Durvalumab Plus Olaparib in Previously Untreated, Platinum-Ineligible Patients With Metastatic Urothelial Carcinoma: A Multicenter, Randomized, Phase II Trial (BAYOU).

PURPOSE: Homologous recombination repair gene mutations (HRRm) are common in urothelial carcinoma (UC), rendering tumor cells sensitive to poly (ADP-ribose) polymerase (PARP) inhibition. We assessed efficacy and safety of durvalumab (anti-programmed cell death ligand-1) plus olaparib (PARP inhibitor) in patients with metastatic UC (mUC). METHODS: This randomized, multicenter, double-blind, phase II trial enrolled untreated, platinum-ineligible patients with mUC. Patients (N = 154) were randomly assigned 1:1 to receive durvalumab (1,500 mg intravenously once every 4 weeks) plus olaparib (300 mg orally, twice daily) or durvalumab plus placebo. The primary end point was progression-free survival (PFS) assessed by investigators per RECIST version 1.1. Secondary end points included overall survival in all patients and PFS in patients with HRRm. RESULTS: Overall, median PFS was 4.2 months (95% CI, 3.6 to 5.6) for durvalumab plus olaparib and 3.5 months (95% CI, 1.9 to 5.1) for durvalumab plus placebo (hazard ratio [HR], 0.94; 95% CI, 0.64 to 1.39; log-rank P value, .789). Median overall survival was 10.2 months (95% CI, 7.0 to 13.9) and 10.7 months (95% CI, 7.2 to 17.3), respectively (HR, 1.07; 95% CI, 0.72 to 1.61). In the 20% of patients with HRRm, median PFS was 5.6 months (95% CI, 1.9 to 8.1) and 1.8 months (95% CI, 1.7 to 2.2), respectively (HR, 0.18; 95% CI, 0.06 to 0.47). Treatment-related grade 3 or 4 adverse events occurred in 18% and 9% of patients, respectively. CONCLUSION: Adding olaparib to durvalumab did not improve survival outcomes in an unselected mUC population. Efficacy outcomes with durvalumab were similar to those reported for other anti-programmed cell death-1/programmed cell death ligand-1 agents. However, the results of secondary analyses suggest a potential role for PARP inhibition in patients with UC harboring HRRm.

Genetic and phenotypic characteristics of Clostridium (Clostridioides) difficile from canine, bovine, and pediatric populations.

OBJECTIVES: Carriage of Clostridioides difficile by different species of animals has led to speculation that animals could represent a reservoir of this pathogen for human infections. The objective of this study was to compare C. difficile isolates from humans, dogs, and cattle from a restricted geographic area. METHODS: C. difficile isolates from 36 dogs and 15 dairy calves underwent whole genome sequencing, and phenotypic assays assessing growth and virulence were performed. Genomes of animal-derived isolates were compared to 29 genomes of isolates from a pediatric population as well as 44 reference genomes. RESULTS: Growth rates and relative cytotoxicity of isolates were significantly higher and lower, respectively, in bovine-derived isolates compared to pediatric- and canine-derived isolates. Analysis of core genes showed clustering by host species, though in a few cases, human strains co-clustered with canine or bovine strains, suggesting possible interspecies transmission. Geographic differences (e.g., farm, litter) were small compared to differences between species. In an analysis of accessory genes, the total number of genes in each genome varied between host species, with 6.7% of functional orthologs differentially present/absent between host species and bovine-derived strains having the lowest number of genes. Canine-derived isolates were most likely to be non-toxigenic and more likely to carry phages. A targeted study of episomes identified in local pediatric strains showed sharing of a methicillin-resistance plasmid with dogs, and historic sharing of a wide range of episomes across hosts. Bovine-derived isolates harbored the widest variety of antibiotic-resistance genes, followed by canine CONCLUSIONS: While C. difficile isolates mostly clustered by host species, occasional co-clustering of canine and pediatric-derived isolates suggests the possibility of interspecies transmission. The presence of a pool of resistance genes in animal-derived isolates with the potential to appear in humans given sufficient pressure from antibiotic use warrants concern.

Brine-driven destruction of clay minerals in Gale crater, Mars.

