In Japan, 14 opioids with a variety of dosage forms, such as oral preparation, injection, transdermal patch, transmucosal and suppository are clinically available as analgesics, and their use properly depends on the type and degree of pain and patient condition. Fundamentally, strong opioids are restricted to only treatment of cancer pain, while the indication of fentanyl transdermal patch and oxycodone tamper resistant tablet has been expanded for the treatment of non-cancer chronic pain. In US, the additional indication of opioids to chronic pain has led to prolongation and generalization of opioid use, which may contribute to “opioid crisis” in which opioid-related death strikingly increased due to opioid abuse and overdose-induced respiratory depression. Currently, opioid-related abuse and death have not been evident in Japan, while abuse of antitussive opioids (codeine and dihydrocodeine) are recently seen as a problem, which may suggest the sense of guilt to abuse legally-uncontrolled drugs (ex. weak opioids, antitussives or hypnotics) may be low in Japanese. In this review, I will summarize current status and issues in clinical use of opioid analgesics, and will talk about the necessity and expectation for development of novel analgesics without serious adverse reactions such as addiction and respiratory depression.

Many studies suggest opioid receptor (OPr) dimerization modulates the pharmacological properties of opiates. Specifically, heteromerization between OPr types has been reported to lead to changes in intracellular signaling. Thus, ligands targeting heteromers are expected to be novel therapeutic targets with reduced side effects. The heteromers of mu (MOPr) and delta (DOPr) are detected in brain regions involved in pain processing. The bivalent ligand or small molecule were identified as a MOPr-DOPr targeting ligand. These ligands exhibit antinociceptive properties similar to that of morphine with lesser antinociceptive tolerance as compared to morphine. Studies exploring the in vivo regulation of MOPr-DOPr heteromers, showed chronic morphine administration leads to an upregulation of these heteromers in select brain regions. Exploration of mechanisms underlying this phenomenon led us to the G protein-coupled receptor chaperone, RTP4, that is induced by chronic morphine and facilitates the heteromerization of MOPr and DOPr. In this review, I will introduce the simulated structure or property of MOPr-DOPr heteromer, its targeting ligands, and its intracellular regulatory mechanism that include a key molecule like RTP4 that could serve as a scaffold for the development of novel therapeutic drugs with reduced adverse effects, and hence may take place of the conventional clinical opioids.

After the identification of nociceptin/orphanin FQ (N/OFQ) peptide (NOP) and its cognate receptor, the unique functional profiles of the N/OFQ-NOP receptor system have been uncovered. NOP receptors are distributed in the key regions that regulate pain and reward processing in the central nervous system. In non-human primates (NHPs), activation of the NOP receptor causes antinociception and anti-hypersensitivity via spinal and supraspinal effects. Moreover, activation of the NOP receptor attenuates dopaminergic transmission and potentiates mu-opioid peptide (MOP) receptor-mediated analgesia. Here, we highlight the functional profiles of bifunctional NOP and MOP receptor agonists based on their promising effects for the treatment of pain and drug abuse. Bifunctional NOP/MOP receptor “partial” agonists, such as AT-121, BU08028, and BU10038, exert potent analgesic effects without MOP receptor-related side effects such as abuse liability, respiratory depression, physical dependence, and itching in NHPs. These novel NOP/MOP receptor agonists reduce rewarding and the reinforcing effects of abused drugs. Furthermore, a mixed NOP/opioid receptor “full” agonist, cebranopadol, is undergoing several clinical trials, and the therapeutic advantage of the coactivation of NOP and MOP receptors has also been confirmed in humans. Therefore, this class of drugs that coactivate NOP and MOP receptors proposes a wide therapeutic range with fewer side effects, indicating a greater potential for the development of novel safer opioid analgesics.

Doxorubicin (DOX)-induced cardiomyopathy has a poor prognosis. No early detection or effective treatment methods are available in clinical. The mechanisms of cardiotoxicity were considered as oxidative stress and apoptosis in cardiomyocytes. However, the effect of DOX on cardiac fibroblasts remains to be developed. We investigated the direct effect of DOX on the function of human cardiac fibroblasts (HCFs) independently of cell death pathway. Animal study showed that lower dose of DOX (4 mg/kg/week for 3 weeks, i.p.) than a toxic cumulate dose, induced perivascular fibrosis without cell death in hear of mice. DOX increased the protein expression of α-SMA (a marker of trans-differentiation) in HCFs culture cells, indicating that DOX promoted the trans-differentiation of HCFs into myofibroblast. DOX also increased the mRNA and protein expression of matrix metalloproteinase (MMP)-1 in less than 0.1 μM which did not induce cell apoptosis of HCFs cells via PI3K/Akt pathway in HCFs. DOX increased Interleukin-6 (IL-6) via transforming growth factor (TGF)-β/Smad pathway. In addition, DOX induced the mitochondrial damage and increased the expression of Interleukin-1 (IL-1) via stress-activated protein kinases (SAPK)/ c-Jun NH-2termial kinase (JNK). A peroxisome proliferator-activated receptor gamma (PPARγ) agonist, pioglitazone hydrochloride attenuated the expression of fibrotic marker such as α-SMA and galectin-3 and collagen1 via SAPK/JNK signaling. Pioglitazone also suppressed DOX-induced early fibrotic response in vivo. In conclusion, these findings suggested that low dose DOX induced reactive fibrotic change of cardiac fibroblasts via cell death-independent pathway. There may be potentially new mechanisms of DOX induced cardiotoxicity in clinical usage.

