Antiviral Activity and Resistance Profile of the Novel HIV-1 Non-Catalytic Site Integrase Inhibitor JTP-0157602.

HIV-1 integrase (IN) is an essential enzyme for viral replication. Non-catalytic site integrase inhibitors (NCINIs) are allosteric HIV-1 IN inhibitors and a potential new class of antiretrovirals. In this report, we identified a novel NCINI, JTP-0157602, with an original scaffold. JTP-0157602 exhibited potent antiviral activity against HIV-1 and showed a serum-shifted 90% effective concentration (EC90) of 138 nM, which is comparable to those of the FDA-approved IN strand transfer inhibitors (INSTIs). This compound was fully potent against a wide range of recombinant viruses with IN polymorphisms, including amino acids 124/125, a hot spot of IN polymorphisms. In addition, JTP-0157602 retained potent antiviral activity against a broad panel of recombinant viruses with INSTI-related resistance mutations, including multiple substitutions that emerged in clinical studies of INSTIs. Resistance selection experiments of JTP-0157602 led to the emergence of A128T and T174I mutations, which are located at the lens epithelium-derived growth factor/p75 binding pocket of IN. JTP-0157602 inhibited HIV-1 replication mainly during the late phase of the replication cycle, and HIV-1 virions produced by reactivation from HIV-1 latently infected Jurkat cells in the presence of JTP-0157602 were noninfectious. These results suggest that JTP-0157602 and analog compounds can be used to treat HIV-1 infectious diseases. IMPORTANCE Non-catalytic site integrase inhibitors (NCINIs) are allosteric HIV-1 integrase (IN) inhibitors that bind to the lens epithelium-derived growth factor (LEDGF)/p75 binding pocket of IN. NCINIs are expected to be a new class of anti-HIV-1 agents. In this study, we present a novel NCINI, JTP-0157602, which has potent activity against a broad range of HIV-1 strains with IN polymorphisms. Furthermore, JTP-0157602 shows strong antiviral activity against IN strand transfer inhibitor-resistant mutations, suggesting that JTP-0157602 and its analogs are potential agents for treating HIV-1 infections. Structural modeling indicated that JTP-0157602 binds to the LEDGF/p75 binding pocket of IN, and the results of in vitro resistance induction revealed the JTP-0157602 resistance mechanism of HIV-1. These data shed light on developing novel NCINIs that exhibit potent activity against HIV-1 with broad IN polymorphisms and multidrug-resistant HIV-1 variants.

Fluorine Modifications Contribute to Potent Antiviral Activity against Highly Drug-Resistant HIV-1 and Favorable Blood-Brain Barrier Penetration Property of Novel Central Nervous System-Targeting HIV-1 Protease Inhibitors In Vitro.

To date, there are no specific treatment regimens for HIV-1-related central nervous system (CNS) complications, such as HIV-1-associated neurocognitive disorders (HAND). Here, we report that two newly generated CNS-targeting HIV-1 protease (PR) inhibitors (PIs), GRL-08513 and GRL-08613, which have a P1-3,5-bis-fluorophenyl or P1-para-monofluorophenyl ring and P2-tetrahydropyrano-tetrahydrofuran (Tp-THF) with a sulfonamide isostere, are potent against wild-type HIV-1 strains and multiple clinically isolated HIV-1 strains (50% effective concentration [EC50]: 0.0001 to ∼0.0032 µM). As assessed with HIV-1 variants that had been selected in vitro to propagate at a 5 µM concentration of each HIV-1 PI (atazanavir, lopinavir, or amprenavir), GRL-08513 and GRL-08613 efficiently inhibited the replication of these highly PI-resistant variants (EC50: 0.003 to ∼0.006 µM). GRL-08513 and GRL-08613 also maintained their antiviral activities against HIV-2ROD as well as severely multidrug-resistant clinical HIV-1 variants. Additionally, when we assessed with the in vitro blood-brain barrier (BBB) reconstruction system, GRL-08513 and GRL-08613 showed the most promising properties of CNS penetration among the evaluated compounds, including the majority of FDA-approved combination antiretroviral therapy (cART) drugs. In the crystallographic analysis of compound-PR complexes, it was demonstrated that the Tp-THF rings at the P2 moiety of GRL-08513 and GRL-08613 form robust hydrogen bond interactions with the active site of HIV-1 PR. Furthermore, both the P1-3,5-bis-fluorophenyl- and P1-para-monofluorophenyl rings sustain greater contact surfaces and form stronger van der Waals interactions with PR than is the case with darunavir-PR complex. Taken together, these results strongly suggest that GRL-08513 and GRL-08613 have favorable features for patients infected with wild-type/multidrug-resistant HIV-1 strains and might serve as candidates for a preventive and/or therapeutic agent for HAND and other CNS complications.

