In vitro to in vivo extrapolation to derive a metabolism factor for estimating the aggregate exposure to salicylic acid after dermal exposure of its esters.

As part of the safety assessment of salicylate esters in cosmetics, we developed a metabolism factor based on in vitro to in vivo extrapolation (IVIVE) to provide a better estimation of the aggregate internal exposure to the common metabolite, salicylic acid. Optimal incubation conditions using human liver S9 were identified before measuring salicylic acid formation from 31 substances. Four control substances, not defined as salicylic esters but which could be mistaken as such due to their nomenclature, did not form salicylic acid. For the remaining substances, higher in vitro intrinsic clearance (CLint, in vitro) values generally correlated with lower LogP values. A "High-Throughput Pharmacokinetic" (HTPK) model was used to extrapolate CLint, in vitro values to human in vivo clearance and half-lives. The latter were used to calculate the percentage of substance metabolised to salicylic acid in 24 h in vivo following human exposure to the ester, i.e. the "metabolism factor". The IVIVE model correctly reproduced the observed elimination rate of 3 substances using in silico or in vitro input parameters. For other substances, in silico only-based predictions generally resulted in lower metabolism factors than when in vitro values for plasma binding and liver S9 CLint, in vitro were used. Therefore, in vitro data input provides the more conservative metabolism factors compared to those derived using on in silico input. In conclusion, these results indicate that not all substances contribute equally (or at all) to the systemic exposure to salicylic acid. Therefore, we propose a realistic metabolism correction factor by which the potential contribution of salicylate esters to the aggregate consumer exposure to salicylic acid from cosmetic use can be estimated.

Evaluation of oral and perioral irritation and sensitization potential of a whitening gel and a whitening toothpaste containing potassium monopersulfate.

PURPOSE: Two clinical trials were conducted to investigate the oral and perioral irritation and sensitization potential of a tooth whitening leave-on-gel alone and in combination with a whitening toothpaste, each containing 1.0% of the active ingredient potassium monopersulfate (MPS). METHODS: Both clinical trials were Institutional Review Board (IRB) approved, double-blind, randomized, and parallel group designed studies. For the MPS leave-on gel study, 200 qualifying and consented subjects were randomly assigned to two groups: (1) 0.1% hydrogen peroxide (H2O2) gel pen (34 subjects); and (2) 0.1% H2O2 + 1.0% MPS gel pen (166 subjects). Subjects used the assigned products according to instructions provided and returned on Days 22 and 36 for oral and perioral tissue examination (pre-challenge). At the Day 36 visit, the subject applied the assigned gel on site (challenge) and received oral and perioral tissue examinations 1 and 24 hours following the application to detect any post-challenge tissue reactions. For the MPS toothpaste/MPS gel pen study, 200 qualifying and consented subjects were randomly assigned to three groups: (1) Placebo toothpaste + placebo gel pen (66 subjects); (2) 1.0% MPS toothpaste + 1.0% MPS gel pen (67 subjects); and (3) 1.0% MPS toothpaste + placebo gel pen (67 subjects). The study design and procedures were the same as those for the MPS gel pen study described above. RESULTS: For the MPS gel pen study, 192 subjects completed the study. None of the eight dropouts was related to the product use. The demographic data were comparable between the two groups. No evidence of tissue irritation and sensitization was detected in any subjects at any visit, and the findings were comparable between the groups. The detected and self-reported tissue issues were minimal and minor, and they were comparable between the two groups. For the MPS toothpaste/MPS gel pen study, 200 subjects were enrolled with 12 dropped from the study, resulting in an overall dropout rate of 6%. Of the 12 that did not complete the study, none were due to product-related use. The demographic data were comparable among the three groups. The detected and self-reported tissue issues were minimal and minor, and they were comparable among the three groups. CLINICAL SIGNIFICANCE: Potassium monopersulfate (MPS) at the active concentration of 1.0% in the tooth whitening leave-on-gel and the toothpaste plus the gel did not cause oral/perioral irritation nor sensitization.

