Detection of retinal dysfunction induced by HCN channel inhibitors using multistep light stimulus and long-duration light stimulus ERG in rats.

Ivabradine, a hyperpolarization-activated cyclic nucleotide-gated (HCN) channel inhibitor, has been reported to induce photosensitivity-related visual disturbances such as phosphene in humans. Ivabradine-induced visual disturbances are caused by inhibition of HCN channels in the retina, and the mechanisms have been verified using HCN channel knockout mice and electroretinography (ERG). However, in rats, classical ERG using single flash light stimulus with standard analyses of waveform amplitude and latency has not revealed abnormal retinal function after administration of ivabradine. To verify whether retinal dysfunction after ivabradine administration was detectable in rats, we performed ERG using multistep flash light stimulation at the time when plasma concentration of ivabradine was high. Furthermore, the mechanism of the change in the waveform that appeared after the b-wave was investigated. Ivabradine and cilobradine, a selective HCN channel inhibitor, were administered subcutaneously to rats at 4-40 mg/kg as a single dose, and flash or long-duration ERG recordings at each light stimulus luminance were conducted 1.5 h after administration. Plasma and retinal concentrations of both compounds were measured immediately after the ERG recordings. In the flash ERG, prolongation of a- and/or b-wave latencies were detected at each light stimulus, and dose-dependent waveform changes after the b-wave were recorded at the specific light stimulus luminance for both compounds. These ERG changes increased in response to increasing plasma and retinal concentrations for both ivabradine and cilobradine. In the long-duration light stimulus ERG, a change in the waveform of the b-wave trough and attenuation of the c-wave were recorded, suggesting that the feedback control in the photoreceptor cells may be inhibited. This study revealed that the retinal dysfunction by HCN channel inhibitors in rats can be detected by multistep light stimulus ERG. Additionally, we identified that the inhibition of feedback current and the sustained responses in the photoreceptor cells cause the retinal dysfunction of HCN channel inhibitors in rats.

Structural analysis reveals TLR7 dynamics underlying antagonism.

Toll-like receptor 7 (TLR7) recognizes both microbial and endogenous RNAs and nucleosides. Aberrant activation of TLR7 has been implicated in several autoimmune diseases including systemic lupus erythematosus (SLE). Here, by modifying potent TLR7 agonists, we develop a series of TLR7-specific antagonists as promising therapeutic agents for SLE. These compounds protect mice against lethal autoimmunity. Combining crystallography and cryo-electron microscopy, we identify the open conformation of the receptor and reveal the structural equilibrium between open and closed conformations that underlies TLR7 antagonism, as well as the detailed mechanism by which TLR7-specific antagonists bind to their binding pocket in TLR7. Our work provides small-molecule TLR7-specific antagonists and suggests the TLR7-targeting strategy for treating autoimmune diseases.

A Novel Phosphodiesterase 1 Inhibitor DSR-141562 Exhibits Efficacies in Animal Models for Positive, Negative, and Cognitive Symptoms Associated with Schizophrenia.

In our drug discovery program, we identified a novel orally available and brain-penetrant phosphodiesterase (PDE) 1 inhibitor, 3-methyl-7-(tetrahydro-2H-pyran-4-yl)-2-{[trans-4-(trifluoromethyl)cyclohexyl]-methoxy}imidazo[5,1-f][1,2,4]triazin-4(3H)-one (DSR-141562). In the present study, we characterized the preclinical profile of DSR-141562. This compound has preferential selectivity for predominantly brain-expressed PDE1B over other PDE1 family members, and high selectivity for the PDE1 family over other PDE families and 65 other tested biologic targets. Oral administration of DSR-141562 at 10 mg/kg slightly elevated the cGMP concentration, and it potently enhanced the increase of cGMP induced by a dopamine D1 receptor agonist in mouse brains. The cGMP level in monkey cerebrospinal fluid was also elevated after treatment with DSR-141562 at 30 and 100 mg/kg and could be used as a translational biomarker. Since PDE1B is believed to regulate dopaminergic and glutamatergic signal transduction, we evaluated the effects of this compound using schizophrenia-related behavioral assays. DSR-141562 at 3-30 mg/kg potently inhibited methamphetamine-induced locomotor hyperactivity in rats, while it had only minimal effects on the spontaneous locomotor activity. Furthermore, DSR-141562 at 1-100 mg/kg did not induce any signs of catalepsy in rats. DSR-141562 at 0.3-3 mg/kg reversed social interaction and novel object recognition deficits induced by repeated treatment with an N-methyl-D-aspartate receptor antagonist, phencyclidine, in mice and rats, respectively. In common marmosets, DSR-141562 at 3 and 30 mg/kg improved the performance in object retrieval with detour tasks. These results suggest that DSR-141562 is a therapeutic candidate for positive, negative, and cognitive symptoms in schizophrenia. SIGNIFICANCE STATEMENT: This is the first paper showing that a phosphodiesterase 1 inhibitor is efficacious in animal models for positive and negative symptoms associated with schizophrenia. Furthermore, we demonstrated that this compound improved cognitive function in the common marmoset, a nonhuman primate.

Availability of multistep light stimulus method for evaluation of visual dysfunctions.

In the field of drug safety research, electroretinography (ERG) is commonly conducted according to the international standard method propounded by the International Society for Clinical Electrophysiology of Vision (ISCEV) in recent years. However, various ERG methods other than the ISCEV standard method are also utilized depending on the intended purpose of the evaluation. In this study, we investigated the availability of a multistep light stimulus method for evaluation of rod function in Long-Evans rats using sildenafil, which is known to inhibit phosphodiesterase 6 (PDE6) in phototransduction and induce visual dysfunctions in humans. Sildenafil was orally administered to female Long-Evans rats at doses of 15, 50, and 150 mg/kg, and ERG was recorded at 1.5 h after treatment. In addition to a - 2.0 log cd·s/m2 stimulus corresponding to dark-adapted 0.01 ERG in the ISCEV standard method, light stimulus intensities of -4.5, -4.0, -3.0, -1.0, 0.0, and +1.0 log cd·s/m2 were applied for multistep ERG recording. The amplitude and implicit time of the a-wave were decreased and prolonged, respectively, at doses of ≥50 mg/kg. The amplitude and implicit time of the b-wave were decreased and prolonged, respectively, at all doses. However, the b-wave at 15 mg/kg was only diminished or attenuated at ≤ - 3.0 log cd·s/m2, as weaker stimuli than dark-adapted 0.01 ERG in the ISCEV standard protocol. These findings suggest that sildenafil triggers visual dysfunctions through PDE6 inhibition, and indicate that the multistep light stimulus method is highly sensitive for detection of phototransduction abnormalities in retinal rod cells.