Ramucirumab in patients with previously treated advanced hepatocellular carcinoma: Impact of liver disease aetiology.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is a common complication of chronic liver disease with diverse underlying aetiologies. REACH/REACH-2 were global phase III studies investigating ramucirumab in advanced HCC (aHCC) following sorafenib treatment. We performed an exploratory analysis of outcomes by liver disease aetiology and baseline serum viral load. METHODS: Meta-analysis was conducted in patients with aHCC and alpha-fetoprotein (AFP) ≥400 ng/mL (N = 542) from REACH/REACH-2 trials. Individual patient-level data were pooled with results reported by aetiology subgroup (hepatitis B [HBV] or C [HCV] and Other). Pre-treatment serum HBV DNA and HCV RNA were quantified using Roche COBAS AmpliPrep/COBAS TaqMan. Overall survival (OS) and progression-free survival (PFS) were evaluated using the Kaplan-Meier method and Cox proportional hazard model (stratified by study). RESULTS: Baseline characteristics were generally balanced between arms in each subgroup (HBV: N = 225, HCV: N = 127, Other: N = 190). No significant difference in treatment effect by aetiology subgroup was detected (OS interaction P-value = .23). Median OS (ramucirumab vs placebo) in months was 7.7 versus 4.5 (HR 0.74, 95% CI 0.55-0.99) for HBV, 8.2 versus 5.5 (HR 0.82, 95% CI 0.55-1.23) for HCV and 8.5 versus 5.4 (HR 0.56, 95% CI 0.40-0.79) for Other. Ramucirumab showed similar overall safety profiles across subgroups. Worst outcomes were noted in patients with a detectable HBV load. Use of HBV antiviral therapy, irrespective of viral load, was beneficial for survival, liver function and liver-specific adverse events. CONCLUSIONS: Ramucirumab improved survival across aetiology subgroups with a tolerable safety profile, supporting its use in patients with aHCC and elevated AFP.

Optimizing treatment duration with ramucirumab and paclitaxel by managing chemotherapy-associated toxicity: Review of four cases.

Gastric and gastroesophageal junction adenocarcinomas have poor prognoses. Ramucirumab is considered a second-line standard of care for patients with these cancers. Patients may develop chemotherapy-induced adverse events, and physicians may benefit from greater familiarity with treatment management in the setting of common adverse events. We report four cases of metastatic gastric or gastric and gastroesophageal junction adenocarcinoma treated with second-line ramucirumab plus paclitaxel. All patients developed chemotherapy-associated grade ⩾2 neutropenia and/or neuropathy, and one experienced recurrence of neurotoxicity, during second-line therapy. These adverse events were successfully managed by withholding or reducing the paclitaxel dose, without modifying the ramucirumab dosage schedule, and allowed administration of additional therapy cycles. In all patients, second-line therapy was associated with a best overall response of complete or partial response ranging from 2.2 to 12.4 months. These four cases demonstrate that paclitaxel-associated adverse events can be managed with dose modifications, thereby allowing continued therapy and potential survival benefits.

A diagnostic real-time PCR assay for the rapid identification of the tomato-potato psyllid, Bactericera cockerelli (Sulc, 1909) and development of a psyllid barcoding database.

The accurate and rapid identification of insect pests is an important step in the prevention and control of outbreaks in areas that are otherwise pest free. The potato-tomato psyllid Bactericera cockerelli (Sulc, 1909) is the main vector of 'Candidatus Liberibacter solanacearum' on potato and tomato crops in North America and New Zealand; and is considered a threat for introduction in Europe and other pest-free regions. This study describes the design and validation of the first species-specific TaqMan probe-based real-time PCR assay, targeting the ITS2 gene region of B. cockerelli. The assay detected B. cockerelli genomic DNA from adults, immatures, and eggs, with 100% accuracy. This assay also detected DNA from cloned plasmids containing the ITS2 region of B. cockerelli with 100% accuracy. The assay showed 0% false positives when tested on genomic and cloned DNA from 73 other psyllid species collected from across Europe, New Zealand, Mexico and the USA. This included 8 other species in the Bactericera genus and the main vectors of 'Candidatus Liberibacter solanacearum' worldwide. The limit of detection for this assay at optimum conditions was 0.000001ng DNA (~200 copies) of ITS2 DNA which equates to around a 1:10000 dilution of DNA from one single adult specimen. This assay is the first real-time PCR based method for accurate, robust, sensitive and specific identification of B. cockerelli from all life stages. It can be used as a surveillance and monitoring tool to further study this important crop pest and to aid the prevention of outbreaks, or to prevent their spread after establishment in new areas.