Assessment of dermal absorption of aluminium from a representative antiperspirant formulation using a (26Al)Al microtracer approach: a follow-up study in humans.

A follow-up study was performed in 12 healthy women to evaluate systemic exposure to aluminium following topical application of a representative antiperspirant formulation under real-life use conditions (part A) and to assess the local fate of topically applied aluminium by taking additional tape strips and skin biopsies (Part B). A simple roll-on formulation, containing the maximal possible radioactive dose, was prepared with [26Al] aluminium-labeled chlorohydrate (ACH). The microtracer of [26Al] was used to distinguish aluminium from the natural background, using accelerator mass spectrometry. [26Al] aluminiumcitrate was administered intravenously to estimate the dermal fraction absorbed. Despite the 25-fold increase of the topical dose compared with the previous study, only 12 blood samples gave results above the lower limit of quantitation (0.118 fg/mL). The most reliable estimates of the dermal fraction absorbed are derived from noncompartmental analysis with the urine data. By using the intravenous dose to normalize the urinary excretion to 100% bioavailability, the best estimate of the fraction absorbed of [26Al] from a topical application of [26Al]-aluminium-labeled chlorohydrate in an antiperspirant formulation was 0.00052%. Part B of the study demonstrated that the majority of the aluminium in the formulation remained associated with the external layers of the skin without penetration through the skin.

The BlueScreen HC assay to predict the genotoxic potential of fragrance materials.

BlueScreen HC is a mammalian cell-based assay for measuring the genotoxicity and cytotoxicity of chemical compounds and mixtures. The BlueScreen HC assay has been utilized at the Research Institute for Fragrance Materials in a safety assessment program as a screening tool to prioritize fragrance materials for higher-tier testing, as supporting evidence when using a read-across approach, and as evidence to adjust the threshold of toxicological concern. Predictive values for the BlueScreen HC assay were evaluated based on the ability of the assay to predict the outcome of in vitro and in vivo mutagenicity and chromosomal damage genotoxicity assays. A set of 371 fragrance materials was assessed in the BlueScreen HC assay along with existing or newly generated in vitro and in vivo genotoxicity data. Based on a weight-of-evidence approach, the majority of materials in the data set were deemed negative and concluded not to have the potential to be genotoxic, while only a small proportion of materials were determined to show genotoxic effects in these assays. Analysis of the data set showed a combination of high positive agreement but low negative agreement between BlueScreen HC results, in vitro regulatory genotoxicity assays, and higher-tier test results. The BlueScreen HC assay did not generate any false negatives, thereby providing robustness when utilizing it as a high-throughput screening tool to evaluate the large inventory of fragrance materials. From the perspective of protecting public health, it is desirable to have no or minimal false negatives, as a false-negative result may incorrectly indicate the lack of a genotoxicity hazard. However, the assay did have a high percentage of false-positive results, resulting in poor positive predictivity of the in vitro genotoxicity test battery outcome. Overall, the assay generated 100% negative predictivity and 3.9% positive predictivity. In addition to the data set of 371 fragrance materials, 30 natural complex substances were evaluated for BlueScreen HC, Ames, and in vitro micronucleus assay, and a good correlation in all three assays was observed. Overall, while a positive result may have to be further investigated, these findings suggest that the BlueScreen HC assay can be a valuable screening tool to detect the genotoxic potential of fragrance materials and mixtures.

Hemoglobin, Body Mass Index, and Age as Risk Factors for Paclitaxel- and Oxaliplatin-Induced Peripheral Neuropathy.