Disseminated tuberculosis with paradoxical reactions caused by a Mycobacterium tuberculosis strain belonging to the Indo-Oceanic lineage: An imported case in Japan.

Tuberculosis remains a major public health concern. Millions of tuberculosis cases and associated deaths have been reported worldwide. The Indo-Oceanic lineage Mycobacterium tuberculosis is common in Southeast Asia and causes extrapulmonary lesions. Only a few case studies on this lineage with genetic analysis using whole-genome sequencing have been reported in the literature. We present a case of disseminated tuberculosis, characterized by a variety of extrapulmonary lesions and paradoxical reactions, caused by the Indo-Oceanic lineage M. tuberculosis in a woman in Myanmar. A 22-year-old Burmese woman had arthritis in the right knee, with unknown aetiology, and was referred to our hospital. Computed tomography of the trunk revealed multiple nodular shadows in both lungs; swollen mediastinal lymph nodes; and small, low-density areas in the spleen. M. tuberculosis was detected in the sputum sample, joint aspirate, subcutaneous tumor, and exudate. She experienced a variety of paradoxical reactions together with aggressive tuberculosis dissemination in all areas of the body. Whole-genome sequencing of the DNA of MTB obtained from sputum and the right cervical subcutaneous abscess confirmed the Indo-Oceanic lineage of M. tuberculosis, the predominant strain in Myanmar. The Indo-Oceanic lineage M. tuberculosis causes disseminated tuberculosis all over the body including the periungual region. When patients show unusual symptoms, physicians should consider the introduction of new strains from foreign countries. Genetic analyses of the strains are recommended to define and confirm the lineages.

Analysis of perioperative glucose metabolism using an artificial pancreas.

Hyperglycemia associated with insulin resistance is common in surgical patients with and without diabetes and is associated with poor surgical outcomes. Several studies have recently shown that a closed-loop blood glucose monitoring system in the form of an artificial pancreas is safe and effective for surgical patients. In this study, we analyzed the risk factors for insulin resistance in patients using an artificial pancreas. We investigated 109 patients who underwent surgical management by an artificial pancreas for 24 hours from the start of surgery during either major hepatectomy (MH), defined as resection of more than two liver segments, or pancreaticoduodenectomy (PD). The target glucose range was from 80 to 110 mg/dL using an artificial pancreas. We analyzed the risk factors for and predictors of a high insulin dose, including sarcopenia markers, according to the median 24-hour total insulin infusion. The median total insulin dose and glycemic control rate (GCR), which is the rate of achieving the target blood glucose range, per 24 hours were 78.0 IU and 30.4% in the MH group and 82.6 IU and 23.5% in the PD group, respectively. The muscle volume was the only independent factor in the high-dose subgroup, and the GCR was significantly lower in the high-dose subgroup despite a high insulin dose in both the MH and PD groups. The results of this study suggest that preoperative sarcopenia is closely associated with insulin resistance in the perioperative period. Clinicians must effectively manage sarcopenia, which may result in improved perioperative glycemic control and reduced postoperative complications.

[Laparoscopy Assisted Subtotal Gastrectomy(Billrothâ Reconstruction)for Gastric Cancer in the Upper Stomach-A Report of Three Cases].

We experienced 3 cases of upper gastric cancer who underwent BillrothⅠ reconstruction in laparoscopy assisted subtotal gastrectomy. Two cases were female and 1 was male. The postoperative course was uneventful in all cases without heartburn, and the surgical margin was negative. The body weight loss rate was 5.8-12.6%, and the short-term results were relatively acceptable. Although the number of cases in this study was small, reconstruction with BillrothⅠ/delta-shaped anastomosis after laparoscopy assisted subtotal gastrectomy were considered to be useful.

Freshwater mussels (Bivalvia: Unionidae) from the rising sun (Far East Asia): phylogeny, systematics, and distribution.