Mars' sedimentary rock record preserves information on geological (and potential astrobiological) processes that occurred on the planet billions of years ago. The Curiosity rover is exploring the lower reaches of Mount Sharp, in Gale crater on Mars. A traverse from Vera Rubin ridge to Glen Torridon has allowed Curiosity to examine a lateral transect of rock strata laid down in a martian lake ~3.5 billion years ago. We report spatial differences in the mineralogy of time-equivalent sedimentary rocks <400 meters apart. These differences indicate localized infiltration of silica-poor brines, generated during deposition of overlying magnesium sulfate-bearing strata. We propose that destabilization of silicate minerals driven by silica-poor brines (rarely observed on Earth) was widespread on ancient Mars, because sulfate deposits are globally distributed.

Evidence for Multiple Diagenetic Episodes in Ancient Fluvial-Lacustrine Sedimentary Rocks in Gale Crater, Mars.

The Curiosity rover's exploration of rocks and soils in Gale crater has provided diverse geochemical and mineralogical data sets, underscoring the complex geological history of the region. We report the crystalline, clay mineral, and amorphous phase distributions of four Gale crater rocks from an 80-m stratigraphic interval. The mineralogy of the four samples is strongly influenced by aqueous alteration processes, including variations in water chemistries, redox, pH, and temperature. Localized hydrothermal events are evidenced by gray hematite and maturation of amorphous SiO2 to opal-CT. Low-temperature diagenetic events are associated with fluctuating lake levels, evaporative events, and groundwater infiltration. Among all mudstones analyzed in Gale crater, the diversity in diagenetic processes is primarily captured by the mineralogy and X-ray amorphous chemistry of the drilled rocks. Variations indicate a transition from magnetite to hematite and an increase in matrix-associated sulfates suggesting intensifying influence from oxic, diagenetic fluids upsection. Furthermore, diagenetic fluid pathways are shown to be strongly affected by unconformities and sedimentary transitions, as evidenced by the intensity of alteration inferred from the mineralogy of sediments sampled adjacent to stratigraphic contacts.

Retrospective study on mixed neuroendocrine non-neuroendocrine neoplasms from five European centres.

BACKGROUND: Mixed neuroendocrine non-neuroendocrine neoplasm (MiNEN) is a rare diagnosis, mainly encountered in the gastro-entero-pancreatic tract. There is limited knowledge of its epidemiology, prognosis and biology, and the best management for affected patients is still to be defined. AIM: To investigate clinical-pathological characteristics, treatment modalities and survival outcomes of a retrospective cohort of patients with a diagnosis of MiNEN. METHODS: Consecutive patients with a histologically proven diagnosis of MiNEN were identified at 5 European centres. Patient data were retrospectively collected from medical records. Pathological samples were reviewed to ascertain compliance with the 2017 World Health Organisation definition of MiNEN. Tumour responses to systemic treatment were assessed according to the Response Evaluation Criteria in Solid Tumours 1.1. Kaplan-Meier analysis was applied to estimate survival outcomes. Associations between clinical-pathological characteristics and survival outcomes were explored using Log-rank test for equality of survivors functions (univariate) and Cox-regression analysis (multivariable). RESULTS: Sixty-nine consecutive patients identified; Median age at diagnosis: 64 years. Males: 63.8%. Localised disease (curable): 53.6%. Commonest sites of origin: colon-rectum (43.5%) and oesophagus/oesophagogastric junction (15.9%). The neuroendocrine component was; predominant in 58.6%, poorly differentiated in 86.3%, and large cell in 81.25%, of cases analysed. Most distant metastases analysed (73.4%) were occupied only by a poorly differentiated neuroendocrine component. Ninety-four percent of patients with localised disease underwent curative surgery; 53% also received perioperative treatment, most often in line with protocols for adenocarcinomas from the same sites of origin. Chemotherapy was offered to most patients (68.1%) with advanced disease, and followed protocols for pure neuroendocrine carcinomas or adenocarcinomas in equal proportion. In localised cases, median recurrence free survival (RFS); 14.0 mo (95%CI: 9.2-24.4), and median overall survival (OS): 28.6 mo (95%CI: 18.3-41.1). On univariate analysis, receipt of perioperative treatment (vs surgery alone) did not improve RFS (P = 0.375), or OS (P = 0.240). In advanced cases, median progression free survival (PFS); 5.6 mo (95%CI: 4.4-7.4), and median OS; 9.0 mo (95%CI: 5.2-13.4). On univariate analysis, receipt of palliative active treatment (vs best supportive care) prolonged PFS and OS (both, P < 0.001). CONCLUSION: MiNEN is most commonly driven by a poorly differentiated neuroendocrine component, and has poor prognosis. Advances in its biological understanding are needed to identify effective treatments and improve patient outcomes.