Importance: Chemotherapy-induced peripheral neuropathy (CIPN) is a debilitating adverse effect of neurotoxic cancer treatments including taxanes and platinum agents. Limited knowledge exists of potential prechemotherapy factors associated with CIPN development. Objective: To identify the association of pretreatment blood-based and clinical factors with CIPN persistence in patients who received paclitaxel or oxaliplatin. Design, Setting, and Participants: This cohort study assessed pretreatment blood-based clinical factors and demographic characteristics of 333 patients treated with paclitaxel and oxaliplatin chemotherapy at urban multicenter cancer clinics and academic institutions in Australia between September 2015 and February 2020. Comprehensive neuropathy assessments were undertaken 3 to 12 months posttreatment. Posttreatment CIPN severity was compared with blood-based factors within 30 days prior to commencing chemotherapy. Data were analyzed between March and December 2020. Exposures: Paclitaxel or oxaliplatin chemotherapy. Main Outcomes and Measures: CIPN was measured using composite neurological grading scales, nerve conduction studies, and assessments of fine motor skills (grooved pegboard test), sensory function (grating orientation test and 2-point discrimination), and patient-reported outcomes. Independent samples t tests and Mann-Whitney U tests with post hoc Bonferroni correction were used to compare CIPN between patients according to blood-based factor normative ranges. Linear regression was used to identify blood-based and clinical associations with CIPN development. Results: The study included 333 participants (266 [79.9%] women; median [interquartile range] age, 58 [18] years) who were consecutively recruited and referred (228 treated with paclitaxel, 105 treated with oxaliplatin; 138 [41.4%] with breast cancer, 83 [24.9%] with colorectal cancer). Most participants had grade 1 CIPN or higher (238 [71.5%] participants). Participants with low hemoglobin pretreatment had worse CIPN posttreatment (median [IQR] composite neurological grading scale score, 5 [2-8] vs 4 [1-6]; P = .002; grooved pegboard mean [SD] time, 84.2 [28.7] vs 72.9 [21.1] seconds; P = .002; grating orientation task, 4.8 [2.8] vs 3.9 [1.8] mm; P = .03; 2-point discrimination, 45% vs 28%; P = .01), with no other impairments outside normative ranges associated with CIPN. In the multivariable model, several factors were associated with worse CIPN (F4,315 = 18.6; P < .001; r2 = .19) including for lower hemoglobin (ß = -0.47; 95% CI, -0.73 to -0.21; P < .001), higher body mass index (ß = 0.08; 95% CI, 0.02 to 0.12; P = .007), older age (ß = 0.08; 95% CI, 0.06 to 0.11; P < .001), and female sex (ß = -1.08; 95% CI, -1.76 to -0.16; P = .01). Conclusions and Relevance: The results of this cohort study suggest that participants with low pretreatment hemoglobin, higher body mass index, older age, and female sex were more likely to develop paclitaxel- or oxaliplatin-induced CIPN posttreatment. Future research should investigate prospectively whether these risk factors are associated with a higher incidence of CIPN development.

Skin sensitization in silico protocol.

The assessment of skin sensitization has evolved over the past few years to include in vitro assessments of key events along the adverse outcome pathway and opportunistically capitalize on the strengths of in silico methods to support a weight of evidence assessment without conducting a test in animals. While in silico methods vary greatly in their purpose and format; there is a need to standardize the underlying principles on which such models are developed and to make transparent the implications for the uncertainty in the overall assessment. In this contribution, the relationship between skin sensitization relevant effects, mechanisms, and endpoints are built into a hazard assessment framework. Based on the relevance of the mechanisms and effects as well as the strengths and limitations of the experimental systems used to identify them, rules and principles are defined for deriving skin sensitization in silico assessments. Further, the assignments of reliability and confidence scores that reflect the overall strength of the assessment are discussed. This skin sensitization protocol supports the implementation and acceptance of in silico approaches for the prediction of skin sensitization.

Participation of xCT in melanoma cell proliferation in vitro and tumorigenesis in vivo.