Freshwater mussels (Bivalvia: Unionidae) is a diverse family with around 700 species being widespread in the Northern Hemisphere and Africa. These animals fulfill key ecological functions and provide important services to humans. Unfortunately, populations have declined dramatically over the last century, rendering Unionidae one of the world's most imperiled taxonomic groups. In Far East Asia (comprising Japan, Korea, and Eastern Russia), conservation actions have been hindered by a lack of basic information on the number, identity, distribution and phylogenetic relationships of species. Available knowledge is restricted to studies on national and sub-national levels. The present study aims to resolve the diversity, biogeography and evolutionary relationships of the Far East Asian Unionidae in a globally comprehensive phylogenetic and systematic context. We reassessed the systematics of all Unionidae species in the region, including newly collected specimens from across Japan, South Korea, and Russia, based on molecular (including molecular species delineation and a COI + 28S phylogeny) and comparative morphological analyses. Biogeographical patterns were then assessed based on available species distribution data from the authors and previous reference works. We revealed that Unionidae species richness in Far East Asia is 30% higher than previously assumed, counting 43 species (41 native + 2 alien) within two Unionidae subfamilies, the Unioninae (32 + 1) and Gonideinae (9 + 1). Four of these species are new to science, i.e. Beringiana gosannensissp. nov., Beringiana fukuharaisp. nov., Buldowskia kamiyaisp. nov., and Koreosolenaia sitgyensisgen. & sp. nov. We also propose a replacement name for Nodularia sinulata, i.e. Nodularia breviconchanom. nov. and describe a new tribe (Middendorffinaiini tribe nov.) within the Unioninae subfamily. Biogeographical patterns indicate that this fauna is related to that from China south to Vietnam until the Mekong River basin. The Japanese islands of Honshu, Shikoku, Kyushu, Hokkaido, and the Korean Peninsula were identified as areas of particularly high conservation value, owing to high rates of endemism, diversity and habitat loss. The genetically unique species within the genera Amuranodonta, Obovalis, Koreosolenaiagen. nov., and Middendorffinaia are of high conservation concern.

[A Case of a Huge Gastrointestinal Stromal Tumor with Extraluminal Growth Arising in the Small Intestine Diagnosed as an Ovarian Tumor].

We report a case of a gastrointenstinal stromal tumor(GIST)of the small intestine with extraluminal growth that was difficult to distinguish from an ovarian tumor. A 73-year-old woman presented to a nearby hospital for lower abdominal pain. A computed tomography(CT)scan showed a 17 cm ovarian tumor in the pelvis, and she was referred to the gynecology department of our hospital. Following examinations(enhanced CT and magnetic resonance imaging), she was referred to our department in suspicion of a small intestinal GIST in which the superior mesenteric artery/vein was the feeding blood vessel, and intraperitoneal tumor resection was performed. A large cystic tumor occupied the abdominal cavity and was in contact with the small intestinal wall. As the tumor was not in contact with the uterus or bilateral adnexa, only partial resection of the small intestine was performed. Histopathological examination showed c-kit positivity and she was diagnosed with small intestinal GIST; as a result, a course of imatinib was started.

Novel p97/VCP inhibitor induces endoplasmic reticulum stress and apoptosis in both bortezomib-sensitive and -resistant multiple myeloma cells.

p97/VCP is an endoplasmic reticulum (ER)-associated protein that belongs to the AAA (ATPases associated with diverse cellular activities) ATPase family. It has a variety of cellular functions including ER-associated protein degradation, autophagy, and aggresome formation. Recent studies have shown emerging roles of p97/VCP and its potential as a therapeutic target in several cancer subtypes including multiple myeloma (MM). We conducted a cell-based compound screen to exploit novel small compounds that have cytotoxic activity in myeloma cells. Among approximately 2000 compounds, OSSL_325096 showed relatively strong antiproliferative activity in MM cell lines (IC50 , 100-500 nmol/L). OSSL_325096 induced apoptosis in myeloma cell lines, including a bortezomib-resistant cell line and primary myeloma cells purified from patients. Accumulation of poly-ubiquitinated proteins, PERK, CHOP, and IREα, was observed in MM cell lines treated with OSSL_325096, suggesting that it induces ER stress in MM cells. OSSL_325096 has a similar chemical structure to DBeQ, a known p97/VCP inhibitor. Knockdown of the gene encoding p97/VCP induced apoptosis in myeloma cells, accompanied by accumulation of poly-ubiquitinated protein. IC50 of OSSL_325096 to myeloma cell lines were found to be lower (0.1-0.8 µmol/L) than those of DBeQ (2-5 µmol/L). In silico protein-drug-binding simulation suggested possible binding of OSSL_325096 to the ATP binding site in the D2 domain of p97/VCP. In cell-free ATPase assays, OSSL_325096 showed dose-dependent inhibition of p97/VCP ATPase activity. Finally, OSSL_325096 inhibited the growth of subcutaneous myeloma cell tumors in vivo. The present data suggest that OSSL_325096 exerts anti-myeloma activity, at least in part through p97/VCP inhibition.