Competitive Testing of the WHO 2010 versus the WHO 2017 Grading of Pancreatic Neuroendocrine Neoplasms: Data from a Large International Cohort Study.

BACKGROUND: The World Health Organization (WHO) and the American Joint Cancer Committee (AJCC) modified the grading of pancreatic neuroendocrine neoplasms from a three-tier (WHO-AJCC 2010) to a four-tier system by introducing the novel category of NET G3 (WHO-AJCC 2017). OBJECTIVES: This study aims at validating the WHO-AJCC 2017 and identifying the most effective grading system. METHOD: A total of 2,102 patients were enrolled; entry criteria were: (i) patient underwent surgery; (ii) at least 2 years of follow-up; (iii) observation time up to 2015. Data from 34 variables were collected; grading was assessed and compared for efficacy by statistical means including Kaplan-Meier method, Cox regression analysis, Harrell's C statistics, and Royston's explained variation in univariable and multivariable analyses. RESULTS: In descriptive analysis, the two grading systems demonstrated statistically significant differences for the major category sex but not for age groups. In Cox regression analysis, both grading systems showed statistically significant differences between grades for OS and EFS; however, no statistically significant difference was observed between the two G3 classes of WHO-AJCC 2017. In multivariable analysis for the two models fitted to compare efficacy, the two grading systems performed equally well with substantially similar optimal discrimination and well-explained variation for both OS and EFS. The WHO-AJCC 2017 grading system retained statistically significant difference between the two G3 classes for OS but not for EFS. CONCLUSIONS: The WHO-AJCC 2017 grading system is at least equally performing as the WHO-AJCC 2010 but allows the successful identification of the most aggressive PanNET subgroup. Grading is confirmed as probably the most powerful tool for predicting patient survival.

The mouse wellhaarig (we) mutations result from defects in epidermal-type transglutaminase 3 (Tgm3).

The recessive wellhaarig (we) mutations, named for the wavy coat and curly whiskers they generate in homozygotes, have previously been mapped on mouse Chromosome 2. To further limit the possible location of the we locus, we crossed hybrid (C57BL/6×AKR)F1, we(4J)/+ females with AKR, we(4J)/we(4J) mutant males to create a large backcross family that was typed for various microsatellite markers and single-nucleotide polymorphisms (SNPs) that distinguish strains AKR and B6. This analysis restricted the location of we(4J) between sites that flank only one gene known to be expressed in skin: epidermal-type transglutaminase 3 (Tgm3). To test Tgm3 as a candidate for the basis of the wellhaarig phenotype we took two approaches. First, we sequenced all Tgm3 coding regions in mice homozygous for four independent, naturally-occurring wellhaarig alleles (we, we(Bkr), we(3J) and we(4J)) and found distinct defects in three of these mutants. Second, we crossed mice homozygous for an induced mutant allele of Tgm3 (Tgm3(Btlr)) with mice heterozygous for one of the wellhaarig alleles we possess (we(4J) or we(Bkr)) to test for complementation. Because the progeny inheriting both a recessive we allele and a recessive Tgm3(Btlr) allele displayed wavy hair, we conclude that the classic wellhaarig mutations result from defects in Tgm3.

Double-layer video transmission over decode-and-forward wireless relay networks using hierarchical modulation.

We consider a wireless relay network with a single source, a single destination, and a multiple relay. The relays are half-duplex and use the decode-and-forward protocol. The transmit source is a layered video bitstream, which can be partitioned into two layers, a base layer (BL) and an enhancement layer (EL), where the BL is more important than the EL in terms of the source distortion. The source broadcasts both layers to the relays and the destination using hierarchical 16-QAM. Each relay detects and transmits successfully decoded layers to the destination using either hierarchical 16-QAM or QPSK. The destination can thus receive multiple signals, each of which can include either only the BL or both the BL and the EL. We derive the optimal linear combining method at the destination, where the uncoded bit error rate is minimized. We also present a suboptimal combining method with a closed-form solution, which performs very close to the optimal. We use the proposed double-layer transmission scheme with our combining methods for transmitting layered video bitstreams. Numerical results show that the double-layer scheme can gain 2-2.5 dB in channel signal-to-noise ratio or 5-7 dB in video peak signal-to-noise ratio, compared with the classical single-layer scheme using conventional modulation.

