A Step Forward for the Treatment of Localized Prostate Cancer Using Gold Nanoparticles Combined with Laser Irradiation.

Prostate cancer (PCA) is the second most common cancer diagnosis in men and the fifth leading cause of death worldwide. The conventional treatments available are beneficial to only a few patients and, in those, some present adverse side effects that eventually affect the quality of life of most patients. Thus, there is an urgent need for effective, less invasive and targeted specific treatments for PCA. Photothermal therapy (PTT) is a minimally invasive therapy that provides a localized effect for tumour cell ablation by activating photothermal agents (PTA) that mediate the conversion of the light beam's energy into heat at the site. As tumours are unable to easily dissipate heat, they become more susceptible to temperature increases. In the PTT field, gold nanoparticles (AuNPs) have been attracting interest as PTA. The aim of this study was to formulate AuNPs capable of remaining retained in the tumour and subsequently generating heat at the tumour site. AuNPs were synthesized and characterized in terms of size, polydispersity index (PdI), zeta potential (ZP), morphology and the surface plasmon resonance (SPR). The safety of AuNPs and their efficacy were assessed using in vitro models. A preliminary in vivo safety assessment of AuNPs with a mean size lower than 200 nm was confirmed. The morphology was spherical-like and the SPR band showed good absorbance at the laser wavelength. Without laser, AuNPs proved to be safe both in vitro (>70% viability) and in vivo. In addition, with laser irradiation, they proved to be relatively effective in PCA cells. Overall, the formulation appears to be promising for use in PTT.

Emergence and rapid dissemination of highly resistant NDM-14-producing Klebsiella pneumoniae ST147, France, 2022.

BackgroundSince 2021, an emergence of New Delhi metallo-ß-lactamase (NDM)-14-producing Klebsiella pneumoniae has been identified in France. This variant with increased carbapenemase activity was not previously detected in Enterobacterales.AimWe investigated the rapid dissemination of NDM-14 producers among patients in hospitals in France.MethodsAll NDM-14-producing non-duplicate clinical isolates identified in France until June 2022 (n = 37) were analysed by whole genome sequencing. The phylogeny of NDM-14-producers among all K. pneumoniae sequence type (ST) 147 reported in France since 2014 (n = 431) was performed. Antimicrobial susceptibility testing, conjugation experiments, clonal relationship and molecular clock analysis were performed.ResultsThe 37 NDM-14 producers recovered in France until 2022 belonged to K. pneumoniae ST147. The dissemination of NDM-14-producing K. pneumoniae was linked to a single clone, likely imported from Morocco and responsible for several outbreaks in France. The gene bla NDM-14 was harboured on a 54 kilobase non-conjugative IncFIB plasmid that shared high homology with a known bla NDM-1-carrying plasmid. Using Bayesian analysis, we estimated that the NDM-14-producing K. pneumoniae ST147 clone appeared in 2020. The evolutionary rate of this clone was estimated to 5.61 single nucleotide polymorphisms per genome per year. The NDM-14 producers were highly resistant to all antimicrobials tested except to colistin, cefiderocol (minimum inhibitory concentration 2 mg/L) and the combination of aztreonam/avibactam.ConclusionHighly resistant NDM-14 producing K. pneumoniae can rapidly spread in healthcare settings. Surveillance and thorough investigations of hospital outbreaks are critical to evaluate and limit the dissemination of this clone.

A Step Forward in Breast Cancer Research: From a Natural-Like Experimental Model to a Preliminary Photothermal Approach.

Breast cancer is one of the most frequently diagnosed malignancies and common causes of cancer death in women. Recent studies suggest that environmental exposures to certain chemicals, such as 7,12-Dimethylbenzanthracene (DMBA), a chemical present in tobacco, may increase the risk of developing breast cancer later in life. The first-line treatments for breast cancer (surgery, chemotherapy or a combination of both) are generally invasive and frequently associated with severe side effects and high comorbidity. Consequently, novel approaches are strongly required to find more natural-like experimental models that better reflect the tumors' etiology, physiopathology and response to treatments, as well as to find more targeted, efficient and minimally invasive treatments. This study proposes the development and an in deep biological characterization of an experimental model using DMBA-tumor-induction in Sprague-Dawley female rats. Moreover, a photothermal therapy approach using a near-infrared laser coupled with gold nanoparticles was preliminarily assessed. The gold nanoparticles were functionalized with Epidermal Growth Factor, and their physicochemical properties and in vitro effects were characterized. DMBA proved to be a very good and selective inductor of breast cancer, with 100% incidence and inducing an average of 4.7 tumors per animal. Epigenetic analysis showed that tumors classified with worst prognosis were hypomethylated. The tumor-induced rats were then subjected to a preliminary treatment using functionalized gold nanoparticles and its activation by laser (650-900 nm). The treatment outcomes presented very promising alterations in terms of tumor histology, confirming the presence of necrosis in most of the cases. Although this study revealed encouraging results as a breast cancer therapy, it is important to define tumor eligibility and specific efficiency criteria to further assess its application in breast cancer treatment on other species.