Halogen Bond Interactions of Novel HIV-1 Protease Inhibitors (PI) (GRL-001-15 and GRL-003-15) with the Flap of Protease Are Critical for Their Potent Activity against Wild-Type HIV-1 and Multi-PI-Resistant Variants.

We generated two novel nonpeptidic HIV-1 protease inhibitors (PIs), GRL-001-15 and GRL-003-15, which contain unique crown-like tetrahydropyranofuran (Crn-THF) and P2'-cyclopropyl-aminobenzothiazole (Cp-Abt) moieties as P2 and P2' ligands, respectively. GRL-001-15 and GRL-003-15 have meta-monofluorophenyl and para-monofluorophenyl at the P1 site, respectively, exert highly potent activity against wild-type HIV-1 with 50% effective concentrations (EC50s) of 57 and 50 pM, respectively, and have favorable cytotoxicity profiles with 50% cytotoxic concentrations (CC50s) of 38 and 11 µM, respectively. The activity of GRL-001-15 against multi-PI-resistant HIV-1 variants was generally greater than that of GRL-003-15. The EC50 of GRL-001-15 against an HIV-1 variant that was highly resistant to multiple PIs, including darunavir (DRV) (HIV-1DRVRP30), was 0.17 nM, and that of GRL-003-15 was 3.3 nM, while DRV was much less active, with an EC50 of 216 nM. The emergence of HIV-1 variants resistant to GRL-001-15 and GRL-003-15 was significantly delayed compared to that of variants resistant to selected PIs, including DRV. Structural analyses of wild-type protease (PRWT) complexed with the novel PIs revealed that GRL-001-15's meta-fluorine atom forms halogen bond interactions (2.9 and 3.0 Å) with Gly49 and Ile50, respectively, of the protease flap region and with Pro81' (2.7 and 3.2 Å), which is located close to the protease active site, and that two fluorine atoms of GRL-142-13 form multiple halogen bond interactions with Gly49, Ile50, Pro81', Ile82', and Arg8'. In contrast, GRL-003-15 forms halogen bond interactions with Pro81' alone, suggesting that the reduced antiviral activity of GRL-003-15 is due to the loss of the interactions with the flap region.

Novel Protease Inhibitors Containing C-5-Modified bis-Tetrahydrofuranylurethane and Aminobenzothiazole as P2 and P2' Ligands That Exert Potent Antiviral Activity against Highly Multidrug-Resistant HIV-1 with a High Genetic Barrier against the Emergence of Drug Resistance.

Combination antiretroviral therapy has achieved dramatic reductions in the mortality and morbidity in people with HIV-1 infection. Darunavir (DRV) represents a most efficacious and well-tolerated protease inhibitor (PI) with a high genetic barrier to the emergence of drug-resistant HIV-1. However, highly DRV-resistant variants have been reported in patients receiving long-term DRV-containing regimens. Here, we report three novel HIV-1 PIs (GRL-057-14, GRL-058-14, and GRL-059-14), all of which contain a P2-amino-substituted-bis-tetrahydrofuranylurethane (bis-THF) and a P2'-cyclopropyl-amino-benzothiazole (Cp-Abt). These PIs not only potently inhibit the replication of wild-type HIV-1 (50% effective concentration [EC50], 0.22 nM to 10.4 nM) but also inhibit multi-PI-resistant HIV-1 variants, including highly DRV-resistant HIVDRVRP51 (EC50, 1.6 nM to 30.7 nM). The emergence of HIV-1 variants resistant to the three compounds was much delayed in selection experiments compared to resistance to DRV, using a mixture of 11 highly multi-PI-resistant HIV-1 isolates as a starting HIV-1 population. GRL-057-14 showed the most potent anti-HIV-1 activity and greatest thermal stability with wild-type protease, and potently inhibited HIV-1 protease's proteolytic activity (Ki value, 0.10 nM) among the three PIs. Structural models indicate that the C-5-isopropylamino-bis-THF moiety of GRL-057-14 forms additional polar interactions with the active site of HIV-1 protease. Moreover, GRL-057-14's P1-bis-fluoro-methylbenzene forms strong hydrogen bonding and effective van der Waals interactions. The present data suggest that the combination of C-5-aminoalkyl-bis-THF, P1-bis-fluoro-methylbenzene, and P2'-Cp-Abt confers highly potent activity against wild-type and multi-PI-resistant HIV strains and warrant further development of the three PIs, in particular, that of GRL-057-14, as potential therapeutic for HIV-1 infection and AIDS.