Our research group demonstrated that riluzole, an inhibitor of glutamatergic signaling reduced melanoma cell proliferation in vitro and tumor progression in vivo. The underlying mechanisms of riluzole are largely unknown. Microarray analyses on two human melanoma cell lines revealed that riluzole stimulates expression of the cystine-glutamate amino acid antiporter, xCT (SLC7A11). Western immunoblot analysis from cultured human melanoma or normal melanocytic cells showed that xCT was significantly overexpressed in most melanomas, but not normal cells. Studies using human tumor biopsy samples demonstrated that overexpression of xCT was correlated with cancer stage and progression. To further investigate if xCT is involved in melanoma cell growth, we derived several stable clones through transfection of exogenous xCT to melanoma cells that originally showed very low expression of xCT. The elevated xCT expression promoted cell proliferation in vitro and inversely, these melanoma clones showed a dose-dependent decrease in cell proliferation in response to riluzole treatment. Xenograft studies showed that these clones formed very aggressive tumors at a higher rate compared to vector controls. Conversely, treatment of xenograft-bearing animals with riluzole down-regulated xCT expression suggesting that xCT is a molecular target of riluzole. Furthermore, protein lysates from tumor biopsies of patients that participated in a riluzole monotherapy phase II clinical trial showed a reduction in xCT levels in post-treatment specimens from patients with stable disease. Taken together, our results show that xCT may be utilized as a marker to monitor patients undergoing riluzole-based chemotherapies.

Pointing to One's Moving Hand: Putative Internal Models Do Not Contribute to Proprioceptive Acuity.

We can easily and without sight bring our fingertip to our nose, or swat a mosquito on our arm. These actions rely on proprioception, also known as kinesthesia, which classically has been attributed to processing of sensory inflow by the CNS. However, internal model theories of sensorimotor neuroscience propose that proprioceptive localization also involves a contribution from estimates of limb kinematics derived from motor commands. We tested this prediction in 19 subjects who moved the right index finger tip to touch the moving left index finger tip under three conditions: (1) vision allowed, active movement of the left hand (2) vision blocked, active movement of the left hand, and (3) vision blocked, passive movement of the left hand imposed by the experimenter. The target left index finger tip was moved in a wide range of directions by unrestricted movements of the arm. Mean errors in apposition of the right to the left index finger tips were small, averaging <2 cm between sensors fixed to the finger nails. Note that the average distance between the sensors was ~1.7 cm when the fingertips were brought together in "perfect" apposition under visual guidance. The 3D mean distance and variable distance errors were marginally lower by some 2 mm with eyes open compared to the eyes closed active condition. However, mean distance and variable distance errors did not differ between the active and passive conditions with eyes closed. Thus, proprioceptive localization of one's moving hand is very accurate, essentially as accurate as when vision is allowed. More importantly, our results demonstrate that hypothesized internal model derived estimates of arm kinematics do not contribute to localization accuracy beyond that provided by sensory signals, casting doubt on their existence.

Activation of Grm1 expression by mutated BRaf (V600E) in vitro and in vivo.

Our laboratory previously showed that ectopic expression of Grm1 is sufficient to induce spontaneous melanoma formation with 100% penetrance in transgenic mouse model, TG-3, which harbors wild-type BRaf. Studies identified Grm1 expression in human melanoma cell lines and primary to secondary metastatic melanoma biopsies having wild-type or mutated BRaf, but not in normal melanocytes or benign nevi. Grm1 expression was detected in tissues from mice genetically engineered with inducible melanocyte-specific BRafV600E. Additionally, stable clones derived from introduction of exogenous BRafV600E in mouse melanocytes also showed Grm1 expression, which was not detected in the parental or empty vector-derived cells, suggesting that expression of BRafV600E could activate Grm1 expression. Despite aberrant Grm1 expression in the inducible, melanocyte-specific BRafV600E mice, no tumors formed. However, in older mice, the melanocytes underwent senescence, as demonstrated previously by others. It was proposed that upregulated p15 and TGFß contributed to the senescence phenotype. In contrast, in older TG-3 mice the levels of p15 and TGFß remained the same or lower. Taken together, these results suggest the temporal regulation on the expression of "oncogenes" such as Grm1 or BRafV600E is critical in the future fate of the cells. If BRafV600E is turned on first, Grm1 expression can be induced, but this is not sufficient to result in development of melanoma; the cells undergo senescence. In contrast, if ectopic expression of Grm1 is turned on first, then regardless of wild-type or mutated BRaf in the melanocytes melanoma development is the consequence.