A selective CD28 antagonist and rapamycin synergise to protect against spontaneous autoimmune diabetes in NOD mice.

AIMS/HYPOTHESIS: The CD28/B7 interaction is critical for both effector T cell activation and forkhead box P3 (FOXP3)+ regulatory T cell (Treg) generation and homeostasis, which complicates the therapeutic use of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4)-immunoglobulin fusion protein (CTLA-4Ig) in autoimmunity. Here, we evaluated the impact of a simultaneous and selective blockade of the CD28 and mammalian target of rapamycin (mTOR) pathways in the NOD mouse model of type 1 diabetes. METHODS: NOD mice were treated with PEGylated anti-CD28 Fab' antibody fragments (PV1-polyethylene glycol [PEG], 10 mg/kg i.p., twice weekly), rapamycin (1 mg/kg i.p., twice weekly) or a combination of both drugs. Diabetes incidence, pancreatic islet infiltration and autoreactive T cell responses were analysed. RESULTS: We report that 4 week administration of PV1-PEG combined with rapamycin effectively controlled the progression of autoimmune diabetes in NOD mice at 10 weeks of age by reducing T cell activation and migration into the pancreas. Treatment with rapamycin alone was without effect, as was PV1-PEG monotherapy initiated at 4, 6 or 10 weeks of age. Prolonged PV1-PEG administration (for 10 weeks) accelerated diabetes development associated with impaired peripheral Treg homeostasis. This effect was not observed with the combined treatment. CONCLUSIONS/INTERPRETATION: CD28 antagonist and rapamycin treatment act in a complementary manner to limit T cell activation and infiltration of pancreatic islets and diabetes development. These data provide new perspectives for the treatment of autoimmune diabetes and support the therapeutic potential of protocols combining antagonists of CD28 (presently in clinical development) and the mTOR pathway.

Oral histone deacetylase inhibitor synergises with T cell targeted immunotherapy to preserve beta cell metabolic function and induce stable remission of new-onset autoimmune diabetes in NOD mice.

AIM/HYPOTHESIS: Combination therapy targeting the major actors involved in the immune-mediated destruction of pancreatic beta cells appears to be an indispensable approach to treat type 1 diabetes effectively. We hypothesised that the combination of an orally active pan-histone deacetylase inhibitor (HDACi: givinostat) with subtherapeutic doses of CD3 antibodies may provide ideal synergy to treat ongoing autoimmunity. METHODS: NOD mice transgenic for the human CD3ε (also known as CD3E) chain (NOD-huCD3ε) were treated for recent-onset diabetes with oral givinostat, subtherapeutic doses of humanised CD3 antibodies (otelixizumab, 50 µg/day, 5 days, i.v.) or a combination of both drugs. Disease remission, metabolic profiles and autoreactive T cell responses were analysed in treated mice. RESULTS: We demonstrated that givinostat synergised with otelixizumab to induce durable remission of diabetes in 80% of recently diabetic NOD-huCD3ε mice. Remission was obtained in only 47% of mice treated with otelixizumab alone. Oral givinostat monotherapy did not reverse established diabetes but reduced the in situ production of inflammatory cytokines (IL-1ß, IL-6, TNF-α). Importantly, the otelixizumab + givinostat combination strongly improved the metabolic status of NOD-huCD3ε mice; the mice recovered the capacity to appropriately produce insulin, control hyperglycaemia and sustain glucose tolerance. Finally, diabetes remission induced by the combination therapy was associated with a significant reduction of insulitis and autoantigen-specific CD8+ T cell responses. CONCLUSIONS/INTERPRETATION: HDACi and low-dose CD3 antibodies synergised to abrogate in situ inflammation and thereby improved pancreatic beta cell survival and metabolic function leading to long-lasting diabetes remission. These results support the therapeutic potential of protocols combining these two drugs, both in clinical development, to restore self-tolerance and insulin independence in type 1 diabetes.