Detection of Nepovirus Vector and Nonvector Xiphinema Species in Grapevine.

Fanleaf degeneration is considered the most damaging viral disease of grapevine. The two major nepoviruses involved are Grapevine fanleaf virus (GFLV) and Arabis mosaic virus (ArMV) which are respectively and specifically transmitted by the dagger nematodes Xiphinema index and X. diversicaudatum. The methods described below are aimed at detecting four prevalent grapevine Xiphinema species: the vector species previously mentioned and two nonvector species X. vuittenezi and X. italiae.

A prospective audit of early stoma complications in colorectal cancer treatment throughout the Greater Manchester and Cheshire colorectal cancer network.

AIM: The study aimed to identify the incidence of early stoma problems after surgery for colorectal cancer to identify predisposing factors and to assess the effect on discharge from hospital and the greater need for community stoma care. METHOD: A prospective study of 192 patients was carried out over a six-month period in the 13 units of the Greater Manchester and Cheshire Cancer Network. Stoma problems were categorized into fistula, leakage, pancaking, necrosis, retraction, separation, stenosis, skin problems, parastomal hernia, suboptimal stoma site and need for resiting or refashioning. Differences in incidence between units (anonymized) were analysed, and the effect of stoma complications on length of hospital stay and the need for additional community stoma care was determined. RESULTS: One hundred and ninety-two patients with stomas were included, of which 52 (27.1%) were identified as being problematic (range 0-66.7% between units). Significant risk factors included stoma type (colostomy) (P < 0.05), short stoma length (P = 0.006), higher BMI (P = 0.043), emergency surgery (P = 0.002) and lack of preoperative site marking (P < 0.001). Problematic stomas were associated with longer hospital stay (P < 0.001) and increased community care (P < 0.001). CONCLUSION: Stoma type, stoma length, body mass index, emergency surgery and lack of preoperative marking were significant risk factors. Overall complication rates compare favourably with other studies.

Decreased internalisation of erbB1 mutants in lung cancer is linked with a mechanism conferring sensitivity to gefitinib.

A majority of gefitinib (IRESSA)-responsive tumours in non-small cell lung cancer have been found to carry mutations in ErbB1. Previously, it has been observed that internalisation-deficient ErbB1 receptors are strong drivers of oncogenesis. Using a computational model of ErbB1 trafficking and signalling, it is found that a deficiency in ErbB1 internalisation is sufficient to explain the observed signalling phenotype of these gefitinib-responsive ErbB1 mutants in lung cancer cell lines. Experimental tests confirm that gefitinib-sensitive cell lines with and without ErbB1 mutations exhibit markedly slower internalisation rates than gefitinib-insensitive cell lines. Moreover, the computational model demonstrates that reduced ErbB1 internalisation rates are mechanistically linked to upregulated AKT signalling. Experimentally it is confirmed that impaired internalisation of ErbB1 is associated with increased AKT activity, which can be blocked by gefitinib. On the basis of these experimental and computational results, it is surmised that gefitinib sensitivity is a marker of a reliance on AKT signalling for cell survival that may be brought about by impaired ErbB1 internalisation.

Cytoskeleton/endoplasmic reticulum collapse induced by prostaglandin J2 parallels centrosomal deposition of ubiquitinated protein aggregates.