Differential Gene Regulation and Tumor-Inhibitory Activities of Alpha-, Delta-, and Gamma-Tocopherols in Estrogen-Mediated Mammary Carcinogenesis.

Despite experimental evidence elucidating the antitumor activities of tocopherols, clinical trials with α-tocopherol (α-T) have failed to demonstrate its beneficial effects in cancer prevention. This study compared the chemopreventive efficacy of individual tocopherols (α-, δ-, and γ-T) and a γ-T-rich tocopherol mixture (γ-TmT) in the August-Copenhagen Irish (ACI) rat model of estrogen-mediated mammary cancer. Female ACI rats receiving 17ß-estradiol (E2) implants were administered with 0.2% α-T, δ-T, γ-T, or γ-TmT for 30 weeks. Although α-T had no significant effects on mammary tumor growth in ACI rats, δ-T, γ-T, and γ-TmT reduced mammary tumor volume by 51% (P < 0.05), 60% (P < 0.01), and 59% (P < 0.01), respectively. Immunohistochemical analysis revealed that δ-T, γ-T, and γ-TmT reduced levels of the cell proliferation marker, proliferating cell nuclear antigen, in the rat mammary tumors. To gain further insight into the biological functions of different forms of tocopherols, RNA-seq analysis of the tumors was performed. Treatment with γ-T induced robust gene expression changes in the mammary tumors of ACI rats. Ingenuity Pathway Analysis identified "Cancer" as a top disease pathway and "Tumor growth" and "Metastasis" as the top signaling pathways modulated by γ-T. Although the results need further functional validation, this study presents an unbiased attempt to understand the differences between biological activities of individual forms of tocopherols at the whole transcriptome level. In conclusion, δ-T and γ-T have superior cancer preventive properties compared to α-T in the prevention of estrogen-mediated mammary carcinogenesis. Cancer Prev Res; 10(12); 694-703. ©2017 AACR.

Amino Acid Block Copolymers with Broad Antimicrobial Activity and Barrier Properties.

Antimicrobial properties of a long-chain, synthetic, cationic, and hydrophobic amino acid block copolymer are reported. In 5 and 60 min time-kill assays, solutions of K100 L40 block copolymers (poly(l-lysine·hydrochloride)100 -b-poly(l-leucine)40 ) at concentrations of 10-100 µg mL-1 show multi-log reductions in colony forming units of Gram-positive and Gram-negative bacteria, as well as yeast, including multidrug-resistant strains. Driven by association of hydrophobic segments, K100 L40 copolymers form viscous solutions and self-supporting hydrogels in water at concentrations of 1 and 2 wt%, respectively. These K100 L40 preparations provide an effective barrier to microbial contamination of wounds, as measured by multi-log decreases of tissue-associated bacteria with deliberate inoculation of porcine skin explants, porcine open wounds, and rodent closed wounds with foreign body. Based on these findings, amino acid copolymers with the features of K100 L40 can combine potent, direct antimicrobial activity and barrier properties in one biopolymer for a new approach to prevention of wound infections.

Capnography versus standard monitoring for emergency department procedural sedation and analgesia.