Combined effects of oregano essential oil and salt on the growth of Escherichia coli in salad dressing.

There is a broad research interest in the search for alternatives to chemical additives for use as natural food preservatives. Although many natural compounds have biological in vitro properties evidenced, in situ studies are still scarce. This study evaluated the effect of oregano essential oil (OEO) and salt (NaCl) concentrations against Escherichia coli (ATCC 8739), in salad dressing, using the response surface methodology. The experiment included a 22 central composite rotatable design (CCRD) in a total of 11 formulations of salad dressings. Oregano essential oil was characterized by gas chromatography and salad dressings by ash, lipids, proteins and moisture. OEO was composed mainly by carvacrol (65.1%) and p-cymene (12.0%). Salad dressings showed similar chemical profiles. A mathematical model for the prediction of the antibacterial activity in salad dressing was obtained. The results revealed that the interaction between OEO and salt showed effect on the bacterial count. However, the effect of salt was negative suggesting that the highest NaCl concentrations decreases the bacterial count. Therefore, within the parameters studied, the use of OEO to control E. coli in salad dressing can be considered promising and allows reduction in the levels of salt to be incorporated in food.

Revisiting autoimmune gastritis in children and adolescents with type 1 diabetes.

OBJECTIVE: Assess the frequency of anti-H+ /K+ adenosine triphosphatase (ATPase) autoantibodies (AAB) and symptoms of autoimmune gastritis in children and adolescents with type 1 diabetes (T1D). RESEARCH DESIGN AND METHODS: Anti-H+ /K+ ATPase AAB were measured in 402 children and adolescents (210 boys and 192 girls, 11.1 ± 4.5 years) treated for T1D (screened positive for ß-cell AAB), along with search of symptoms of anemia (hemoglobin, serum iron, and ferritin levels) and gastric pain. The AAB specific for thyroperoxydase, thyroglobulin, and transglutaminase were also measured. RESULTS: Anti-H+ /K+ ATPase AAB were present in 6.5% of children. Their frequency increased with age: 4% at 10 years, 10% at 15 years, and 20% at 20 years. Iron deficiency (45% vs 3.8%), iron deficiency anemia (36% vs 3.8%), antithyroid AAB (24% vs 9.7%), and family history of Graves' disease (25% vs 5.6%) were more frequent in patients with anti-H+ /K+ ATPase AAB. Two patients, a 13-year-old girl and a 11-year-old boy, experienced symptoms (iron deficiency anemia and epigastric pain) which led to diagnosis of autoimmune gastritis confirmed upon fibroscopy. Both showed high levels of anti-H+ /K+ ATPase AAB and atrophic gastritis. CONCLUSIONS: Autoimmune gastritis presents an age-dependent frequency in children and adolescents with T1D but is rarely symptomatic. Screening for anti-H+ /K+ ATPase AAB should thus target patients with iron deficiency, anemia, epigastralgia, autoimmune thyroiditis, or age over 15 years.

Measurement properties of the incremental step test for people with chronic obstructive pulmonary disease: a cross-sectional study.

OBJECTIVES: The new incremental step test (IST) is a field test that was developed for people with chronic obstructive pulmonary disease (COPD), based on the characteristics of the incremental shuttle walk test (ISWT); however, its measurement properties still need to be determined. We aimed, first, to assess the construct validity (through the comparison with the ISWT), within-day reliability and measurement error of the IST in people with COPD; and, second, to identify whether the participants have a learning effect in the IST. DESIGN: Cross-sectional study, conducted according to COnsensus-based Standards for the selection of health status Measurement INstruments guidelines. SETTING: A family health unit in Portugal, April 2022 to June 2023. PARTICIPANTS AND ANALYSIS: 63 participants (67.5±10.5 years) attended two sessions to perform two IST and two ISWT, separately. Spearman's correlations were used to compare the best performances between the IST and the ISWT. Intraclass correlation coefficient (ICC2,1) was used for reliability, and the SE of measurement (SEM), minimal detectable change at 95% CI (MDC95) and Bland and Altman 95% limits of agreement (LoA) were used for measurement error. The learning effect was explored with the Wilcoxon signed-rank test. RESULTS: The IST was significant and strongly correlated with the ISWT (0.72<ρ<0.74, p<0.001), presented an ICC2,1 of 0.95 (95% CI 0.92 to 0.97), SEM=11.7 (18.9%), MDC95=32.4 (52.2%) and the LoA were -33.61 to 31.48 for the number of steps. No difference was observed between the number of steps of the two attempts of the IST (p>0.05). CONCLUSIONS: The IST can be suggested as a valid and reliable test to assess exercise capacity in people with COPD, with no learning effect when two IST are performed on the same day. The measurement error of the IST is considered indeterminate. TRIAL REGISTRATION NUMBER: NCT04715659.

