Amine-free melanin-concentrating hormone receptor 1 antagonists: Novel non-basic 1-(2H-indazole-5-yl)pyridin-2(1H)-one derivatives and mitigation of mutagenicity in Ames test.

To develop non-basic melanin-concentrating hormone receptor 1 (MCHR1) antagonists with a high probability of target selectivity and therapeutic window, we explored neutral bicyclic motifs that could replace the previously reported imidazo[1,2-a]pyridine or 1H-benzimidazole motif. The results indicated that the binding affinity of a chemically neutral 2H-indazole derivative 8a with MCHR1 (hMCHR1: IC50=35nM) was comparable to that of the imidazopyridine and benzimidazole derivatives (1 and 2, respectively) reported so far. However, 8a was positive in the Ames test using TA1537 in S9- condition. Based on a putative intercalation of 8a with DNA, we introduced a sterically-hindering cyclopropyl group on the indazole ring to decrease planarity, which led to the discovery of 1-(2-cyclopropyl-3-methyl-2H-indazol-5-yl)-4-{[5-(trifluoromethyl)thiophen-3-yl]methoxy}pyridin-2(1H)-one 8l without mutagenicity in TA1537. Compound 8l exerted significant antiobesity effects in diet-induced obese F344 rats and exhibited promising safety profile.

Amine-free melanin-concentrating hormone receptor 1 antagonists: Novel 1-(1H-benzimidazol-6-yl)pyridin-2(1H)-one derivatives and design to avoid CYP3A4 time-dependent inhibition.

Melanin-concentrating hormone (MCH) is an attractive target for antiobesity agents, and numerous drug discovery programs are dedicated to finding small-molecule MCH receptor 1 (MCHR1) antagonists. We recently reported novel pyridine-2(1H)-ones as aliphatic amine-free MCHR1 antagonists that structurally featured an imidazo[1,2-a]pyridine-based bicyclic motif. To investigate imidazopyridine variants with lower basicity and less potential to inhibit cytochrome P450 3A4 (CYP3A4), we designed pyridine-2(1H)-ones bearing various less basic bicyclic motifs. Among these, a lead compound 6a bearing a 1H-benzimidazole motif showed comparable binding affinity to MCHR1 to the corresponding imidazopyridine derivative 1. Optimization of 6a afforded a series of potent thiophene derivatives (6q-u); however, most of these were found to cause time-dependent inhibition (TDI) of CYP3A4. As bioactivation of thiophenes to form sulfoxide or epoxide species was considered to be a major cause of CYP3A4 TDI, we introduced electron withdrawing groups on the thiophene and found that a CF3 group on the ring or a Cl adjacent to the sulfur atom helped prevent CYP3A4 TDI. Consequently, 4-[(5-chlorothiophen-2-yl)methoxy]-1-(2-cyclopropyl-1-methyl-1H-benzimidazol-6-yl)pyridin-2(1H)-one (6s) was identified as a potent MCHR1 antagonist without the risk of CYP3A4 TDI, which exhibited a promising safety profile including low CYP3A4 inhibition and exerted significant antiobesity effects in diet-induced obese F344 rats.

The GLP-1 receptor agonist exenatide improves recovery from spinal cord injury by inducing macrophage polarization toward the M2 phenotype.

Although a wide variety of mechanisms take part in the secondary injury phase of spinal cord injury (SCI), inflammation is the most important factor implicated in the sequelae after SCI. Being central to the inflammation reaction, macrophages and their polarization are a topic that has garnered wide interest in the studies of SCI secondary injury. The glucagon-like peptide 1 (GLP-1) receptor agonist exenatide has been shown to enhance the endoplasmic reticulum stress response and improve motor function recovery after spinal cord injury (SCI). Since exenatide has also been reported to induce the production of M2 cells in models of cerebral infarction and neurodegenerative diseases, this study was conducted to examine the effects of exenatide administration on the inflammation process that ensues after spinal cord injury. In a rat contusion model of spinal cord injury, the exenatide group received a subcutaneous injection of 10 µg exenatide immediately after injury while those in the control group received 1 mL of phosphate-buffered saline. Quantitative RT-PCR and immunohistochemical staining were used to evaluate the effects of exenatide administration on the macrophages infiltrating the injured spinal cord, especially with regard to macrophage M1 and M2 profiles. The changes in hind limb motor function were assessed based on Basso, Beattie, Bresnahan locomotor rating scale (BBB scale) scores. The improvement in BBB scale scores was significantly higher in the exenatide group from day 7 after injury and onwards. Quantitative RT-PCR revealed an increase in the expression of M2 markers and anti-inflammatory interleukins in the exenatide group that was accompanied by a decrease in the expression of M1 markers and inflammatory cytokines. Immunohistochemical staining showed no significant difference in M1 macrophage numbers between the two groups, but a significantly higher number of M2 macrophages was observed in the exenatide group on day 3 after injury. Our findings suggest that exenatide administration promoted the number of M2-phenotype macrophages after SCI, which may have led to the observed improvement in hind limb motor function in a rat model of SCI.