BACKGROUND: Procedural sedation and analgesia (PSA) is used frequently in the emergency department (ED) to facilitate painful procedures and interventions. Capnography, a monitoring modality widely used in operating room and endoscopy suite settings, is being used more frequently in the ED setting with the goal of reducing cardiopulmonary adverse events. As opposed to settings outside the ED, there is currently no consensus on whether the addition of capnography to standard monitoring modalities reduces adverse events in the ED setting. OBJECTIVES: To assess whether capnography in addition to standard monitoring (pulse oximetry, blood pressure and cardiac monitoring) is more effective than standard monitoring alone to prevent cardiorespiratory adverse events (e.g. oxygen desaturation, hypotension, emesis, and pulmonary aspiration) in ED patients undergoing PSA. SEARCH METHODS: We searched the Cochrane Central Register of Controlled Trials (2016, Issue 8), and MEDLINE, Embase, and CINAHL to 9 August 2016 for randomized controlled trials (RCTs) and quasi-randomized trials of ED patients requiring PSA with no language restrictions. We searched meta-registries (www.controlled-trials.com, www.clinicalstudyresults.org, and clinicaltrials.gov) for ongoing trials (February 2016). We contacted the primary authors of included studies as well as scientific advisors of capnography device manufacturers to identify unpublished studies (February 2016). We handsearched conference abstracts of four organizations from 2010 to 2015. SELECTION CRITERIA: We included any RCT or quasi-randomized trial comparing capnography and standard monitoring to standard monitoring alone for ED patients requiring PSA. DATA COLLECTION AND ANALYSIS: Two authors independently performed study selection, data extraction, and assessment of methodological quality for the 'Risk of bias' tables. An independent researcher extracted data for any included studies that our authors were involved in. We contacted authors of included studies for incomplete data when applicable. We used Review Manager 5 to combine data and calculate risk ratios (RR) and 95% confidence intervals (CI) using both random-effects and fixed-effect models. MAIN RESULTS: We identified three trials (κ = 1.00) involving 1272 participants. Comparing the capnography group to the standard monitoring group, there were no differences in the rates of oxygen desaturation (RR 0.89, 95% CI 0.48 to 1.63; n = 1272, 3 trials; moderate quality evidence) and hypotension (RR 2.36, 95% CI 0.98 to 5.69; n = 986, 1 trial; moderate quality evidence). There was only one episode of emesis recorded without significant difference between the groups (RR 3.10, 95% CI 0.13 to 75.88, n = 986, 1 trial; moderate quality evidence). The quality of evidence for the primary outcomes was moderate with downgrades primarily due to heterogeneity and reporting bias.There were no differences in the rate of airway interventions performed (RR 1.26, 95% CI 0.94 to 1.69; n = 1272, 3 trials; moderate quality evidence). In the subgroup analysis, we found a higher rate of airway interventions for adults in the capnography group (RR 1.44, 95% CI 1.16 to 1.79; n = 1118, 2 trials; moderate quality evidence) with a number needed to treat for an additional harmful outcome of 12. Although statistical heterogeneity was reduced, there was moderate quality of evidence due to outcome definition heterogeneity and limited reporting bias. None of the studies reported recovery time. AUTHORS' CONCLUSIONS: There is a lack of convincing evidence that the addition of capnography to standard monitoring in ED PSA reduces the rate of clinically significant adverse events. Evidence was deemed to be of moderate quality due to population and outcome definition heterogeneity and limited reporting bias. Our review was limited by the small number of clinical trials in this setting.

Metabotropic glutamate receptors in cancer.

Metabotropic glutamate receptors (mGluRs) are widely known for their roles in synaptic signaling. However, accumulating evidence suggests roles of mGluRs in human malignancies in addition to synaptic transmission. Somatic cell homeostasis presents intriguing possibilities of mGluRs and glutamate signaling as novel targets for human cancers. More recently, aberrant glutamate signaling has been shown to participate in the transformation and maintenance of various cancer types, including glioma, melanoma skin cancer, breast cancer, and prostate cancer, indicating that genes encoding mGluRs, GRMs, can function as oncogenes. Here, we provide a review on the interactions of mGluRs and their ligand, glutamate, in processes that promote the growth of tumors of neuronal and non-neuronal origins. Further, we discuss the evolution of riluzole, a glutamate release inhibitor approved for amyotrophic lateral sclerosis (ALS), but now fashioned as an mGluR1 inhibitor for melanoma therapy and as a radio-sensitizer for tumors that have metastasized to the brain. With the success of riluzole, it is not far-fetched to believe that other drugs that may act directly or indirectly on other mGluRs can be beneficial for multiple applications. This article is part of the Special Issue entitled 'Metabotropic Glutamate Receptors, 5 years on'.

