacity of AhRA-expanded HSCs. In this study, we confirm the protective effect of two commercially available AhRA compounds on HSCs from both cord blood (CB) and adult samples and assess the T-lymphocyte potential of the expanded cells. METHODS: Adult mobilized peripheral blood and CB samples were purified to CD34(+) cells, which were expanded in vitro with cytokines and AhRAs. After 14 d, CD34(+) cells were re-isolated and then grown on in OP9Delta co-culture under conditions that allow T-lymphocyte differentiation. Cells were monitored weekly for T-lineage markers by flow cytometry. RESULTS: Both AhRA compounds promoted maintenance of CD34 expression during 2 weeks of proliferation with growth factors, although adult cells proliferated markedly less than CB cells. AhRA-expanded CD34(+) cells from CB differentiated to T cells on OP9Delta co-culture with the same rate and time course as untreated cells. Adult cells, by contrast, had reduced differentiation to T cells, with donor-dependent variable responses. CONCLUSIONS: This study shows that whereas AhRA treatment is effective in CB samples, expansion of adult HSCs is less successful and reflects their inherent poor potential in T-cell generation.

Antioxidant responses in Phanerochaete chrysosporium exposed to Astrazone Red FBL textile dye.

Interest in environmental-pollutant-induced oxidative stress and knowledge of the interactions between reactive oxygen species and cellular systems have increased in toxicology and microbial ecology considerably in recent decades. These reactive oxidants are produced by a variety of environmental sources: ionizing radiations, ultraviolet light, redox cycling drugs, hyperoxia, ischemia and redox-active xenobiotics or during metabolism of environmental pollutants, such as heavy metals in mining industries, dyes in wastewater of textile industries, pesticides and polycyclic hydrocarbons, i.e. foreign materials. In this study, the effect of dye on the antioxidative defence system of Phanerochaete chrysosporium was investigated, and we showed the ability of Phanerochaete chrysosporium to antioxidative response and defence system exposed to Astrazone Red FBL. Catalase, glutathione reductase, glutathione s-transferase activities and level of glutathione decreased, depending on the period of growth in each exposure to low and high concentration group (20 and 50 ppm) compared with the control group.

Sensitizing capacity of Disperse Orange 1 and its potential metabolites from azo reduction and their cross-reactivity pattern.

BACKGROUND: Simultaneous contact allergies to Disperse Orange 1, 4-nitroaniline and p-aminodiphenylamine (PADPA), as well as to other disperse azo dyes and to p-phenylenediamine (PPD), have been reported. Cross-reactivity is one of the possible explanations for simultaneous reactions between PPD and disperse azo dyes. Some metabolites from the azo reduction of these disperse azo dyes could be sensitizers, as human skin bacteria produce azo reductases. OBJECTIVES: To investigate the sensitizing capacity of Disperse Orange 1, PADPA and 4-nitroaniline, and the cross-reactivity between these substances and Disperse Yellow 3, its potential metabolites from azo reduction (4-aminoacetanilide and 2-amino-p-cresol), and PPD. METHOD: The guinea-pig maximization test was used. RESULTS: It was found that both Disperse Orange 1 and PADPA are strong sensitizers and cross-react with each other. We were unable to sensitize guinea-pigs with 4-nitroaniline tested in equimolar concentrations to Disperse Orange 1. CONCLUSIONS: The results indicate that patients sensitized primarily to Disperse Orange 1 will also react to PADPA, which can be found mainly in hair dyes. PPD, 4-nitroaniline, 4-aminoacetanilide, 2-amino-p-cresol and Disperse Yellow 3 did not show any cross-reactivity with Disperse Orange 1 or PADPA.

Growth-inhibitory and chemosensitizing effects of the glutathione-S-transferase-π-activated nitric oxide donor PABA/NO in malignant gliomas.

Glutathione-S-transferases (GSTs) are upregulated in malignant gliomas and contribute to their chemoresistance. The nitric oxide (NO) donor PABA/NO (O(2) -{2,4-dinitro-5-[4-(N-methylamino)benzoyloxy]phenyl} 1-(N,N-dimethylamino)diazen-1-ium-1,2-diolate) generates NO upon selective enzymatic activation by GST-π-inducing selective biological effects in tumors. Tumor cell killing and chemosensitization were observed in a variety of tumors after exposure to GST-activated NO donor drugs. In our project, cytotoxic and chemosensitizing effects of PABA/NO in combination with carboplatin (CPT) and temozolomide (TMZ) were studied in human U87 glioma cells in vitro and in vivo. U87 glioma cells were exposed to PABA/NO alone or in combination with CPT or TMZ for 24 hr. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay after 24-hr incubation and 48 hr after drug removal. The antiproliferative effect of PABA/NO was assessed in an intracranial U87 glioma nude rat model comparing subcutaneous administration and intratumoral delivery by convection-enhanced delivery. PABA/NO monotherapy showed a strong dose-dependent growth-inhibitory effect in U87 glioma cells in vitro, and a strong synergistic effect was observed after concomitant treatment with TMZ, but not with CPT. Systemic and intratumoral PABA/NO administration significantly reduced cell proliferation, but this did not result in prolonged survival in nude rats with intracranial U87 gliomas. PABA/NO has potent antiproliferative effects, sensitizes U87 glioma cells to TMZ in vitro and shows some in vivo efficacy. Further studies are still required to consolidate the role of NO donor therapy in glioma treatment.

P2Y1 and P2Y13 purinergic receptors mediate Ca2+ signaling and proliferative responses in pulmonary artery vasa vasorum endothelial cells.