Rational approach to design gold nanoparticles for photothermal therapy: the effect of gold salt on physicochemical, optical and biological properties.

Among the unique characteristics associated to gold nanoparticles (AuNPs) in biomedicine, their ability to convert light energy into heat opens ventures for improved cancer therapeutic options, such as photothermal therapy (PTT). PTT relies on the local hyperthermia of tumor cells upon irradiation with light beams, and the association of AuNPs with radiation within the near infrared (NIR) range constitutes an advantageous strategy to potentially improve PTT efficacy. Herein, it was explored the effect of the gold salt on the AuNPs' physicochemical and optical properties. Mostly spherical-like negatively charged AuNPs with variable sizes and absorbance spectra were obtained. In addition, photothermal features were assessed using in vitro phantom models. The best formulation showed the ability to increase their temperature in aqueous solution up to 19 °C when irradiated with a NIR laser for 20 min. Moreover, scanning transmission electron microscopy confirmed the rearrangement of the gold atoms in a face-centered cubic structure, which further allowed to calculate the photothermal conversion efficiency upon combination of theoretical and experimental data. AuNPs also showed local retention after being locally administered in in vivo models. These last results obtained by computerized tomography allow to consider these AuNPs as promising elements for a PTT system. Moreover, AuNPs showed high potential for PTT by resulting in in vitro cancer cells' viability reductions superior to 70 % once combine with 5 min of NIR irradiation.

Pluronic® F127 Hydrogel Containing Silver Nanoparticles in Skin Burn Regeneration: An Experimental Approach from Fundamental to Translational Research.

Presently, skin burns are considered one of the main public health problems and lack therapeutic options. In recent years, silver nanoparticles (AgNPs) have been widely studied, playing an increasingly important role in wound healing due to their antibacterial activity. This work is focused on the production and characterization of AgNPs loaded in a Pluronic® F127 hydrogel, as well as assessing its antimicrobial and wound-healing potential. Pluronic® F127 has been extensively explored for therapeutic applications mainly due to its appealing properties. The developed AgNPs had an average size of 48.04 ± 14.87 nm (when prepared by method C) and a negative surface charge. Macroscopically, the AgNPs solution presented a translucent yellow coloration with a characteristic absorption peak at 407 nm. Microscopically, the AgNPs presented a multiform morphology with small sizes (~50 nm). Skin permeation studies revealed that no AgNPs permeated the skin after 24 h. AgNPs further demonstrated antimicrobial activity against different bacterial species predominant in burns. A chemical burn model was developed to perform preliminary in vivo assays and the results showed that the performance of the developed AgNPs loaded in hydrogel, with smaller silver dose, was comparable with a commercial silver cream using higher doses. In conclusion, hydrogel-loaded AgNPs is potentially an important resource in the treatment of skin burns due to their proven efficacy by topical administration.

Safety of Gold Nanoparticles: From In Vitro to In Vivo Testing Array Checklist.

In recent years, gold nanoparticles (AuNPs) have aroused the interest of many researchers due to their unique physicochemical and optical properties. AuNPs are being explored in a variety of biomedical fields, either in diagnostics or therapy, particularly for localized thermal ablation of cancer cells after light irradiation. Besides the promising therapeutic potential of AuNPs, their safety constitutes a highly important issue for any medicine or medical device. For this reason, in the present work, the production and characterization of physicochemical properties and morphology of AuNPs coated with two different materials (hyaluronic and oleic acids (HAOA) and bovine serum albumin (BSA)) were firstly performed. Based on the above importantly referred issue, the in vitro safety of developed AuNPs was evaluated in healthy keratinocytes, human melanoma, breast, pancreatic and glioblastoma cancer cells, as well as in a three-dimensional human skin model. Ex vivo and in vivo biosafety assays using, respectively, human red blood cells and Artemia salina were also carried out. HAOA-AuNPs were selected for in vivo acute toxicity and biodistribution studies in healthy Balb/c mice. Histopathological analysis showed no significant signs of toxicity for the tested formulations. Overall, several techniques were developed in order to characterize the AuNPs and evaluate their safety. All these results support their use for biomedical applications.

