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 (IC 50 5 103 nM), selective TRPM8 antagonist, PF-05105679It demonstrated activity in vivo in the guinea pig bladder ice water and menthol challenge tests with an IC 50 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 IC 50 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

Evidence of emerging Plasmodium falciparum resistance to artemisinin-based combination therapies, documented in western Cambodia, underscores the continuing need to identify new antimalarial combinations. Given recent reports of the resurgence of chloroquine-sensitive P. falciparum parasites in Malawi, after the enforced and prolonged withdrawal of this drug, and indications of a possible synergistic interaction with the macrolide azithromycin, we sought to further characterize chloroquine-azithromycin combinations for their in vitro and in vivo antimalarial properties. In vitro 96-h susceptibility testing of chloroquine-azithromycin combinations showed mostly additive interactions against freshly cultured P. falciparum field isolates obtained from Mali. Some evidence of synergy, however, was apparent at the fractional 90% inhibitory concentration level. Additional in vitro testing highlighted the resistance reversal properties of amlodipine for both chloroquine and quinine. In vivo experiments, using the Peters 4-day suppressive test in a P. yoelii mouse model, revealed up to 99.9% suppression of parasitemia following treatment with chloroquine-azithromycin plus the R enantiomer of amlodipine. This enantiomer was chosen because it does not manifest the cardiac toxicities observed with the racemic mixture. Pharmacokinetic/pharmacodynamic analyses in this rodent model and subsequent extrapolation to a 65-kg adult led to the estimation that 1.8 g daily of R-amlodipine would be required to achieve similar efficacy in humans, for whom this is likely an unsafe dose. While these data discount amlodipine as an additional partner for chloroquine-based combination therapy, our studies continue to support azithromycin as a safe and effective addition to antimalarial combination therapies.In the last decade, artemisinin-based combination therapies (ACTs) have become widely adopted as first-line treatment in almost all countries where malaria is endemic (90). These drug combinations display excellent clinical efficacy against Plasmodium falciparum infection, yet recent studies from western Cambodia report decreases in parasite clearance rates following artesunate monotherapy or artesunate-mefloquine combination therapy (18,59,60,76,92). The possible emergence of resistance underscores a clear need to find alternative regimens. While several promising agents are in the pipeline, the development of antimalarial drugs that are effective, well tolerated, and safe remains a very challenging task (6,27,77,89). In addition, the new paradigm of antimalarial therapies based on the combination of drugs that have additive or preferably synergistic properties raises the threshold for drug discovery even further.Chloroquine (CQ) was the most important drug for the treatment of malaria for many decades, until widespread resistance led to its replacement, most recently by ACTs (21,93). Recent data from Malawi and Kenya have shown that in those regions, the removal of CQ from local use has led to the resurgence of CQ-sensitive (CQS) strain...

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. Hepatitis C virus (HCV) is a leading cause of chronic liver disease, liver cancer, and cirrhosis, with an infection prevalence of 2 to 3% worldwide (9). Approximately 70% of people who become infected with HCV are unable to clear the virus naturally and develop a chronic infection. The current treatment regimen requires 6 to 12 months of combination therapy with pegylated interferon (pegIFN) (weekly injections) and ribavirin (RBV) (daily oral dosing). This standard of care results in cure rates of ϳ50% in patients with the most common genotype virus (genotype 1) and is associated with multiple side effects (2, 6, 13). Therefore, new, improved treatments are urgently needed.Multiple candidate small-molecule therapeutics that specifically target the virus (direct-acting antivirals [DAAs]) are currently in development, including inhibitors of the HCV NS3 protease, NS5A protein, and NS5B RNA-dependent RNA polymerase (10,22). The most advanced compounds, telaprevir and boceprevir, are NS3 protease inhibitors and have been approved for use in combination with pegIFN and RBV. In phase 3 clinical studies, these compounds significantly increased rates of sustained viral response (SVR) compared to the standard of care alone in both treatment-naïve and treatment-experienced patient populations (16). However, both compounds have significant side effects, including severe rash and anemia, which contribute to the overall adverse-event burden associated with treatment. Patients failing therapy also typically develop drug class resistance that may prevent effective retreatment with other HCV protease inhibitors (19). Therefore, more tolerable and efficacious DAAs with unique mechanisms of a...