Administration of the GLP-1 receptor agonist exenatide in rats improves functional recovery after spinal cord injury by reducing endoplasmic reticulum stress.

After spinal cord injury (SCI), endoplasmic reticulum (ER) stress has been reported to be an integral part of the secondary injury process that causes apoptosis of glial cells, leading to remyelination failure. This report focuses on exenatide, a glucagon-like peptide-1 (GLP-1) receptor agonist widely used to treat diabetes, as a potential agent to improve functional outcome after SCI by improving the ER stress response. Exenatide administered subcutaneously immediately after injury and 7 days later in a rat model of moderate contusive SCI revealed significant improvement in hindlimb function without any hypoglycemia. Changes in the expression of glucose regulatory protein 78 (GRP78), an endoplasmic reticulum chaperone that protects against ER stress, and C/EBP homologous transcription factor protein (CHOP), a pro-apoptotic transcription factor in the apoptosis pathway were examined as indices of ER stress. We found that administration of exenatide after SCI suppressed CHOP while increasing GRP78 in the injured spinal cord, leading to a significant decrease in tissue damage and a significant increase in oligodendrocyte progenitor cell survival. This study suggests that administration of exenatide after SCI decreases ER stress and improves functional recovery without any apparent side-effects.

Hemostasis using a covered self-expandable metal stent for pseudoaneurysm bleeding from the perihilar bile duct.

Although there are many reports of hemostasis with covered self-expandable metal stent (CSEMS) for bleeding from the papilla of Vater and the intrapapillary and distal bile duct, there are rare reports of its use for hemostasis in the perihilar bile duct. We report the case of a patient undergoing supportive care for perihilar cholangiocarcinoma with acute cholecystitis after side-by-side placement of uncovered SEMS for perihilar bile duct obstruction. Percutaneous transhepatic gallbladder aspiration was performed upon admission, and hematemesis occurred the next day. Since computed tomography scanning showed a pseudoaneurysm in the right uncovered SEMS, hemostasis by interventional radiology (IVR) was performed thrice for massive bleeding; however, hemostasis could not be achieved. When endoscopic retrograde cholangiopancreatography was performed for scrutiny and treatment of melena and increased hepatobiliary enzyme, the endoscopic visual field could not be secured by bleeding, and changes in hemodynamics were observed; thus, IVR was required, but it was difficult to perform. Since bleeding from the right bile duct was expected, hemostasis was performed using CSEMS. This is the first report of hemostasis performed by placing a covered SEMS for bleeding from a pseudoaneurysm of the intrahepatic bile duct.

Primary endoscopic bile duct stone removal for severe acute cholangitis: a retrospective study.

INTRODUCTION: While the Tokyo Guidelines 2018 suggest primary stone removal for mild to moderate cholangitis, a guideline for severe acute cholangitis is not mentioned. We, therefore, investigated the clinical outcomes of patients with severe acute cholangitis to confirm the usefulness and safety of primary stone removal. METHOD: This study included 104 severe acute cholangitis patients without gallstone pancreatitis diagnosed at our institution between January 2014 and December 2020. Patients with percutaneous transhepatic biliary drainage as the primary drainage, bile duct stenosis, and endoscopically unidentified bile duct stones were excluded from this study. The clinical results of 14 patients with primary stone removal (primary group) and 23 patients with elective stone removal (elective group) were then retrospectively examined (excluding abnormal values due to underlying diseases). RESULTS: Upon comparing the patient characteristics between groups, the elective group had significantly higher cardiovascular dysfunction (57% vs 7%; p = 0.004), septic shock (39% vs 0%; p = 0.006), disseminated intravascular coagulation treatment (57% vs 14%; p = 0.016), and positive blood cultures (91% vs 43%; p = 0.006) than those in the primary group. Endoscopic sphincterotomy for naïve papilla (90% vs 21%; p = 0.01) and endoscopic nasobiliary drainage (50% vs 9%; p = 0.014) were higher in the primary group, while endoscopic biliary stenting (7% vs 87%; p < 0.001) was lower than that in the elective group. DISCUSSION: There were no significant differences in adverse events or complete stone removal rates between the two groups. In the primary group, the period from the first endoscopic retrograde cholangiopancreatography to stone removal (0 days vs 12 days; p < 0.001) and hospitalization period (12 days vs 26 days; p = 0.012) were significantly shorter and the hospitalization cost ($7731 vs $18758; p < 0.001) was significantly lower than those in the elective group. CONCLUSION: If patients are appropriately selected, bile duct stones may be safely removed for the treatment of severe acute cholangitis.