Many neurodegenerative disorders, such as Parkinson disease, exhibit inclusion bodies containing ubiquitinated proteins. The mechanisms implicated in this aberrant protein deposition remain elusive. In these disorders signs of inflammation are also apparent in the affected central nervous system areas. We show that prostaglandin J2 (PGJ2), an endogenous product of inflammation, disrupts the cytoskeleton in neuronal cells. Furthermore, PGJ2 perturbed microtubule polymerization in vitro and decreased the number of free sulfhydryl groups on tubulin cysteines. A direct effect of PGJ2 on actin was not apparent, although actin filaments were altered in cells treated with PGJ2. This cyclopentenone prostaglandin triggered endoplasmic reticulum (ER) collapse and the redistribution of ER proteins, such as calnexin and catechol-O-methyltransferase, into a large centrosomal aggregate containing ubiquitinated proteins and alpha-synuclein. The PGJ2-dependent cytoskeletal rearrangement paralleled the development of the large centrosomal aggregate. Both of these events were replicated by treating cells with colchicine, which disrupts the microtubule/ER network, but not with brefeldin A, which impairs ER/Golgi transport. PGJ2 also perturbed 26 S proteasome assembly and activity, which preceded the accumulation of ubiquitinated proteins as detergent/salt-insoluble aggregates. Our data support a mechanism by which, upon PGJ2 treatment, cytoskeleton/ER collapse coincides with the relocation of ER proteins, other potentially neighboring proteins, and ubiquitinated proteins into centrosomal aggregates. Development of these large perinuclear aggregates is associated with disruption of the microtubule/ER network. This aberrant protein deposition, triggered by a product of inflammation, may be common to other compounds that disrupt microtubules and induce protein aggregation, such as MPP+ and rotenone, found to be associated with neurodegeneration.

Prostaglandin J2 alters pro-survival and pro-death gene expression patterns and 26 S proteasome assembly in human neuroblastoma cells.

Many neurodegenerative disorders are characterized by two pathological hallmarks: progressive loss of neurons and occurrence of inclusion bodies containing ubiquitinated proteins. Inflammation may be critical to neurodegeneration associated with ubiquitin-protein aggregates. We previously showed that prostaglandin J2 (PGJ2), one of the endogenous products of inflammation, induces neuronal death and the accumulation of ubiquitinated proteins into distinct aggregates. We now report that temporal microarray analysis of human neuroblastoma SK-N-SH revealed that PGJ2 triggered a "repair" response including increased expression of heat shock, protein folding, stress response, detoxification and cysteine metabolism genes. PGJ2 also decreased expression of cell growth/maintenance genes and increased expression of apoptotic genes. Over time pro-death responses prevailed over pro-survival responses, leading to cellular demise. Furthermore, PGJ2 increased the expression of proteasome and other ubiquitin-proteasome pathway genes. This increase failed to overcome PGJ2 inhibition of 26 S proteasome activity. Ubiquitinated proteins are degraded by the 26 S proteasome, shown here to be the most active proteasomal form in SK-N-SH cells. We demonstrate that PGJ2 impairs 26 S proteasome assembly, which is an ATP-dependent process. PGJ2 perturbs mitochondrial function, which could be critical to the observed 26 S proteasome disassembly, suggesting a cross-talk between mitochondrial and proteasomal impairment. In conclusion neurotoxic products of inflammation, such as PGJ2, may play a role in neurodegenerative disorders associated with the aggregation of ubiquitinated proteins by impairing 26 S proteasome activity and inducing a chain of events that culminates in neuronal cell death. Temporal characterization of these events is relevant to understanding the underlying mechanisms and to identifying potential early biomarkers.

Prostaglandin J2 reduces catechol-O-methyltransferase activity and enhances dopamine toxicity in neuronal cells.