Novel Central Nervous System (CNS)-Targeting Protease Inhibitors for Drug-Resistant HIV Infection and HIV-Associated CNS Complications.

There is currently no specific therapeutics for the HIV-1-related central nervous system (CNS) complications. Here we report that three newly designed CNS-targeting HIV-1 protease inhibitors (PIs), GRL-083-13, GRL-084-13, and GRL-087-13, which contain a P1-3,5-bis-fluorophenyl or P1-para-monofluorophenyl ring, and P2-bis-tetrahydrofuran (bis-THF) or P2-tetrahydropyrano-tetrahydrofuran (Tp-THF), with a sulfonamide isostere, are highly active against wild-type HIV-1 strains and primary clinical isolates (50% effective concentration [EC50], 0.0002 to ∼0.003 µM), with minimal cytotoxicity. These CNS-targeting PIs efficiently suppressed the replication of HIV-1 variants (EC50, 0.002 to ∼0.047 µM) that had been selected to propagate at high concentrations of conventional HIV-1 PIs. Such CNS-targeting PIs maintained their antiviral activity against HIV-2ROD as well as multidrug-resistant clinical HIV-1 variants isolated from AIDS patients who no longer responded to existing antiviral regimens after long-term therapy. Long-term drug selection experiments revealed that the emergence of resistant-HIV-1 against these CNS-targeting PIs was substantially delayed. In addition, the CNS-targeting PIs showed the most favorable CNS penetration properties among the tested compounds, including various FDA-approved anti-HIV-1 drugs, as assessed with the in vitro blood-brain barrier reconstruction system. Crystallographic analysis demonstrated that the bicyclic rings at the P2 moiety of the CNS-targeting PIs form strong hydrogen-bond interactions with HIV-1 protease (PR) active site. Moreover, both the P1-3,5-bis-fluorophenyl and P1-para-monofluorophenyl rings sustain greater van der Waals contacts with PR than in the case of darunavir (DRV). The data suggest that the present CNS-targeting PIs have desirable features for treating patients infected with wild-type and/or multidrug-resistant HIV-1 strains and might serve as promising preventive and/or therapeutic candidates for HIV-1-associated neurocognitive disorders (HAND) and other CNS complications.

Maximum Walking Speed at Discharge Could Be a Prognostic Factor for Vascular Events in Patients With Mild Stroke: A Cohort Study.

OBJECTIVE: To identify the prognostic value of physical activity-related factors as well as known vascular risk factors for vascular events in mild ischemic stroke (MIS). DESIGN: Single-center prospective cohort study. SETTING: University hospital. PARTICIPANTS: Consecutive patients (N=255) (175 men, median age 70.0y) with acute ischemic stroke and transient ischemic attack (TIA) with modified Rankin scale scores ranging from 0 to 2 were enrolled in this study. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Enrolled patients were followed up for composite vascular events as primary outcomes up to 3 years postdischarge. Primary outcomes included stroke and cardiovascular death, hospitalization due to stroke or TIA recurrence, cardiovascular disease, and peripheral artery disease. During hospitalization, known vascular risk factors such as previous history of vascular events, stroke subtype, white matter lesions, and ankle-brachial index were assessed. Moreover, at the time of discharge, physical activity-related factors such as maximum walking speed (MWS), handgrip strength, knee extensor isometric muscle strength, anxiety, and depression were assessed as potential predictors. RESULTS: The Kaplan-Meier estimates of cumulative risk of composite vascular events at 1, 2, and 3 years were 9.6%, 14.4%, and 15.2%, respectively. After multivariate analysis, cerebral white matter lesions of periventricular hyperintensity (PVH) (grade=3; hazard ratio: 2.904; 95% confidence interval: 1.160 to 7.266; P=.023) and MWS (<1.45m/s; hazard ratio: 2.232; 95% confidence interval: 1.010 to 4.933; P=.047) were identified as significant independent predictors of composite vascular events. CONCLUSIONS: The results of this study indicate that MWS could be an independent prognostic factor for composite vascular events in MIS.