Endothelial nitric oxide synthase and heme oxygenase-1 act independently in liver ischemic preconditioning.

BACKGROUND: ischemic preconditioning (IPC) protects against liver ischemia-reperfusion (IR) injury. The mechanism involves nitric oxide metabolism but the importance of endothelial nitric oxide synthase (eNOS) has not been established. Heme oxygenase-1 (HO-1) protects against liver IR but it is unclear if this depends on nitric oxide synthase. MATERIALS AND METHODS: A mouse model of IPC with liver IR using wild-type (WT) and eNOS transgenic knockout (eNOS-/-) mice was developed to study the role of eNOS and its relationship to HO-1. Serum alanine aminotransferase level, liver histopathologic injury scores, and liver microcirculatory blood flow were measured. Western blots measured liver HO-1/2, eNOS, phosphorylated eNOS, inducible nitric oxide synthase, and reverse transcription-polymerase chain reaction (HO-1). A set of 24-h recovery experiments was undertaken on WT mice with measurement of serum alanine aminotransferase level, histologic injury score, and HO-1 by Western blot. RESULTS: In WT animals, IPC preceding IR resulted in a reduction in hepatocellular and histologic injury, and improvement in parenchymal perfusion. In contrast, IPC in the eNOS-/- model did not protect the animals from IR injury. There was no difference between the eNOS and phosphorylated eNOS expression in all the WT groups. HO-1 protein was not detected in the nonrecovery groups but HO-1 messenger RNA was detected in all groups. In WT recovery experiments, IPC was protective against IR injury. HO-1 protein was detected in the IPC + IR and IR only groups but not in the sham group. CONCLUSIONS: This study developed and used an eNOS-/- model to demonstrate that eNOS mediates protection against liver IR injury by IPC. The eNOS expression and activity and HO-1 expression are increased independently in liver IPC and IR, with HO-1 expression increased in the later stages of IPC and IR.

National influenza surveillance in Vietnam, 2006-2007.

In 2006, national influenza surveillance was implemented in Vietnam. Epidemiologic and demographic data and a throat swab for influenza testing were collected from a subset of outpatients with influenza-like illness (ILI). During January 1, 2006 through December 31, 2007, of 184,521 ILI cases identified at surveillance sites, 11,082 were tested and 2112 (19%) were positive for influenza by reverse transcription polymerase chain reaction. Influenza viruses were detected year-round, and similar peaks in influenza activity were observed in all surveillance regions, coinciding with cooler and rainy periods. Studies are needed to ascertain the disease burden and impact of influenza in Vietnam.

An off-seasonal amantadine-resistant H3N2 influenza outbreak in Japan.

An off-season community influenza outbreak with high prevalence of amantadine-resistant influenza A/H3N2 occurred during September-October 2005 in Nagasaki Prefecture, Japan, prior to standard influenza circulation. A total of 48 patients with influenza-like-illness (ILI) visited a clinic during the outbreak and 27 (69.2%) of 39 ILI patients were positive for influenza A with rapid antigen testing (Quick Vue Rapid SP Influ). Nine patients were not tested because their symptoms were compatible for influenza without examination. Nasopharyngeal swabs were obtained from 4 of 27 rapid test positive patients, and influenza H3N2 strain was isolated from one out of four. The 4 nasopharyngeal samples were positive for influenza A M2 gene in polymerase chain reaction, and sequencing results all showed identical mutation at position 31, serine to asparagine (S31N) in the gene, conferring amantadine resistance. The phylogenetic tree analysis demonstrated that the hemagglutinin (HA) gene sequences of the 4 samples formed a distinct cluster (named clade N) from recent circulating H3N2 strains, characterized by dual mutations at position 193, serine to phenylalanine (S193F), and at position 225, asparatic acid to asparagine (D225N). Our findings suggested that an off-season community influenza outbreak in Nagasaki was caused by a distinct clade in H3N2 (named clade N), which possessed characteristics of amantadine resistance.