Reference Equation of a New Incremental Step Test to Assess Exercise Capacity in the Portuguese Adult Population.

Step tests are important in community- and home-based rehabilitation programs to assess patients' exercise capacity. A new incremental step test was developed for this purpose, but its clinical interpretability is currently limited. This study aimed to establish a reference equation for this new incremental step test (IST) for the Portuguese adult population. A cross-sectional study was conducted on people without disabilities. Sociodemographic (age and sex), anthropometric (weight, height, and body mass index), smoking status, and physical activity (using the brief physical activity assessment tool) data were collected. Participants performed two repetitions of the IST and the best test was used to establish the reference equation with a forward stepwise multiple regression. An analysis comparing the results from the reference equation with the actual values was conducted with the Wilcoxon test. A total of 155 adult volunteers were recruited (60.6% female, 47.8 ± 19.7 years), and the reference equation was as follows: steps in IST = 475.52 - (4.68 × age years) + (30.5 × sex), where male = 1 and female = 0, and r2 = 60%. No significant differences were observed between the values performed and those obtained by the equation (p = 0.984). The established equation demonstrated that age and sex were the determinant variables for the variability of the results.

The Role of Rosmarinic Acid on the Bioproduction of Gold Nanoparticles as Part of a Photothermal Approach for Breast Cancer Treatment.

Breast cancer is a high-burden malignancy for society, whose impact boosts a continuous search for novel diagnostic and therapeutic tools. Among the recent therapeutic approaches, photothermal therapy (PTT), which causes tumor cell death by hyperthermia after being irradiated with a light source, represents a high-potential strategy. Furthermore, the effectiveness of PTT can be improved by combining near infrared (NIR) irradiation with gold nanoparticles (AuNPs) as photothermal enhancers. Herein, an alternative synthetic method using rosmarinic acid (RA) for synthesizing AuNPs is reported. The RA concentration was varied and its impact on the AuNPs physicochemical and optical features was assessed. Results showed that RA concentration plays an active role on AuNPs features, allowing the optimization of mean size and maximum absorbance peak. Moreover, the synthetic method explored here allowed us to obtain negatively charged AuNPs with sizes favoring the local particle accumulation at tumor site and maximum absorbance peaks within the NIR region. In addition, AuNPs were safe both in vitro and in vivo. In conclusion, the synthesized AuNPs present favorable properties to be applied as part of a PTT system combining AuNPs with a NIR laser for the treatment of breast cancer.

Biological Thermal Performance of Organic and Inorganic Aerogels as Patches for Photothermal Therapy.

Aerogels are materials with unique properties, among which are low density and thermal conductivity. They are also known for their exquisite biocompatibility and biodegradability. All these features make them attractive for biomedical applications, such as their potential use in photothermal therapy (PTT). This technique is, yet, still associated with undesirable effects on surrounding tissues which emphasizes the need to minimize the exposure of healthy regions. One way to do so relies on the use of materials able to block the radiation and the heat generated. Aerogels might be potentially useful for this purpose by acting as insulators. Silica- and pectin-based aerogels are reported as the best inorganic and organic thermal insulators, respectively; thus, the aim of this work relies on assessing the possibility of using these materials as light and thermal insulators and delimiters for PTT. Silica- and pectin-based aerogels were prepared and fully characterized. The thermal protection efficacy of the aerogels when irradiated with a near-infrared laser was assessed using phantoms and ex vivo grafts. Lastly, safety was assessed in human volunteers. Both types presented good textural properties and safe profiles. Moreover, thermal activation unveils the better performance of silica-based aerogels, confirming the potential of this material for PTT.

Nanogold-based materials in medicine: from their origins to their future.

The properties of gold-based materials have been explored for centuries in several research fields, including medicine. Multiple published production methods for gold nanoparticles (AuNPs) have shown that the physicochemical and optical properties of AuNPs depend on the production method used. These different AuNP properties have allowed exploration of their usefulness in countless distinct biomedical applications over the last few years. Here we present an extensive overview of the most commonly used AuNP production methods, the resulting distinct properties of the AuNPs and the potential application of these AuNPs in diagnostic and therapeutic approaches in biomedicine.

