The transient receptor potential (subfamily M, member 8; TRPM8) is a nonselective cation channel localized in primary sensory neurons, and is a candidate for cold thermosensing, mediation of cold ...pain, and bladder overactivity. Studies with TRPM8 knockout mice and selective TRPM8 channel blockers demonstrate a lack of cold sensitivity and reduced cold pain in various rodent models. Furthermore, TRPM8 blockers significantly lower body temperature. We have identified a moderately potent (IC50 = 103 nM), selective TRPM8 antagonist, PF-05105679 (R)-3-(1-(4-fluorophenyl)ethyl)(quinolin-3-ylcarbonyl)aminomethylbenzoic acid. It demonstrated activity in vivo in the guinea pig bladder ice water and menthol challenge tests with an IC50 of 200 nM and reduced core body temperature in the rat (at concentrations >1219 nM). PF-05105679 was suitable for acute administration to humans and was evaluated for effects on core body temperature and experimentally induced cold pain, using the cold pressor test. Unbound plasma concentrations greater than the IC50 were achieved with 600- and 900-mg doses. The compound displayed a significant inhibition of pain in the cold pressor test, with efficacy equivalent to oxycodone (20 mg) at 1.5 hours postdose. No effect on core body temperature was observed. An unexpected adverse event (hot feeling) was reported, predominantly periorally, in 23 and 36% of volunteers (600- and 900-mg dose, respectively), which in two volunteers was nontolerable. In conclusion, this study supports a role for TRPM8 in acute cold pain signaling at doses that do not cause hypothermia.
Glaucoma is one of the leading causes of blindness, and there is an ongoing need for new therapies. Recent studies indicate that cell transplantation using Müller glia may be beneficial, but there is ...a need for novel sources of cells to provide therapeutic benefit. In this study, we have isolated Müller glia from retinal organoids formed by human induced pluripotent stem cells (hiPSCs) in vitro and have shown their ability to partially restore visual function in rats depleted of retinal ganglion cells by NMDA. Based on the present results, we suggest that Müller glia derived from retinal organoids formed by hiPSC may provide an attractive source of cells for human retinal therapies, to prevent and treat vision loss caused by retinal degenerative conditions. Stem Cells Translational Medicine 2019;8:775&784
Purified populations of Müller glial cells were isolated from retinal organoids derived from human iPSC. Intra‐vitreal transplantation of these cells into rat eyes depleted of RGC by NMDA caused partial restoration of visual function, as assessed by improvement of the retinal electrophysiological responses. These findings highlight the potential of iPSC as a potential source of therapeutic cells for retinal degeneration.
Filibuvir (PF-00868554) is an investigational nonnucleoside inhibitor of the hepatitis C virus (HCV) nonstructural 5B (NS5B) RNA-dependent RNA polymerase currently in development for treating chronic ...HCV infection. The aim of this study was to characterize the selection of filibuvir-resistant variants in HCV-infected individuals receiving filibuvir as short (3- to 10-day) monotherapy. We identified amino acid M423 as the primary site of mutation arising upon filibuvir dosing. Through bulk cloning of clinical NS5B sequences into a transient-replicon system, and supported by site-directed mutagenesis of the Con1 replicon, we confirmed that mutations M423I/T/V mediate phenotypic resistance. Selection in patients of an NS5B mutation at M423 was associated with a reduced replicative capacity in vitro relative to the pretherapy sequence; consistent with this, reversion to wild-type M423 was observed in the majority of patients following therapy cessation. Mutations at NS5B residues R422 and M426 were detected in a small number of patients at baseline or the end of therapy and also mediate reductions in filibuvir susceptibility, suggesting these are rare but clinically relevant alternative resistance pathways. Amino acid variants at position M423 in HCV NS5B polymerase are the preferred pathway for selection of viral resistance to filibuvir in vivo.