Uncoupling the senescent phenotype from telomere shortening in hydrogen peroxide-treated fibroblasts.

Normal human cells have a limited replicative potential and inevitably reach replicative senescence in culture. Replicatively senescent cells show multiple molecular changes, some of which are related to the irreversible growth arrest in culture, whereas others resemble the changes occurring during the process of aging in vivo. Telomeres shorten as a result of cell replication and are thought to serve as a replicometer for senescence. Recent studies show that young cells can be induced to develop features of senescence prematurely by damaging agents, chromatin remodeling, and overexpression of ras or the E2F1 gene. Accelerated telomere shortening is thought to be a mechanism of premature senescence in some models. In this work, we test whether the acquisition of a senescent phenotype after mild-dose hydrogen peroxide (H(2)O(2)) exposure requires telomere shortening. Treating young HDFs with 150 microM H(2)O(2) once or 75 microM H(2)O(2) twice in 2 weeks causes long-term growth arrest, an enlarged morphology, activation of senescence-associated beta-galactosidase, and elevated expression of collagenase and clusterin mRNAs. No significant telomere shortening was observed with H(2)O(2) at doses ranging from 50 to 200 microM. Weekly treatment with 75 microM H(2)O(2) also failed to induce significant telomere shortening. Failure of telomere shortening correlated with an inability to elevate p16 protein or mRNA in H(2)O(2)-treated cells. In contrast, p21 mRNA was elevated over 40-fold and remained at this level for at least 2 weeks after a pulse treatment of H(2)O(2). The role of cell cycle checkpoints centered on p21 in premature senescence induced by H(2)O(2) is discussed here.

Molecular mechanisms of cardiac hypertrophy induced by toxicants.

Cardiac hypertrophy is an end point of chronic cardiac toxicity from a number of toxicants. Doxorubicin, cocaine, acetaldehyde, monocrotaline, and azide are examples of these toxicants, which may induce hypertrophy by increasing oxidants, circulating levels of catecholamines, and hemodynamic load or by inducing hypoxia. We summarize here the major signal transduction pathways and common changes in gene expression found with the classical hypertrophy inducers angiotensin II, endothelin 1, and catecholamines. Activation of G-proteins, calcium signaling, phosphoinositide 3-kinase (PI3K), certain family members of protein kinase Cs (PKCs), and three branches of mitogenactivated protein kinases (MAPKs), i.e. extracellular signal-regulated kinases (ERKs), p38, and c-Jun N-terminal kinases (JNKs), are important for developing a hypertrophic phenotype in cardiomyocytes. Characteristic changes of gene expression in hypertrophy include the elevated transcription of atrial natriuretic factor (ANF), beta-myosin heavy chain (beta MHC), skeletal alpha-actin (SkA), certain variants of integrins and perhaps tubulin genes, and reduced expression of the sarcoplasmic reticulum proteins phospholamban and sarco(endo)plasmic reticulum Ca2+-ATPase 2 alpha (SERCA2 alpha), and of the ryanodine receptors. Although which toxicants induce these molecular changes remains to be tested, increasing lines of evidence support that oxidants play a central role in cardiac hypertrophy. Oxidants activate small G-proteins, calcium signaling, PI3K, PKCs, and MAPKs. Oxidants cause cardiomyocytes to enlarge in vitro. Recent developments in transgenic, genomic, and proteomic technologies will provide needed tools to reveal the mechanism of chronic cardiac toxicity at the cellular and molecular levels.

Involvement of Rb family proteins, focal adhesion proteins and protein synthesis in senescent morphogenesis induced by hydrogen peroxide.