The current management of brain metastasis in melanoma: a focus on riluzole.

Brain metastasis is a common endpoint in human malignant melanoma, and the prognosis for patients remains poor despite advancements in therapy. Current treatment for melanoma metastatic to the brain is grouped into those providing symptomatic relief such as corticosteroids and antiepileptic agents, to those that are disease modifying. Related to the latter group, recent studies have demonstrated that aberrant glutamate signaling plays a role in the transformation and maintenance of various cancer types, including melanoma. Glutamate secretion from these and surrounding cells have been found to stimulate regulatory pathways that control tumor growth, proliferation and survival in vitro and in vivo. The antiglutamatergic actions of an inhibitor of glutamate release, riluzole, have been detected by its ability to clear glutamate from the synapse, and it has been shown to inhibit glutamate release rather than directly inhibiting glutamate receptors. Preclinical studies have demonstrated the ability of riluzole to act as a radiosensitizing agent in melanoma. The effect of riluzole on downstream glutamatergic signaling has pointed to cross talk between the metabotropic G-protein-coupled glutamate receptors implicated in a subset of human melanomas with other signaling pathways, including apoptotic, angiogenic, ROS and cell invasion mechanisms, thus establishing its potential to be further explored in combination therapy regimens for both primary human melanoma and melanoma metastatic to the brain.

Riluzole is a radio-sensitizing agent in an in vivo model of brain metastasis derived from GRM1 expressing human melanoma cells.

Approximately 50% of patients having metastatic melanoma develop brain metastases during the course of their illness. Evidence exists that melanoma cells have increased aptitude for the repair of sublethal DNA damage caused by ionizing radiation therapy. To address the radio-resistance of melanoma, many groups adopted radiotherapy schedules that deliver larger daily fractions of radiation, but due to the risk of neurotoxicity, these large fractions cannot be delivered to the whole brain for patients with brain metastases. Here, we used orthotopic implanted GRM1 expressing human melanoma cell xenografts in mice, to demonstrate that animals receiving concurrent glutamate signaling blockade (riluzole) and radiation led to a decrease in intracranial tumor growth compared to either modality alone. These preclinical results suggest riluzole may cause radio-sensitization that offers enhanced efficacy for a subset of human melanoma patients undergoing radiotherapy for brain metastasis.

Tocopherols inhibit oxidative and nitrosative stress in estrogen-induced early mammary hyperplasia in ACI rats.

Oxidative stress is known to play a key role in estrogen-induced breast cancer. This study assessed the chemopreventive activity of the naturally occurring γ-tocopherol-rich mixture of tocopherols (γ-TmT) in early stages of estrogen-induced mammary hyperplasia in ACI rats. ACI rats provide an established model of rodent mammary carcinogenesis due to their high sensitivity to estrogen. Female rats were implanted with 9 mg of 17ß-estradiol (E2) in silastic tubings and fed with control or 0.3% γ-TmT diet for 1, 3, 7, and 14 d. γ-TmT increased the levels of tocopherols and their metabolites in the serum and mammary glands of the rats. Histological analysis revealed mammary hyperplasia in the E2 treated rats fed with control or γ-TmT diet. γ-TmT decreased the levels of E2-induced nitrosative and oxidative stress markers, nitrotyrosine, and 8-oxo-dG, respectively, in the hyperplastic mammary tissues. 8-Isoprostane, a marker of oxidative stress in the serum, was also reduced by γ-TmT. Noticeably, γ-TmT stimulated Nrf2-dependent antioxidant response in the mammary glands of E2 treated rats, evident from the induced mRNA levels of Nrf2 and its downstream antioxidant enzymes, superoxide dismutase, catalase, and glutathione peroxidase. Therefore, inhibition of nitrosative/oxidative stress through induction of antioxidant response is the primary effect of γ-TmT in early stages of E2-induced mammary hyperplasia. Due to its cytoprotective activity, γ-TmT could be a potential natural agent for the chemoprevention of estrogen-induced breast cancer.