Safety and efficacy assessment of aerogels for biomedical applications.

The unique physicochemical properties of aerogels have made them an attractive class of materials for biomedical applications such as drug delivery, regenerative medicine, and wound healing. Their low density, high porosity, and ability to regulate the pore structure makes aerogels ideal nano/micro-structures for loading of drugs and active biomolecules. As a result of this, the number of in vitro and in vivo studies on the therapeutic efficacy of these porous materials has increased substantially in recent years and continues to be an area of great interest. However, data about their in vivo performance and safety is limited. Studies have shown that polymer-based, silica-based and some hybrid aerogels are generally regarded as safe but given that studies on the acute, subacute, and chronic toxicity for the majority of aerogel types is missing, more work is still needed. This review presents a comprehensive summary of different biomedical applications of aerogels proposed to date as well as new and innovative applications of aerogels in other areas such as decontamination. We have also reviewed their biological effect on cells and living organisms with a focus on therapeutic efficacy and overall safety (in vivo and in vitro).

Proof-of-Concept Study of Multifunctional Hybrid Nanoparticle System Combined with NIR Laser Irradiation for the Treatment of Melanoma.

The global impact of cancer emphasizes the importance of developing innovative, effective and minimally invasive therapies. In the context of superficial cancers, the development of a multifunctional nanoparticle-based system and its in vitro and in vivo safety and efficacy characterization are, herein, proposed as a proof-of-concept. This multifunctional system consists of gold nanoparticles coated with hyaluronic and oleic acids, and functionalized with epidermal growth factor for greater specificity towards cutaneous melanoma cells. This nanoparticle system is activated by a near-infrared laser. The characterization of this nanoparticle system included several phases, with in vitro assays being firstly performed to assess the safety of gold nanoparticles without laser irradiation. Then, hairless immunocompromised mice were selected for a xenograft model upon inoculation of A375 human melanoma cells. Treatment with near-infrared laser irradiation for five minutes combined with in situ administration of the nanoparticles showed a tumor volume reduction of approximately 80% and, in some cases, led to the formation of several necrotic foci, observed histologically. No significant skin erythema at the irradiation zone was verified, nor other harmful effects on the excised organs. In conclusion, these assays suggest that this system is safe and shows promising results for the treatment of superficial melanoma.

Preventive action of benztropine on platinum-induced peripheral neuropathies and tumor growth.

The endogenous cholinergic system plays a key role in neuronal cells, by suppressing neurite outgrowth and myelination and, in some cancer cells, favoring tumor growth. Platinum compounds are widely used as part of first line conventional cancer chemotherapy; their efficacy is however limited by peripheral neuropathy as a major side-effect. In a multiple sclerosis mouse model, benztropine, that also acts as an anti-histamine and a dopamine re-uptake inhibitor, induced the differentiation of oligodendrocytes through M1 and M3 muscarinic receptors and enhanced re-myelination. We have evaluated whether benztropine can increase anti-tumoral efficacy of oxaliplatin, while preventing its neurotoxicity.We showed that benztropine improves acute and chronic clinical symptoms of oxaliplatin-induced peripheral neuropathies in mice. Sensory alterations detected by electrophysiology in oxaliplatin-treated mice were consistent with a decreased nerve conduction velocity and membrane hyperexcitability due to alterations in the density and/or functioning of both sodium and potassium channels, confirmed by action potential analysis from ex-vivo cultures of mouse dorsal root ganglion sensory neurons using whole-cell patch-clamp. These alterations were all prevented by benztropine. In oxaliplatin-treated mice, MBP expression, confocal and electronic microscopy of the sciatic nerves revealed a demyelination and confirmed the alteration of the myelinated axons morphology when compared to animals injected with oxaliplatin plus benztropine. Benztropine also prevented the decrease in neuronal density in the paws of mice injected with oxaliplatin. The neuroprotection conferred by benztropine against chemotherapeutic drugs was associated with a lower expression of inflammatory cytokines and extended to diabetic-induced peripheral neuropathy in mice.Mice receiving benztropine alone presented a lower tumor growth when compared to untreated animals and synergized the anti-tumoral effect of oxaliplatin, a phenomenon explained at least in part by benztropine-induced ROS imbalance in tumor cells.This report shows that blocking muscarinic receptors with benztropine prevents peripheral neuropathies and increases the therapeutic index of oxaliplatin. These results can be rapidly transposable to patients as benztropine is currently indicated in Parkinson's disease in the United States.