Non-alcoholic steatohepatitis (NASH) is a common risk factor for fibrosis, cirrhosis, and a predisposing factor for the development of hepatocellular carcinoma. Recently, incidence of NASH has increased due to an increase in metabolic syndrome. Connexin (Cx)32, a hepatocyte gap-junction protein, plays an important role in liver tissue homeostasis; Cx32 dominant-negative transgenic rat (Cx32ΔTg) has much decreased gap-junctional intercellular communication, and high susceptibility to carcinogens. We found for the first time that Cx32 has play suppressive roles in inflammation and fibrosis of NASH using Cx32ΔTg received methionine-choline deficient diet (MCDD). Elevation of reactive oxygen species (ROS) play important roles in progression of NASH and elimination of ROS by antioxidant luteolin inhibited NASH in the Cx32ΔTg-MCDD model. This model had histological changes similar to those of human NASH, but was not accompanied by the metabolic syndrome such as obesity and insulin resistance. Therefore, we further established an improved NASH model. Cx32ΔTg rats and wild-type rats were fed a high-fat diet (HFD) and dimethylnitrosamine to induce NASH with metabolic syndrome. The HFD and DMN increased body, liver, and visceral fat weights in both genotypes. Serum insulin level and HOMA-IR score in Cx32ΔTg rats were higher than those in wild-type rats. Elevation of serum hepatic enzymes (AST, ALT), inflammatory cytokine expressions, α-smooth muscle actin expression, progression of steatohepatitis and fibrosis were induced by HFD and dimethylnitrosamine especially in Cx32ΔTg rats. These results indicate Cx32 dysfunction promoted the development of NASH and fibrosis accompanied by metabolic syndrome through accumulation of oxidative stress.

Physiologically, urine from the subject with normal kidney function does not contain detectable level of glucose unless otherwise renal glycosuria. Sodium glucose transporter (SGLT) families in proximal tubules of the kidney play detrimental role to reabsorb the filtered glucose. Recently, the inhibitors for the SGLT2 are available for clinical use for purposing the urinary glucose excretion and lowering blood glucose level. Unexpectedly, the SGLT2 inhibitors have become famous for its cardio-renal protective effects with unknown mechanism. We have so far explored how its inhibition changes cell fate, how the drug affects glucose uptake in non-diabetic kidney, and if the drug suppresses the development of fibrosis. In this review, we will summarize our findings and provide the remaining questions.

Pulmonary hypertension (PH) is defined as mean pulmonary arterial pressure at rest ≥25 mmHg. Pulmonary arterial hypertension (PAH) is classified as group 1 of PH and is a progressive and fatal disease of the pulmonary artery. The pathogenesis is sustained pulmonary vasoconstriction and pulmonary vascular remodeling, which cause progressive elevations in pulmonary vascular resistance and pulmonary arterial pressure. Elevated pulmonary arterial pressure leads to right heart failure and finally death. The pulmonary vascular remodeling is triggered by an increase in cytosolic Ca²⁺ concentration ([Ca²⁺]_cyt). [Ca²⁺]_cyt is regulated by the stimulation of vasoconstrictors and growth factors though their receptors and ion channels on the plasma membrane. It has been reported that the epidermal growth factor (EGF), fibroblast growth factor (FGF), insulin-like growth factor (IGF), vascular endothelial growth factor (VEGF), and platelet-derived growth factor (PDGF) are involved in the development of PAH. Upon binding of these growth factors with their specific receptor tyrosine kinases, their receptors activate cytosolic Ca²⁺ signaling and signal transduction cascades to induce cell proliferation, differentiation, and migration. Expressions of some growth factors and their receptors upregulate in PAH patients, which contributes to the formation of vascular remodeling and plexiform lesions in PAH. We have recently found that enhanced Ca²⁺-sensing receptor (CaSR) function is involved the development of PAH and CaSR expression is upregulated by PDGF in pulmonary arterial smooth muscle cells (PASMCs) from idiopathic PAH patients. This review will be discussed the physiological and pathological roles of growth factors in PAH.