GRL-079, a Novel HIV-1 Protease Inhibitor, Is Extremely Potent against Multidrug-Resistant HIV-1 Variants and Has a High Genetic Barrier against the Emergence of Resistant Variants.

We identified four novel nonpeptidic human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs), GRL-078, -079, -077, and -058, containing an alkylamine at the C-5 position of P2 tetrahydropyrano-tetrahydrofuran (Tp-THF) and a P2' cyclopropyl (Cp) (or isopropyl)-aminobenzothiazole (Abt) moiety. Their 50% effective concentrations (EC50s) were 2.5 to 30 nM against wild-type HIV-1NL4-3, 0.3 to 6.7 nM against HIV-2EHO, and 0.9 to 90 nM against laboratory-selected PI-resistant HIV-1 and clinical HIV-1 variants resistant to multiple FDA-approved PIs (HIVMDR). GRL-078, -079, -077, and -058 also effectively blocked the replication of HIV-1 variants highly resistant to darunavir (DRV) (HIVDRVrp51), with EC50s of 38, 62, 61, and 90 nM, respectively, while four FDA-approved PIs examined (amprenavir, atazanavir, lopinavir [LPV], and DRV) had virtually no activity (EC50s of >1,000 nM) against HIVDRVrp51 Structurally, GRL-078, -079, and -058 form strong hydrogen bond interactions between Tp-THF modified at C-5 and Asp29/Asp30/Gly48 of wild-type protease, while the P2' Cp-Abt group forms strong hydrogen bonds with Asp30'. The Tp-THF and Cp-Abt moieties also have good nonpolar interactions with protease residues located in the flap region. For selection with LPV and DRV by use of a mixture of 11 HIVMDR strains (HIV11MIX), HIV11MIX became highly resistant to LPV and DRV over 13 to 32 and 32 to 41 weeks, respectively. However, for selection with GRL-079 and GRL-058, HIV11MIX failed to replicate at >0.08 µM and >0.2 µM, respectively. Thermal stability results supported the highly favorable anti-HIV-1 potency of GRL-079 as well as other PIs. The present data strongly suggest that the P2 Tp-THF group modified at C-5 and the P2' Abt group contribute to the potent anti-HIV-1 profiles of the four PIs against HIV-1NL4-3 and a wide spectrum of HIVMDR strains.

Phosphodiesterase III inhibitor attenuates rat sinusoidal obstruction syndrome through inhibition of platelet aggregation in Disse's space.

BACKGROUND AND AIM: Sinusoidal obstruction syndrome (SOS) is a serious drug-induced liver injury. However, the pathophysiology of the disease remains unclear. This study investigated the effects of cilostazol (CZ), a phosphodiesterase III inhibitor, in a monocrotaline (MCT)-induced rat model of SOS. METHODS: Male Wistar rats were administrated MCT to induce SOS. Rats were divided into control, MCT, and MCT + CZ groups. In the MCT + CZ group, CZ was administered at 48 h, 24 h, and 30 min prior to and 8 h and 24 h after MCT administration. The MCT group was treated with water instead of CZ. At 48 h after MCT administration, blood and liver samples were collected to assess biochemistry and liver histology. Expression of rat endothelial cell antigen, CD34, CD41, P-selectin, and caspase-3 in the liver were analyzed. Plasminogen activator inhibitor-1 (PAI-1) in hepatocytes was analyzed using western blotting and polymerase chain reaction. RESULTS: In the MCT group, macroscopic findings showed a dark-red liver surface. Histological findings showed sinusoidal dilatation, coagulative necrosis of hepatocytes, and endothelial damage of the central vein. These changes were attenuated in the MCT + CZ group. Elevated serum transaminase and decreased platelet counts were observed in the MCT + CZ group compared with those in the MCT group. Treatment with CZ reduced MCT-induced damage to the liver sinusoidal endothelial cells, inhibited extravasated platelet aggregation, and suppressed hepatocyte apoptosis around the central vein. CZ attenuated hepatic PAI-1 protein and mRNA levels. CONCLUSIONS: Cilostazol attenuated MCT-induced SOS by preventing damage to liver sinusoidal endothelial cells and extravasated platelet aggregation. Hepatic PAI-1 levels were suppressed with CZ treatment.