Clinical Features and Outcomes of Bilateral Atypical Femoral Fractures.

Bilateral atypical femoral fractures (AFFs) are relatively rare. In this report, we retrospectively researched clinical features and outcomes of bilateral AFFs treated at our institution. We previously treated 4 patients (8 limbs) with intramedullary nailing for complete AFFs (6 limbs) and incomplete AFFs (2 limbs). The mean age at the first operation was 53.3 years, and all patients were female. Of the 4 patients, two had breast cancer, and another two had systemic lupus erythematosus. Three of them received bisphosphonates, and 2 received denosumab, proton pump inhibitor, or glucocorticoid therapy. Only 2 of 6 cases of incomplete AFFs had prodromal pain before progressing to complete fracture. The mean interval from the first surgery to contralateral fracture or prophylactic surgery was 16 months. Radiographically, complete bone union was achieved in 6 limbs. However, a small gap at the lateral cortex of fracture site remained in 2 limbs. Finally, all of the patients were pain-free and able to walk without a cane. It is absolutely necessary to confirm contralateral femoral conditions; however, prediction of progression to complete fracture based solely on prodromal pain was difficult. Therefore, we should advise patients about the danger of progression to complete AFFs even if they are asymptomatic, and a prophylactic surgery should be performed after obtaining informed consent.

Intra-hepatic arterial administration with miriplatin suspended in an oily lymphographic agent inhibits the growth of human hepatoma cells orthotopically implanted in nude rats.

Miriplatin is a lipophilic platinum complex which contains myristates as leaving groups and diaminocyclohexane as a carrier ligand. In order to examine in vivo the antitumor activities of miriplatin suspended in an oily lymphographic agent (Lipiodol Ultra-Fluide, LPD) against human hepatocellular carcinoma (HCC) after the intra-hepatic arterial administration, we have developed a novel orthotopic model of HCC in which the human hepatoma cell line Li-7 was successively implanted and maintained in the liver of nude rats. Li-7 tumors established in nude rat livers displayed a trabecular structure similar to their original morphology, and were exclusively supplied by the hepatic artery, suggesting that they exhibited in part the conditions of human HCC. Miriplatin suspended in LPD (miriplatin/LPD) administered into the hepatic artery of this model dose-dependently inhibited the growth of Li-7 tumors without markedly enhancing body weight loss and caused a significant reduction in the growth rate at a dose of 400 microg/head compared to LPD alone. In addition, at the therapeutic dose, miriplatin/LPD as well as cisplatin suspended in LPD (400 microg/head) was shown to be more active than zinostatin stimalamer suspended in LPD (20 microg/head) against Li-7 tumors after a single intra-hepatic arterial administration. These results suggest miriplatin to be a suitable candidate for use in transarterial chemoembolization.

Hematological aspects of a novel 9-aminoanthracycline, amrubicin.

Our previous study showed that the myelosuppression induced by a single-bolus intravenous injection of amrubicin into mice was more severe, even at half the maximum tolerated dose (MTD), than that induced by adriamycin (ADR) at the MTD, but that the recovery was more rapid than that after ADR. The present study shows that the administration of amrubicin significantly decreased the number of colony-forming units of granulocytes and monocytes (CFU-GM) from day 1, but that the CFU-GM numbers recovered by day 3. In contrast, the administration of ADR induced a continuous decrease in the numbers of CFU-GM until day 10. The early myelosuppression and rapid recovery of white blood cells (WBC) induced by amrubicin may depend upon the early decrease in the number of CFU-GM and the rapid recovery of CFU-GM. These data suggest that the toxic effects on the peripheral blood and bone marrow induced by amrubicin are more reversible and more controllable than those induced by ADR.

Intra-hepatic arterial administration with miriplatin suspended in an oily lymphographic agent inhibits the growth of tumors implanted in rat livers by inducing platinum-DNA adducts to form and massive apoptosis.

BACKGROUND: Miriplatin (formerly SM-11355), a novel lipophilic platinum complex developed to treat hepatocellular carcinoma, is administered into the hepatic artery using an oily lymphographic agent (Lipiodol Ultra-Fluide) as a carrier. We clarified the usefulness of miriplatin as an agent for transarterial chemoembolization. METHODS: Platinum compounds released from miriplatin into serum, medium and Earle's balanced salt solution were examined. Then, miriplatin and cisplatin were administered to rats bearing hepatoma AH109A tumors in livers. Platinum concentrations in tissues and DNA were assessed. RESULTS: Miriplatin showed a more sustained release than cisplatin. Dichloro[(1R, 2R)-1, 2-cyclohexane diamine-N, N']platinum, the most abundant platinum compound released from miriplatin, was as effective as cisplatin in inhibiting the growth of cells. Miriplatin was selectively disposed of in tumors, maintained in tumors longer than cisplatin and caused apparent tumor regression inducing platinum-DNA adducts to form and massive apoptosis. CONCLUSION: Miriplatin appears to be a suitable chemotherapeutic agent for transarterial chemoembolization.