Sensory processing in the spinal cord during disease states can reveal mechanisms for novel treatments, yet very little is known about pain processing at this level in the most commonly used animal ...models of articular pain. Here we report a test of the prediction that two clinically effective compounds, naproxen (an NSAID) and oxycodone (an opiate), are efficacious in reducing the response of spinal dorsal horn neurons to noxious knee joint rotation in the monosodium iodoacetate (MIA) sensitized rat. The overall objective for these experiments was to develop a high quality in vivo electrophysiology assay to confidently test novel compounds for efficacy against pain. Given the recent calls for improved preclinical experimental quality we also developed and implemented an Assay Capability Tool to determine the quality of our assay and ensure the quality of our results. Spinal dorsal horn neurons receiving input from the hind limb knee joint were recorded in anesthetized rats 14 days after they were sensitized with 1 mg of MIA. Intravenous administered oxycodone and naproxen were each tested separately for their effects on phasic, tonic, ongoing and afterdischarge action potential counts in response to innocuous and noxious knee joint rotation. Oxycodone reduced tonic spike counts more than the other measures, doing so by up to 85%. Tonic counts were therefore designated the primary endpoint when testing naproxen which reduced counts by up to 81%. Both reductions occurred at doses consistent with clinically effective doses for osteoarthritis. These results demonstrate that clinically effective doses of standard treatments for osteoarthritis reduce pain processing measured at the level of the spinal cord for two different mechanisms. The Assay Capability Tool helped to guide experimental design leading to a high quality and robust preclinical assay to use in discovering novel treatments for pain.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Despite the efforts, cost, and extensive use of animals for nonclinical research, only a small number of studies have methodically compared findings from animal toxicology studies to those from human ...clinical trials. Impediments to understanding the translation of nonclinical safety have included the lack of easy access to data and the need for extensive data curation given the diverse terminologies, formats, and data platforms in use. SEND and SDTM study data standards, developed by CDISC and about to become mandated by FDA, can address this and other drug development issues by facilitating access to data in ways that are not currently feasible. A consistent data standard across clinical and nonclinical will discourage the development of data silos, which easily become obstacles to data sharing and maximizing the value of animal and human data. The confluence of rapid scientific advances, increasingly larger quantities of diverse data, technological advances in data mining, and the FDA's requirements for standardized study data create new opportunities for the advancement of drug development and for refinement in the way we use animals.
Gaucher Disease (GD) is an inherited metabolic disorder caused by mutations in the GBA1 gene. It can manifest with severe neurodegeneration and visceral pathology. The most acute neuronopathic form ...(nGD), for which there are no curative therapeutic options, is characterised by devastating neuropathology and death during infancy. In this study, we investigated the therapeutic benefit of systemically delivered AAV9 vectors expressing the human GBA1 gene at two different doses comparing a neuronal-selective promoter with ubiquitous promoters. Our results highlight the importance of a careful evaluation of the promoter sequence used in gene delivery vectors, suggesting a neuron-targeted therapy leading to high levels of enzymatic activity in the brain but lower GCase expression in the viscera, might be the optimal therapeutic strategy for nGD.
Getting our ACT together Gore, Katrina; Stanley, Phil
Significance (Oxford, England),
06/2015, Letnik:
12, Številka:
3
Journal Article
Results that cannot be replicated during drug trials cost pharma companies millions. Katrina Gore and Phil Stanley describe a tool developed by Pfizer to improve the quality of preclinical study ...design and analysis
Katrina Gore and Phil Stanley describe a tool developed by Pfizer to improve the quality of preclinical study design and analysis
Numerous articles in Nature, Science, Pharmacology Research and Perspectives, and other biomedical research journals over the past decade have highlighted that research is plagued by findings that ...are not reliable and cannot be reproduced. Poor experiments can occur, in part, as a consequence of inadequate statistical thinking in the experimental design, conduct and analysis. As it is not feasible for statisticians to be involved in every preclinical experiment many of the same journals have published guidelines on good statistical practice. Here, we outline a tool that addresses the root causes of irreproducibility in preclinical research in the pharmaceutical industry. The Assay Capability Tool uses 13 questions to guide scientists and statisticians during the development of in vitro and in vivo assays. It promotes the absolutely essential experimental design and analysis strategies and documents the strengths, weaknesses, and precision of an assay. However, what differentiates it from other proposed solutions is the emphasis on how the resulting data will be used. An assay can be assigned a low, medium, or high rating to indicate the level of confidence that can be afforded when making important decisions using data from that assay. This provides transparency on the appropriate interpretation of the assay's results in the light of its current capability. We suggest that following a well‐defined process during assay development and use such as that laid out within the Assay Capability Tool means that whatever the results, positive or negative, a researcher can have confidence to make decisions upon and publish their findings.
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