Evaluation of the Spider (Phlogiellus genus) Phlotoxin 1 and Synthetic Variants as Antinociceptive Drug Candidates.

Over the two last decades, venom toxins have been explored as alternatives to opioids to treat chronic debilitating pain. At present, approximately 20 potential analgesic toxins, mainly from spider venoms, are known to inhibit with high affinity the NaV1.7 subtype of voltage-gated sodium (NaV) channels, the most promising genetically validated antinociceptive target identified so far. The present study aimed to consolidate the development of phlotoxin 1 (PhlTx1), a 34-amino acid and 3-disulfide bridge peptide of a Phlogiellus genus spider, as an antinociceptive agent by improving its affinity and selectivity for the human (h) NaV1.7 subtype. The synthetic homologue of PhlTx1 was generated and equilibrated between two conformers on reverse-phase liquid chromatography and exhibited potent analgesic effects in a mouse model of NaV1.7-mediated pain. The effects of PhlTx1 and 8 successfully synthetized alanine-substituted variants were studied (by automated whole-cell patch-clamp electrophysiology) on cell lines stably overexpressing hNaV subtypes, as well as two cardiac targets, the hCaV1.2 and hKV11.1 subtypes of voltage-gated calcium (CaV) and potassium (KV) channels, respectively. PhlTx1 and D7A-PhlTx1 were shown to inhibit hNaV1.1-1.3 and 1.5-1.7 subtypes at hundred nanomolar concentrations, while their affinities for hNaV1.4 and 1.8, hCaV1.2 and hKV11.1 subtypes were over micromolar concentrations. Despite similar analgesic effects in the mouse model of NaV1.7-mediated pain and selectivity profiles, the affinity of D7A-PhlTx1 for the NaV1.7 subtype was at least five times higher than that of the wild-type peptide. Computational modelling was performed to deduce the 3D-structure of PhlTx1 and to suggest the amino acids involved in the efficiency of the molecule. In conclusion, the present structure-activity relationship study of PhlTx1 results in a low improved affinity of the molecule for the NaV1.7 subtype, but without any marked change in the molecule selectivity against the other studied ion channel subtypes. Further experiments are therefore necessary before considering the development of PhlTx1 or synthetic variants as antinociceptive drug candidates.

From identification to functional characterization of cyriotoxin-1a, an antinociceptive toxin from the spider Cyriopagopus schioedtei.

BACKGROUND AND PURPOSE: The NaV 1.7 channel is highly expressed in dorsal root ganglia of the sensory nervous system and plays a central role in the pain signalling process. We investigated a library prepared from original venoms of 117 different animals to identify new selective inhibitors of this target. EXPERIMENTAL APPROACH: We used high throughput screening of a large venom collection using automated patch-clamp experiments on human voltage-gated sodium channel subtypes and then in vitro and in vivo electrophysiological experiments to characterize the active peptides that have been purified, sequenced, and chemically synthesized. Analgesic effects were evaluated in vivo in mice models. KEY RESULTS: We identified cyriotoxin-1a (CyrTx-1a), a novel peptide isolated from Cyriopagopus schioedtei spider venom, as a candidate for further characterization. This 33 amino acids toxin belongs to the inhibitor cystine knot structural family and inhibits hNaV 1.1-1.3 and 1.6-1.7 channels in the low nanomolar range, compared to the micromolar range for hNaV 1.4-1.5 and 1.8 channels. CyrTx-1a was 920 times more efficient at inhibiting tetrodotoxin (TTX)-sensitive than TTX-resistant sodium currents recorded from adult mouse dorsal root ganglia neurons and in vivo electrophysiological experiments showed that CyrTx-1a was approximately 170 times less efficient than huwentoxin-IV at altering mouse skeletal neuromuscular excitability properties. CyrTx-1a exhibited an analgesic effect in mice by increasing reaction time in the hot-plate assay. CONCLUSIONS AND IMPLICATIONS: The pharmacological profile of CyrTx-1a paves the way for further molecular engineering aimed to optimize the potential antinociceptive properties of this peptide.