Extracellular ATP and ADP have been shown to exhibit potent angiogenic effects on pulmonary artery adventitial vasa vasorum endothelial cells (VVEC). However, the molecular signaling mechanisms of extracellular nucleotide-mediated angiogenesis remain not fully elucidated. Since elevation of intracellular Ca(2+) concentration ([Ca(2+)](i)) is required for cell proliferation and occurs in response to extracellular nucleotides, this study was undertaken to delineate the purinergic receptor subtypes involved in Ca(2+) signaling and extracellular nucleotide-mediated mitogenic responses in VVEC. Our data indicate that stimulation of VVEC with extracellular ATP resulted in the elevation of [Ca(2+)](i) via Ca(2+) influx through plasma membrane channels as well as Ca(2+) mobilization from intracellular stores. Moreover, extracellular ATP induced simultaneous Ca(2+) responses in both cytosolic and nuclear compartments. An increase in [Ca(2+)](i) was observed in response to a wide range of purinergic receptor agonists, including ATP, ADP, ATPγS, ADPßS, UTP, UDP, 2-methylthio-ATP (MeSATP), 2-methylthio-ADP (MeSADP), and BzATP, but not adenosine, AMP, diadenosine tetraphosphate, αßMeATP, and ßγMeATP. Using RT-PCR, we identified mRNA for the P2Y1, P2Y2, P2Y4, P2Y13, P2Y14, P2X2, P2X5, P2X7, A1, A2b, and A3 purinergic receptors in VVEC. Preincubation of VVEC with the P2Y1 selective antagonist MRS2179 and the P2Y13 selective antagonist MRS2211, as well as with pertussis toxin, attenuated at varying degrees agonist-induced intracellular Ca(2+) responses and activation of ERK1/2, Akt, and S6 ribosomal protein, indicating that P2Y1 and P2Y13 receptors play a major role in VVEC growth responses. Considering the broad physiological implications of purinergic signaling in the regulation of angiogenesis and vascular homeostasis, our findings suggest that P2Y1 and P2Y13 receptors may represent novel and specific targets for treatment of pathological vascular remodeling involving vasa vasorum expansion.

Antibacterial activity of nitric oxide-releasing carboxymethylcellulose against periodontal pathogens.

The prevalence of periodontal disease poses a significant global health burden. Treatments for these diseases, primarily focused on removal and eradication of dental plaque biofilms, are challenging due to limited access to periodontal pockets where these oral pathogens reside. Herein, we report on the development and characterization of nitric oxide (NO)-releasing carboxymethylcellulose (CMC) derivatives and evaluate their in vitro bactericidal efficacy against planktonic Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, two prominent periodontopathogens. Bactericidal exposure assays revealed that three of the synthesized NO-releasing polymers were capable of reducing bacterial viability of both species by 99.9% in 2 hr at concentrations of 4 mg ml-1 or lower, reflecting NO's potent and rapid bactericidal action. The NO-releasing CMCs elicited minimal toxicity to human gingival fibroblasts at their bactericidal concentrations following 24-hr exposure.

Stabilization of the nitric oxide (NO) prodrugs and anticancer leads, PABA/NO and Double JS-K, through incorporation into PEG-protected nanoparticles.

We report the stabilization of the nitric oxide (NO) prodrugs and anticancer lead compounds, PABA/NO (O(2)-{2,4-dinitro-5-[4-(N-methylamino)benzoyloxy]phenyl} 1-(N,N-dimethylamino)diazen-1-ium-1,2-diolate) and "Double JS-K" 1,5-bis-{1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diol-2-ato}-2,4-dinitrobenzene, through their incorporation into polymer-protected nanoparticles. The prodrugs were formulated in block copolymer-stabilized nanoparticles with sizes from 220 to 450 nm by a novel rapid precipitation process. The block copolymers, with polyethylene glycol (PEG) soluble blocks, provide a steric barrier against NO prodrug activation by glutathione. Too rapid activation and NO release has been a major barrier to effective administration of this class of compounds. The nanoparticle stabilized PABA/NO are protected from attack by glutathione as evidenced by a significant increase in time taken for 50% decomposition from 15 min (unformulated) to 5 h (formulated); in the case of Double JS-K, the 50% decomposition time was extended from 4.5 min (unformulated) to 40 min (formulated). The more hydrophobic PABA/NO produced more stable nanoparticles and correspondingly more extended release times in comparison with Double JS-K. The hydrophobic blocks of the polymer were either polystyrene or polylactide. Both blocks produced nanoparticles of approximately the same size and release kinetics. This combination of PEG-protected nanoparticles with sizes appropriate for cancer targeting by enhanced permeation and retention (EPR) and delayed release of NO may afford enhanced therapeutic benefit.

Delivery Strategies for Skin: Comparison of Nanoliter Jets, Needles and Topical Solutions.

Drug diffusion within the skin with a needle-free micro-jet injection (NFI) device was compared with two well-established delivery methods: topical application and solid needle injection. A permanent make-up (PMU) machine, normally used for dermal pigmentation, was utilized as a solid needle injection method. For NFIs a continuous wave (CW) laser diode was used to create a bubble inside a microfluidic device containing a light absorbing solution. Each method delivered two different solutions into ex vivo porcine skin. The first solution consisted of a red dye (direct red 81) and rhodamine B in water. The second solution was direct red 81 and rhodamine B in water and glycerol. We measured the diffusion depth, width and surface area of the solutions in all the injected skin samples. The NFI has a higher vertical dispersion velocity of 3 × 105µm/s compared to topical (0.1 µm/s) and needle injection (53 µm/s). The limitations and advantages of each method are discussed, and we conclude that the micro-jet injector represents a fast and minimally invasive injection method, while the solid needle injector causes notable tissue damage. In contrast, the topical method had the slowest diffusion rate but causes no visible damage to the skin.