Amrubicin, a novel 9-aminoanthracycline, enhances the antitumor activity of chemotherapeutic agents against human cancer cells in vitro and in vivo.

Amrubicin, a completely synthetic 9-aminoanthracycline derivative, is an active agent in the treatment of untreated extensive disease-small-cell lung cancer and advanced non-small-cell lung cancer. Amrubicin administered intravenously at 25 mg/kg substantially prevented the growth of five of six human lung cancer xenografts established in athymic nude mice, confirming that amrubicin as a single agent was active in human lung tumors. To survey which antitumor agent available for clinical use produces a synergistic interaction with amrubicin, we examined the effects in combinations with amrubicinol, an active metabolite of amrubicin, of several chemotherapeutic agents in vitro using five human cancer cell lines using the combination index (CI) method of Chou and Talalay. Synergistic effects were obtained on the simultaneous use of amrubicinol with cisplatin, irinotecan, gefitinib and trastuzumab, with CI values after 3 days of exposure being <1. Additive effect was observed with the combination containing vinorelbine with CI values indistinguishable from 1, while the combination of amrubicinol with gemcitabine was antagonistic. All combinations tested in vivo were well tolerated. The combinations of cisplatin, irinotecan, vinorelbine, trastuzumab, tegafur/uracil, and to a lesser extent, gemcitabine with amrubicin caused significant growth inhibition of human tumor xenografts without pronouncedly enhancing body weight loss, compared with treatment using amrubicin alone at the maximum tolerated dose. Growth inhibition of tumors by gefitinib was not antagonized by amrubicin. These results suggest that amrubicin appears to be a possible candidate for combined use with cisplatin, irinotecan, vinorelbine, gemcitabine, tegafur/uracil or trastuzumab.

Amrubicin induces apoptosis in human tumor cells mediated by the activation of caspase-3/7 preceding a loss of mitochondrial membrane potential.

Amrubicin, a completely synthetic 9-aminoanthracycline derivative, inhibits cell growth by stabilizing a topoisomerase II-DNA complex. This study was designed to examine the apoptosis induced in human leukemia U937 cells by amrubicin and its active metabolite amrubicinol. Amrubicin, amrubicinol and other antitumor agents, such as daunorubicin and etoposide, induced typical apoptosis with characteristic nuclear morphological change and DNA fragmentation. Measuring the population of sub-G(1) phase cells, it was found that under conditions where cell growth was inhibited by either amrubicin or amrubicinol, U937 cells underwent apoptotic cell death in a dose-dependent manner accompanied by an arrest of the cell cycle at G(2)/M. Furthermore, amrubicin- and amrubicinol-induced apoptosis was mediated by the activation of caspase-3/7, but not caspase-1, preceding a loss of mitochondrial membrane potential. These results indicate that both a reduction in mitochondrial membrane potential and the activation of caspase-3/7 are key events in the apoptosis induced by amrubicin and amrubicinol as well as the other antitumor agents. In addition, studies with oligomycin suggested that the apoptosis induced by amrubicin and amrubicinol involved substantially different pathways from that triggered by daunorubicin and etoposide. Oligomycin blocked the etoposide-induced increase in the number of sub-G(1) phase cells without preventing the activation of caspase-3/7, and had no inhibitory effect on the expansion of the sub-G(1) population in daunorubicin-treated cells, whereas apoptosis-related changes caused by amrubicin and amrubicinol were suppressed in the presence of oligomycin.

[Profile of the anti-tumor effects of amrubicin, a completely synthetic anthracycline].

Amrubicin is a completely synthetic anthracycline derivative. In contrast, however, the anthracyclines used clinically thus far have been produced by fermentation or semisynthesis. Amrubicin is structurally distinguishable from other anthracyclines by the amino group at the 9-position and its unique sugar moiety. Amrubicinol, the C-13 hydroxy- metabolite of amrubicin, is associated with a 5 to 200 times greater cytotoxicity than amrubicin. Amrubicin exhibited superior in vivo antitumor activity to doxorubicin in the human tumor xenograft model. Using this model, the level of amrubicinol (active metabolite) was shown to be higher than that of doxorubicin in tumor tissues, but lower in normal tissues. These results suggest potent therapeutic activity for amrubicin because of the selective distribution of its highly active metabolite, amrubicinol, in tumors. These anti-tumor effects of amrubicin are considered to be induced by DNA topoisomeraseII inhibition. In clinical studies, amrubicin has demonstrated potent single agent activity as compared to a standard regimen in untreated patients with extensive small cell lung cancer. Its major toxicity was myelosuppression (especially neutropenia).