In the brains of patients with Alzheimer’s disease, a decrease in phosphatidylinositol phosphate (PIP) requiring Cl⁻-ATPase activity was found. In cultured rat hippocampal neurons, pathophysiological concentrations of amyloid β proteins (Aβs≤10 nM) lowered PIP levels and Cl⁻-ATPase activity with an increase in intracellular Cl⁻ concentrations, resulting in Cl⁻-dependent enhancements in glutamate neurotoxicity and, ultimately, neuronal cell death. Pathophysiological concentrations of Aβs(0.1-10 nM) directly lowered phosphatidylinositol-4-kinase. Non-toxic peptide fragments of Aβ, such as Ile-Gly-Leu, recovered Aβ-induced inhibition of recombinant human phosphatidylinositol-4-kinase IIα (PI4KIIα) and the intrahippocampally administered Aβ-induced degeneration of hippocampal neurons and impairment of spatial memory in mice. Agents with the potential to block these neurotoxic mechanisms of Aβ were summarized herein as (1) Aβ antagonists, (2) substrates of PI4K, (3) PI4K product, (4) PI4K activators, and (5) GABAc receptor stimulants.

The modified Irwin’s method and functional observational battery (FOB）used in non-clinical studies for predicting side effects that may appear in the central nervous system (CNS）in clinical studies consist of mainly macroscopic observation and largely depend on the observer’s ability. Therefore, appropriate training for the observer and consistency of findings are extremely important, making it necessary for methods and judgment criteria to be standardized. In addition, because of concern for animal welfare as well as an increase in biopharmaceutical and anticancer drug development, there is increasing opportunity to incorporate safety pharmacological evaluation into general toxicity studies. While CNS evaluation can be incorporated into general toxicity studies relatively easily, studies need to be designed in such a way that reliable data can be obtained without reducing the ability to detect neurobehavioral abnormalities. It is therefore important to improve CNS evaluation techniques and to share these techniques with new observers in order to reliably detect the effects on the CNS during drug development.

Selexipag (Uptravi^® tablets) is a novel prostacyclin receptor (IP receptor) agonist designed and synthesized at Nippon Shinyaku Co., Ltd., and approved for the treatment of pulmonary arterial hypertension (PAH). Selexipag is converted to MRE-269 in vivo, and the plasma concentration of MRE-269 is maintained at a therapeutic level for a long time. MRE-269 has selective IP receptor agonist activity and exerts vasodilatory and anti-proliferative effects on pulmonary arterial smooth muscle cells. In a study to investigate its vasodilatory effect in isolated rat pulmonary arteries, MRE-269 showed potent vasodilatory effects not only in extralobar but also in small intralobar pulmonary arteries. In a Sugen 5416/hypoxia rat model of PAH, selexipag significantly improved pulmonary artery obstruction, decreased right ventricular systolic pressure, decreased right ventricular hypertrophy and improved survival rate. In a phase II clinical trial for treatment with PAH conducted in Europe, selexipag showed good tolerability with promising efficacy. In an open-label phase II study in 37 patients with PAH in Japan, selexipag significantly decreased pulmonary vascular resistance compared with baseline. In the GRIPHON (Prostacyclin (PGI₂) Receptor agonist In Pulmonary arterial HypertensiON) study in 1156 patients with PAH, the largest outcome study ever conducted in PAH, the selexipag treatment group showed a significant reduction in the risk of the primary composite endpoint of death or a complication related to PAH compared with placebo. Selexipag has been shown in clinical trials to prevent the progression of PAH, and is expected to contribute to the treatment of patients with PAH.

Roxadustat (Evrenzo^® tablet) is an oral hypoxia-inducible factor prolyl hydroxylase (HIF-PH) inhibitor. Roxadustat has been approved for the treatment of renal anemia in patients on dialysis in September 2019 in Japan. By inhibiting HIF-PH, roxadustat suppresses the degradation of HIF-α, a subunit of the heterodimeric transcription factor HIF, leading to its accumulation and activation of the HIF pathway. Similar to activation of the HIF pathway in response to hypoxia, the production of endogenous erythropoietin is increased and erythropoiesis is stimulated. Moreover, roxadustat stimulates erythropoiesis efficiently by improving iron bioavailability. The efficacy and mechanism of action of roxadustat have been detailed in non-clinical pharmacology studies. Rat models of anemia demonstrated efficacy of roxadustat in correcting anemia and changes in gene expression leading to increased iron bioavailability. Four phase 3 clinical studies in Japan clearly demonstrated the efficacy of roxadustat in patients with renal anemia on dialysis. Roxadustat showed an acceptable safety profile, and the incidences and types of adverse events and serious adverse events reported in the clinical studies were similar with those predicted to occur in these patient population. Since roxadustat is an oral drug, concerns present with erythropoiesis-stimulating agents (ESAs) such as the risk of infection to the medical staff due to accidental needle-stick, pain during ESA injection in patients and burden on patients to visit a hospital, can be avoided or reduced. In November 2020, roxadustat has also been approved for the treatment of renal anemia in patients not on dialysis (data not shown in this article).