There is clear evidence that an inflammatory reaction is mounted within the CNS following trauma, stroke, infection and seizures, thus augmenting brain damage. Furthermore, chronic inflammation of the CNS is implicated in many neurodegenerative disorders. However, the effects of products of inflammation on neuronal cells are poorly understood. Herein, we characterize the effects of a neurotoxic product of inflammation, prostaglandin J2 (PGJ2), on catechol-O-methyltransferase (COMT) in human dopaminergic-like neuroblastoma SK-N-SH cells and rat (P2) cortical neurons. COMT metabolizes catechols and catecholamines, a pathway relevant to neurodegeneration. PGJ2 treatment reduced the expression and activity of COMT, induced its sequestration into perinuclear aggregates and potentiated dopamine toxicity. The large COMT aggregates were co-localized with the centrosome, suggesting an aggresome-like structure. Our results indicate that COMT impairment induced by PGJ2 treatment may increase the concentration of dopamine (or its metabolites) to neurotoxic levels. Thus, COMT impairment following pro-inflammatory events may be a potential risk factor in neurodegeneration.

Managing a low-level mixed waste storage facility: a checklist for compliance to 40 CFR 266.

Do you hold radioactive material for storage and decay before disposal? Does your material contain any hazardous chemical as defined by the EPA? You may need to take a closer look at the Final Rule published by the EPA in the May 16, 2001 Federal Register.

Drug delivery performance of the mometasone furoate dry powder inhaler.

The mometasone furoate dry powder inhaler (MF-DPI) is a multiple-dose, breath-actuated inhaler that uses agglomerates of micronized MF and lactose. In vitro analyses evaluated dose uniformity, variability, and particle size distribution of the MF-DPI. Tests of first, middle, and end doses from 10 inhalers each of the 200-microg MF/inhalation and 400-microg MF/inhalation dose sizes found that delivered doses (doses emitted from the inhaler) ranged from 91% to 112% of claimed doses for all tested DPIs. The mean MF doses delivered at 28.3 L/min were 100% and 94% of the doses delivered at 60 L/min for the 200-microg and 400-microg dose sizes, respectively; the relative standard deviation of doses was < or = 6.1% within this range of inhalation rates. At a flow rate of 60 L/min, the mean delivered doses, compared to claimed doses for inspiration times of 1-3 sec, were 102-104% for the 200-microg dose size and 98.8-102% for the 400-microg dose size. The mean cumulative fraction of dose delivered at 60 L/min for 2 sec which consisted of particles of <6.5 microm in diameter was 39.9% (+/-2.5 SD; n = 9) for the 200-microg dose size and 35.6% (+/-3.4 SD; n = 9) for the 400-microg dose size. All MF-DPI inhalers tested were well within U.S. and European compendial standards and regulatory guidelines for dose uniformity. An appropriate and reproducible fraction of the delivered dose was within the optimal particle size range for therapeutic effectiveness.

The application of real-time PCR to the identification, detection and quantification of Pyrenophora species in barley seed.

Summary A real-time quantitative PCR technique has been used to develop a rapid and sensitive seed health test for Pyrenophora spp. on barley seed. Using the fluorescent reporter dye SYBR Green I for real-time detection of PCR amplification, pathogen DNA extracted from infected seed can be quantified to the picogram level. The amount of Pyrenophora DNA extracted from seed samples of an artificial infection level gradient, constructed by mixing infected and uninfected seed, correlated with good agreement (r = 0.931) to percentage infection levels of the same samples measured by agar plate testing. In addition, a correlation of r = 0.883 was obtained between the two testing methods for naturally infected seed, ranging from 0% to 89% infection. Samples could be quantified to below the 2% voluntary threshold required for deciding on seed treatment. The proposed test was performed in three parts: (i) quantification of Pyrenophora spp. infection using Pyrenophora-specific PCR primers; (ii) test of any negative samples from (i) with barley-specific PCR primers to check the DNA extraction process; (iii) test of positive samples from (i) for the presence of Pyrenophora graminea using P. graminea-specific PCR primers. All PCRs were performed in the LightCycler instrument allowing each PCR run and analysis to be completed within 30 min. With the current daily receipt of samples (batches up to 16) the test can be completed in 8 h, compared to 7 days for the traditional agar plate test.