Early passage human diploid fibroblasts develop senescent morphology prematurely within a week after a 2-hour pulse treatment with low or mild dose H(2)O(2). We test here the role of cell cycle checkpoints, cytoskeletal proteins and de novo protein synthesis in senescent morphogenesis following H(2)O(2) treatment. H(2)O(2) treatment causes transient elevation of p53 protein and prolonged inhibition of Rb hyperphosphorylation. Expression of human papillomaviral E6 gene prevented elevation of p53 but did not affect senescent morphogenesis. Expression of human papillomaviral E7 gene reduced the level of Rb protein and prevented induction of senescent morphology by H(2)O(2). The mutants of the E7 gene, in which the Rb family protein binding site was destroyed, could not reduce Rb protein or prevent H(2)O(2) from inducing senescent morphology. Senescent-like cells showed enhanced actin stress fibers. In untreated cells, vinculin and paxillin preferentially distributed along the edge of the cells. In contrast, vinculin and paxillin distributed randomly and sporadically throughout senescent-like cells. E7 expression prevented enhancement of actin filament formation and redistribution of vinculin or paxillin. Neither wild-type nor E7 cells showed changes in the protein level of actin, vinculin or paxillin measured by western blot after H(2)O(2) treatment. Finally, depletion of methionine in the culture medium after H(2)O(2) treatment prevented senescent morphogenesis without affecting dephosphorylation of Rb protein. Our results suggest that senescent morphology likely develops by a program involving activated Rb family proteins, enhancement of actin stress fibers, redistribution of focal adhesion proteins and de novo protein synthesis.

Hydrogen peroxide dose dependent induction of cell death or hypertrophy in cardiomyocytes.

Cardiomyocyte hypertrophy and cell death are often observed in the end stages of heart failure. The triggers of these two cellular processes are not known under most pathological conditions. Oxidants are by-products of aerobic metabolism. The level of oxidants increases as a result of ischemic reperfusion. Using H9C2 and primary cultured neonatal rat cardiomyocytes, we found that a 2-h pulse treatment with H(2)O(2) at 250 microM or lower caused activation of DEVD sequence specific caspases. The activity of DEVD-ase peaked with 200 microM H(2)O(2) at 24 h. While a fraction of the cells detached and showed nuclear condensation, the majority of the cells (>55%) survived the treatment and appeared to enlarge when cultured for 5 days. These cells showed increases in cell surface area, cell volume, and protein content. With 200 microM H(2)O(2), treated cells appeared to be six times bigger in volume and contained three times more protein per cell than untreated cells. The enlarged cells showed enhanced actin stress fibers and disrupted myofibrils. Our data indicate that while H(2)O(2) can cause cell death, the surviving cardiomyocytes undergo hypertrophy.

Measurements of hydrogen peroxide induced premature senescence: senescence-associated beta-galactosidase and DNA synthesis index in human diploid fibroblasts with down-regulated p53 or Rb.

Human diploid fibroblasts (HDFs) inevitably undergo replicative senescence in culture after serial passages. Recent work indicates that early passage HDFs undergo irreversible growth arrest and develop features of senescence after being treated with oxidants and other agents. Senescence is usually measured by a decrease in DNA synthesis index and an increase in the activity of senescence-associated beta-galactosidase (SA beta-gal). We compared these two measurements here and found that IMR-90 HDFs lost the ability to synthesize DNA immediately but did not activate SA beta-gal until 4 days after the treatment with 75 microM or 0.75 pmol/cell H2O2. Expression of human papillomaviral E6 or/and E7 genes results in reduction of p53 or/and Rb protein levels and increases in ED50 for DNA synthesis inhibition or SA beta-gal expression. A small fraction of wild type and E7 expressing cells could not synthesize DNA and did not express SA beta-gal one week following the treatment with H2O2 at doses lower than 150 microM or 1.5 pmol/cell. The dose response curve of SA beta-gal activation overlapped with that of DNA synthesis inhibition in E6 and E6E7 expressing cells. The results indicate that the expression of SA beta-gal correlates with inability of DNA synthesis in the majority of wild type, E6, E7 or E6E7 expressing cells one week after H2O2 treatment.

The role of macrophages in the protection of mice against leptospirosis: in vitro and in vivo studies.

Balb/c mice are naturally resistant to infection with Leptospira interrogans serovar copenhageni, but leptospires were not phagocytosed by mouse peritoneal macrophages in vitro without added specific antibody. Similar results were obtained irrespective of whether leptospires were viable or killed, virulent or avirulent, or whether macrophages were obtained from normal mice, immunized mice or mice previously infected with BCG. Suppression or stimulation of macrophage function in vivo did not affect the outcome of infection of immunosuppressed mice with leptospires; specific antibody was essential for protection from infection.