Feasibility of dose-dense epirubicin and cyclophosphamide with subcutaneous pegfilgrastim 3.6 mg support: a single-center prospective study in Japan.

BACKGROUND: Dose-dense chemotherapy consisting of a combination of epirubicin and cyclophosphamide (EC) improves the survival of patients with breast cancer. Although pegfilgrastim was used at a subcutaneous dose of 6.0 mg in a pivotal study of dose-dense EC treatment, pegfilgrastim at a dose of 3.6 mg has been approved in Japan. We have assessed the feasibility of dose-dense EC treatment supported with a 3.6 mg dose of pegfilgrastim by evaluating the relative dose intensity (RDI) and safety of the treatment, together with measuring the pegfilgrastim concentrations remaining on the day of starting the next cycle of chemotherapy. METHODS: Patients with primary breast cancer received a total of 4 cycles of dose-dense EC treatment every 2 weeks, together with a subcutaneous injection of 3.6 mg pegfilgrastim on the day after chemotherapy. The serum granulocyte colony-stimulating factor (G-CSF) concentrations were measured on the 15th day of every chemotherapy cycle. RESULTS: From March 2015 through to July 2016, a total of 51 patients (median age 51 years; range 33-73 years) were studied. The mean RDI was 95.2% (range 60.0-100%). Although most adverse events were consistent with those reported in previous studies, pneumocystis pneumonia developed in two patients during the following course of docetaxel treatment. The median serum G-CSF concentration was 92.5 (range 30.4-440) pg/ml. CONCLUSIONS: With support provided by pegfilgrastim injection at a dose of 3.6 mg, dose-dense EC is feasible and associated with maintenance of a high RDI. There was no clinically significant accumulation of serum G-CSF concentrations associated with the use of a 3.6 mg dose of pegfilgrastim at 2-week intervals.

[A Case of Colon Cancer with Brain, Liver and Lung Metastasis Successfully Treated with Bevacizumab plus Xelox Therapy].

The patient was a 57-year-old woman. Preclinical examination of malignant lymphoma revealed 0-I sp type of early rectal cancer in the upper rectum, 20 cm from the anal margin. Endoscopic mucosal resection was performed and positive deep margins were pathologically diagnosed. Additional intestinal resection with lymph node dissection was deemed necessary, but ABVD therapy was initiated because the clinical stage of the malignant lymphoma was Stage III b or higher. Two months after detecting elevated CEA, S8 liver metastasis was pointed out, and examination of weakness of the right upper limb revealed nodular, multifocal brain metastasis. After chemotherapy for malignant lymphoma, bevacizumab(BV)plus Xelox therapy was initiated. After administering 4 courses, partial loss of multiple brain metastases and reduction of the liver metastatic lesion were confirmed; therefore, partial resection of the liver via laparoscopy was performed. After surgery, BV plus Xelox therapy was resumed, but since the lower lobular lung metastasis was confirmed after 8 courses, partial resection of the left lower lobe with thoracoscopy was performed. After lung resection, BV plus FOLFIRI therapy was administered, and 12 months after the onset of treatment for brain metastasis, recurrence was not detected.

[A Case of Spontaneous Esophageal Rupture That Required Operation during Adjuvant Chemotherapy for Transverse Colon Cancer].

A 75-year-old male underwent adjuvant chemotherapy with tegafur uracil(UFT)plus Leucovorin(LV)after surgery for transverse colon cancer(pT3pN0M0, ly1, v2, pStageⅡ). Although he had diarrhea(Grade 3)and vomiting(Grade 2)from day 15, he continued to take the medicine at his own discretion. He visited a hospital because of acute renal failure from severe dehydration. He went into shock after evacuation, and the computed tomography(CT)finding suggested a diagnosis of spontaneous esophageal rupture at the lower esophagus. We made a diagnosis of intrathoracic perforation of the esophagus by using thoracic drainage. Then, we performed an operation for mediastinal drainage via a transabdominal approach and the lesser omentum. He started ingestion from POD36 and transferred to another hospital on POD85. He had no disease recurrence in our outpatient care. We think that the spontaneous esophageal rupture occurred because of the frequent vomiting caused by the continued chemotherapy despite the severe side effects. Treatments must be selected by considering patients' life background and medical compliance, and common guidance in taking medications must be provided to elderly patients at the start of chemotherapy.