Heightening hazard awareness using simple structural models in the real and virtual environments

Heightening awareness to earthquake hazard required the used of diverse educational approaches. The appropiateness of a particular approach depends on the intended audience. techniques used to educate an audience of corporate professionals, building owners, and the general public, may differ from those used with engineering professionals (AU)

Tumorigenicity of adenovirus-transformed cells and their sensitivity to tumor necrosis factor alpha and NK/LAK cell cytolysis.

Sensitivity of a library of cloned adenovirus-transformed rat cell lines of varying tumorigenicity to cytotoxic action of tumor necrosis factor alpha (TNF alpha) was studied and correlated with their sensitivity to NK/LAK cell cytolysis. Our data confirm earlier reports that expression of the E1A oncogene of Ad2 or Ad5 is associated with sensitivity of transformed cells to TNF alpha and also NK/LAK cytotoxicity. Ad2-transformed cell line which expresses the E3 early region in addition to the E1 gene block is resistant to TNF alpha, but remains sensitive to NK/LAK cells. All cell lines which express the E1A oncogene of highly oncogenic Ad12 are resistant to NK but not LAK cells. Their sensitivity to TNF alpha, however, varies over a broad range and does not correlate with either their susceptibility to NK/LAK cytolysis or their tumorigenic potential.

Region E1a of highly oncogenic adenovirus 12 in transformed cells protects against NK but not LAK cytolysis.

The sensitivity of a library of adenovirus-transformed rat cell lines to lysis with highly enriched populations of rat NK cells and LAK cells activated in vitro by culture with recombinant human IL-2 was studied and correlated with the tumorigenic potential of these cell lines. The cell lines studied express the transforming E1 region of highly oncogenic Ad12 or nononcogenic Ad2. Two cell lines express recombinant E1A regions. In one the E1A genes were of Ad12 origin and the E1B region was derived from nononcogenic Ad5. In the other, the E1A region was from Ad5 and the E1B genes from Ad12. All cell lines tested which express the early region E1 of Ad12 are tumorigenic in syngeneic rats. The two cell lines which express only the E1A or the E1B genes of Ad12, and the Ad2-transformed cells did not induce tumors. Transformed cell lines which express the E1A region of nononcogenic Ad2 or Ad5 are efficiently killed by rat NK cells, but cells which express the Ad12 E1A genes are resistant to lysis by NK-enriched cell fractions even at high effector:target ratio; cells containing the Ad12 E1 region are also resistant to IFN-activated NK cells. Although such NK-resistant cells have a uniformly low level of class I MHC antigen, their resistance is not affected by MHC antigen level modulation by rat IFN. Ad12-transformed cells resistant to endogeneous NK cells, however, are efficiently lysed by LAK cells stimulated in vitro by recombinant IL-2. Sensitivity to LAK killing is unaffected by IFN treatment of target cells. These results show that expression of the E1A region of highly oncogenic Ad12 in the transformed cells, which confers resistance to endogeneous NK cells, fails to protect against lysis by LAK cells.

Regional cerebral glucose metabolism of newborn infants measured by positron emission tomography.

The new diagnostic technique, positron emission tomography with 18F-2-fluoro-2-deoxy-D-glucose (18FDG), was used to measure regional cerebral glucose metabolism in five newborn infants with demonstrated structural abnormalities of the brain. 18FDG was synthesized, diluted in normal saline and injected intravenously. After one hour, tomographic slices of the brain were obtained, the level of the slices being defined relative to the cerebral ventricles. Glucose metabolism of grey- and white-matter structures in the brain could be differentiated clearly. Decreased glucose metabolism was identified in regions of the brain shown by computerized axial tomography to be structurally abnormal. Positron emission tomography is a promising new diagnostic tool for the study of newborn infants with suspected abnormalities of brain function.

A positron emission tomograph to study brain metabolism in man.

The construction of a single ring positron emission tomograph with which to examine the brain is described. The ring comprises 160 bismuth germanate detectors that are not separated from each other by high Z septa. The thickness of the slice examined is 2 cm and the inherent resolution is 7 mm in the plane. The performance of the tomograph and representative clinical studies are illustrated.