Variation of formal hydrogen-bonding networks within electronically delocalized π-conjugated oligopeptide nanostructures.

This photophysical study characterizes the generality of intermolecular electronic interactions present within nanomaterials derived from self-assembling oligopeptides with embedded π-conjugated oligophenylenevinylene (OPV) subunits stilbene and distyrylbenzene that in principle present two distinct ß-sheet motifs. Two different synthetic approaches led to oligopeptides that upon self-assembly are expected to self-assemble into multimeric aggregates stabilized by ß-sheet-like secondary structures. The target molecules express either two C-termini linked to the central OPV core (symmetric peptides) or the more common N-termini to C-termini polarity typical of natural oligopeptides (nonsymmetric peptides). Both peptide secondary structures were shown to form extended 1-D peptide aggregates with intimate intermolecular π-electron interactions. Differences in length of the π-conjugated OPV segments resulted in differing extents of intermolecular interactions and the resulting photophysics. The peptides containing the shorter stilbene (OPV2) units showed little ground state interactions and resulted in excimeric emission, while the longer distyrylbenzene (OPV3) peptides had different ground state interactions between adjacent π-conjugated subunits resulting in either perturbed electronic properties arising from exciton coupling or excimer-like excited states. Molecular dynamics simulations of nascent aggregate formation predict peptide dimerization to be a spontaneous process, possessing thermodynamic driving potentials in the range 2-6 kcal/mol for the four molecules considered. Antiparallel stacking of the peptides containing an OPV3 subunit is thermodynamically favored over the parallel orientation, whereas both arrangements are equally favored for the peptides containing an OPV2 subunit. This study validates the generality of peptide-π-peptide self-assembly to provide electronically delocalized supramolecular structures and suggests flexibility in peptide sequence design as a way to tune the material properties of π-conjugated supramolecular polymers.

Supramolecular polymorphism: tunable electronic interactions within π-conjugated peptide nanostructures dictated by primary amino acid sequence.

We present a systematic study of the photophysical properties of one-dimensional electronically delocalized nanostructures assembled from π-conjugated subunits embedded within oligopeptide backbones. The nature of the excited states within these nanostructures is studied as a function of primary amino acid sequence utilizing steady-state and time-resolved spectroscopies, and their atomistic structure is probed by molecular simulation. Variations introduced into the amino acid side chains at specific residue locations along the molecular peptide backbone lead to pronounced changes in the observed photophysical behavior of the fibrillar structures (spanning H-like excitonic coupling and disordered excimeric coupling) that arise from subtle changes in the π-stacking within them. These results indicate that residue modification-in terms of relative size, solvation properties, and with respect to the distance from the central π-electron core-enables the ability to tune chromophore packing and the resulting photophysics of supramolecular assemblies of π-conjugated bioelectronic materials in a rational and systematic manner.

Disruption of GRM1-mediated signalling using riluzole results in DNA damage in melanoma cells.

Gain of function of the neuronal receptor, metabotropic glutamate receptor 1 (Grm1), was sufficient to induce melanocytic transformation in vitro and spontaneous melanoma development in vivo when ectopically expressed in melanocytes. The human form of this receptor, GRM1, has been shown to be ectopically expressed in a subset of human melanomas but not benign nevi or normal melanocytes, suggesting that misregulation of GRM1 is involved in the pathogenesis of certain human melanomas. Sustained stimulation of Grm1 by the ligand, glutamate, is required for the maintenance of transformed phenotypes in vitro and tumorigenicity in vivo. In this study, we investigate the mechanism of an inhibitor of glutamate release, riluzole, on human melanoma cells that express metabotropic glutamate receptor 1 (GRM1). Various in vitro assays conducted show that inhibition of glutamate release in several human melanoma cell lines resulted in an increase of oxidative stress and DNA damage response markers.