Vincristine-induced peripheral neuropathy is driven by canonical NLRP3 activation and IL-1ß release.

Vincristine is an important component of many regimens used for pediatric and adult malignancies, but it causes a dose-limiting sensorimotor neuropathy for which there is no effective treatment. This study aimed to delineate the neuro-inflammatory mechanisms contributing to the development of mechanical allodynia and gait disturbances in a murine model of vincristine-induced neuropathy, as well as to identify novel treatment approaches. Here, we show that vincristine-induced peripheral neuropathy is driven by activation of the NLRP3 inflammasome and subsequent release of interleukin-1ß from macrophages, with mechanical allodynia and gait disturbances significantly reduced in knockout mice lacking NLRP3 signaling pathway components, or after treatment with the NLRP3 inhibitor MCC950. Moreover, treatment with the IL-1 receptor antagonist anakinra prevented the development of vincristine-induced neuropathy without adversely affecting chemotherapy efficacy or tumor progression in patient-derived medulloblastoma xenograph models. These results detail the neuro-inflammatory mechanisms leading to vincristine-induced peripheral neuropathy and suggest that repurposing anakinra may be an effective co-treatment strategy to prevent vincristine-induced peripheral neuropathy.

Pharmacological activity and NMR solution structure of the leech peptide HSTX-I.

The role of voltage-gated sodium (NaV) channels in pain perception is indisputable. Of particular interest as targets for the development of pain therapeutics are the tetrodotoxin-resistant isoforms NaV1.8 and NaV1.9, based on animal as well as human genetic studies linking these ion channel subtypes to the pathogenesis of pain. However, only a limited number of inhibitors selectively targeting these channels have been reported. HSTX-I is a peptide toxin identified from saliva of the leech Haemadipsa sylvestris. The native 23-residue peptide, stabilised by two disulfide bonds, has been reported to inhibit rat NaV1.8 and mouse NaV1.9 with low micromolar activity, and may therefore represent a scaffold for development of novel modulators with activity at human tetrodotoxin-resistant NaV isoforms. We synthetically produced this hydrophobic peptide in high yield using a one-pot oxidation and single step purification and determined the three-dimensional solution structure of HSTX-I using NMR solution spectroscopy. However, in our hands, the synthetic HSTX-I displayed only very modest activity at human NaV1.8 and NaV1.9, and lacked analgesic efficacy in a murine model of inflammatory pain.

High-Throughput Fluorescence Assays for Ion Channels and GPCRs.

Ca2+, Na+ and K+- permeable ion channels as well as GPCRs linked to Ca2+ release are important drug targets. Accordingly, high-throughput fluorescence plate reader assays have contributed substantially to drug discovery efforts and pharmacological characterization of these receptors and ion channels. This chapter describes some of the basic properties of the fluorescent dyes facilitating these assay approaches as well as general methods for establishment and optimisation of fluorescence assays for ion channels and Gq-coupled GPCRs.

Development of a high-throughput fluorescent no-wash sodium influx assay.

Voltage-gated sodium channels (NaVs) are key therapeutic targets for pain, epilepsy and cardiac arrhythmias. Here we describe the development of a no-wash fluorescent sodium influx assay suitable for high-throughput screening and characterization of novel drug leads. Addition of red-violet food dyes (peak absorbance range 495-575 nm) to assays in HEK293 cells heterologously expressing hNaV1.1-1.8 effectively quenched background fluorescence of the sodium indicator dye Asante NaTRIUM Green-2 (ANG-2; peak emission 540 nm), negating the need for a wash step. Ponceau 4R (1 mM) was identified as a suitable quencher, which had no direct effect on NaV channels as assessed by patch-clamp experiments, and did not alter the pharmacology of the NaV1.1-1.7 activator veratridine (EC50 10-29 µM) or the NaV1.1-1.8 inhibitor tetracaine (IC50's 6-66 µM). In addition, we also identified that the food dyes Ponceau 4R, Brilliant Black BN, Allura Red and Amaranth are effective at quenching the background fluorescence of the calcium indicator dyes fluo-4, fura-2 and fura-5F, identifying them as potential inexpensive alternatives to no-wash calcium ion indicator kits. In summary, we have developed a no-wash fluorescent sodium influx assay suitable for high-throughput screening based on the sodium indicator dye ANG-2 and the quencher Ponceau 4R.

Toxins as tools: Fingerprinting neuronal pharmacology.

Toxins have been used as tools for decades to study the structure and function of neuronal ion channels and receptors. The biological origin of these toxins varies from single cell organisms, including bacteria and algae, to complex multicellular organisms, including a wide variety of plants and venomous animals. Toxins are a structurally and functionally diverse group of compounds that often modulate neuronal function by interacting with an ion channel or receptor. Many of these toxins display high affinity and exquisite selectivity, making them valuable tools to probe the structure and function of neuronal ion channels and receptors. This review article provides an overview of the experimental techniques used to assess the effects that toxins have on neuronal function, as well as discussion on toxins that have been used as tools, with a focus on toxins that target voltage-gated and ligand-gated ion channels.

Sodium Channels and Venom Peptide Pharmacology.

Venomous animals including cone snails, spiders, scorpions, anemones, and snakes have evolved a myriad of components in their venoms that target the opening and/or closing of voltage-gated sodium channels to cause devastating effects on the neuromuscular systems of predators and prey. These venom peptides, through design and serendipity, have not only contributed significantly to our understanding of sodium channel pharmacology and structure, but they also represent some of the most phyla- and isoform-selective molecules that are useful as valuable tool compounds and drug leads. Here, we review our understanding of the basic function of mammalian voltage-gated sodium channel isoforms as well as the pharmacology of venom peptides that act at these key transmembrane proteins.

Self-Care for Older People (SCOPE): a cluster randomized controlled trial of self-care training and health outcomes in low-income elderly in Singapore.

UNLABELLED: Population aging is associated with a higher prevalence of chronic health conditions. Previous studies have shown that older persons, specifically those with chronic conditions, often lack sufficient knowledge about their condition and thus frequently have poor self-care skills. Efforts to increase general health screenings and improve access to chronic condition management resources are hampered by a lack of disease and health awareness. Self-Care for Older People (SCOPE) study, a cluster randomized controlled trial in Singapore, was designed to evaluate the impact of a self-care program for chronic disease awareness and management of specific health measures and quality of life of older people over eighteen months. SCOPE provided self-care education targeted at older people with low income and low education in order to improve health-related knowledge. A total of 378 low-income older people with no or minimal disability, defined as having difficulty in one or more activities of daily living (ADL), were recruited from senior activity centers. The measurements taken included self-reported health conditions, health and disease knowledge questions, and biomarkers (HbA1c, blood pressure, peak expiratory flow, lipid panel, albumin, and creatinine). SCOPE was also designed to provide information for policy makers on chronic disease burden and healthcare facility utilization among community-dwelling older adults. TRIAL REGISTRATION NUMBER: NCT01672177.