Dimerization of HIV-1 protease occurs through two steps relating to the mechanism of protease dimerization inhibition by darunavir.

Dimerization of HIV-1 protease (PR) subunits is an essential process for PR's acquisition of proteolytic activity, which plays a critical role in the maturation of HIV-1. Recombinant wild-type PR (PR(WT)) proved to dimerize, as examined with electrospray ionization mass spectrometry; however, two active site interface PR mutants (PR(T26A) and PR(R87K)) remained monomeric. On the other hand, two termini interface PR mutants (PR(1-C95A) and PR(97/99)) took both monomeric and dimeric forms. Differential scanning fluorimetry indicated that PR(1-C95A) and PR(97/99) dimers were substantially less stable than PR(WT) dimers. These data indicate that intermolecular interactions of two monomers occur first at the active site interface, generating unstable or transient dimers, and interactions at the termini interface subsequently occur, generating stable dimers. Darunavir (DRV), an HIV-1 protease inhibitor, inhibits not only proteolytic activity but also PR dimerization. DRV bound to protease monomers in a one-to-one molar ratio, inhibiting the first step of PR dimerization, whereas conventional protease inhibitors (such as saquinavir) that inhibit enzymatic activity but not dimerization failed to bind to monomers. DRV also bound to mutant PRs containing the transframe region-added PR (TFR-PR(D25N) and TFR-PR(D25N-7AA)), whereas saquinavir did not bind to TFR-PR(D25N) or TFR-PR(D25N-7AA). Notably, DRV failed to bind to mutant PR containing four amino acid substitutions (V32I, L33F, I54M, and I84V) that confer resistance to DRV on HIV-1. To our knowledge, the present report represents the first demonstration of the two-step PR dimerization dynamics and the mechanism of dimerization inhibition by DRV, which should help design further, more potent novel PIs.

[A Case of Esophagogastric Junction Adenocarcinoma with Neuroendocrine Differentiation].

Gastric cancer rarely contains neuroendocrine component. This mixed tumor is defined as neuroendocrine carcinoma (NEC), mixed adenoneuroendocrine carcinoma(MANEC)and so on according to the WHO classification. We report a patient with esophagogastric junction cancer with neuroendocrine differentiation(NED). The patient was 54-year-old man who diagnosed of esophagogastric junction cancer at the medical examination. Upper gastrointestinal endoscopy revealed a type 3 tumor at esophagogastric junction, and the pathological findings were poorly differentiated adenocarcinoma with focal positivity of chromogranin A. CT and FDG/PET revealed a metastatic regional lymph node. He had undergone proximal gastrectomy and lower esophagectomy, with dissection of D1+and double-tract reconstruction. Pathological findings revealed moderate to poorly differentiated adenocarcinoma containing chromogranin A-positive tumor cells. Neuroendocrine components were lower than 30%, and we diagnosed adenocarcinoma with NED. Standard treatment for gastric cancer with NED has not been established and more reports and reviews are required.

[A Case of Pneumocystis Pneumonia Developed during Chemotherapy for Sigmoid Colon Cancer].

A 69-year-old man with multiple liver metastases from sigmoid colon cancer received mFOLFOX6 plus cetuximab(Cmab) chemotherapy. A partial response was observed; hence, we performed an extended left hepatectomy, 3 partial liver resections, and a sigmoidectomy. After 4 courses of CapeOX, a recurrent lesion occurred between S8 and S7 of the liver, and we changed the regimen to FOLFIRI plus bevacizumab(BV). Three months later, he had Grade 3 febrile neutropenia and CT scan findings showed ground glass opacity in the superior lobes of both lungs. We diagnosed pneumocystis pneumonia(PCP)and administered steroids and trimethoprim/sulfamethoxazole. The signs of PCP thus improved. PCP during chemotherapy for gastrointestinal cancer is rarely reported, but recently it has increased.