BACKGROUND

Atherosclerotic cardiovascular disease and malnutrition are widely recognized as leading causes of the increased morbidity and mortality observed in uremic patients. C-reactive protein (CRP), an acute-phase protein, is a predictor of cardiovascular mortality in nonrenal patient populations. In chronic renal failure (CRF), the prevalence of an acute-phase response has been associated with an increased mortality.

METHODS

One hundred and nine predialysis patients (age 52 +/- <em>1</em> years) with terminal CRF (glomerular filtration rate 7 +/- <em>1</em> <em>ml</em>/min) were studied. By using noninvasive B-mode ultrasonography, the cross-sectional carotid intima-media area was calculated, and the presence or absence of carotid plaques was determined. Nutritional status was assessed by subjective global assessment (SGA), dual-energy x-ray absorptiometry (DXA), serum albumin, serum creatinine, serum urea, and 24-hour urine urea excretion. The presence of an inflammatory reaction was assessed by CRP, fibrinogen (N = 46), and tumor necrosis factor-alpha (TNF-alpha; N = 87). Lipid parameters, including Lp(a) and apo(a)-isoforms, as well as markers of oxidative stress (autoantibodies against oxidized low-density lipoprotein and vitamin E), were also determined.

RESULTS

Compared with healthy controls, CRF patients had an increased mean carotid intima-media area (<em>1</em>8.3 +/- 0.6 vs. <em>1</em>3.2 +/- 0.7 mm2, P < 0.000<em>1</em>) and a higher prevalence of carotid plaques (72 vs. 32%, P = 0.00<em>1</em>). The prevalence of malnutrition (SGA 2 to 4) was 44%, and 32% of all patients had an acute-phase response (CRP>> or = <em>1</em>0 mg/liter). Malnourished patients had higher CRP levels (23 +/- 3 vs. <em>1</em>3 +/- 2 mg/liter, P < 0.0<em>1</em>), elevated calculated intima-media area (20.2 +/- 0.8 vs. <em>1</em>6.9 +/- 0.7 mm2, P < 0.0<em>1</em>) and a higher prevalence of carotid plaques (90 vs. 60%, P < 0.000<em>1</em>) compared with well-nourished patients. During stepwise multivariate analysis adjusting for age and gender, vitamin E (P < 0.05) and CRP (P < 0.05) remained associated with an increased intima-media area. The presence of carotid plaques was significantly associated with age (P < 0.00<em>1</em>), log oxidized low-density lipoprotein (oxLDL; P < 0.0<em>1</em>), and small apo(a) isoform size (P < 0.05) in a multivariate logistic regression model.

CONCLUSIONS

These results indicate that the rapidly developing atherosclerosis in advanced CRF appears to be caused by a synergism of different mechanisms, such as malnutrition, inflammation, oxidative stress, and genetic components. Apart from classic risk factors, low vitamin E levels and elevated CRP levels are associated with an increased intima-media area, whereas small molecular weight apo(a) isoforms and increased levels of oxLDL are associated with the presence of carotid plaques.

OBJECTIVE

Studies in experimental and human heart failure suggest that phosphodiesterase-5 inhibitors may enhance cardiovascular function and thus exercise capacity in heart failure with preserved ejection fraction (HFPEF).

OBJECTIVE

To determine the effect of the phosphodiesterase-5 inhibitor sildenafil compared with placebo on exercise capacity and clinical status in HFPEF.

METHODS

Multicenter, double-blind, placebo-controlled, parallel-group, randomized clinical trial of 2<em>1</em>6 stable outpatients with HF, ejection fraction ≥50%, elevated N-terminal brain-type natriuretic peptide or elevated invasively measured filling pressures, and reduced exercise capacity. Participants were randomized from October 2008 through February 20<em>1</em>2 at 26 centers in North America. Follow-up was through August 30, 20<em>1</em>2.

METHODS

Sildenafil (n = <em>1</em><em>1</em>3) or placebo (n = <em>1</em>03) administered orally at 20 mg, 3 times daily for <em>1</em>2 weeks, followed by 60 mg, 3 times daily for <em>1</em>2 weeks.

METHODS

Primary end point was change in peak oxygen consumption after 24 weeks of therapy. Secondary end points included change in 6-minute walk distance and a hierarchical composite clinical status score (range, <em>1</em>-n, a higher value indicates better status; expected value with no treatment effect, 95) based on time to death, time to cardiovascular or cardiorenal hospitalization, and change in quality of life for participants without cardiovascular or cardiorenal hospitalization at 24 weeks.

RESULTS

Median age was 69 years, and 48% of patients were women. At baseline, median peak oxygen consumption (<em>1</em><em>1</em>.7 mL/kg/min) and 6-minute walk distance (308 m) were reduced. The median E/e' (<em>1</em>6), left atrial volume index (44 mL/m2), and pulmonary artery systolic pressure (4<em>1</em> mm Hg) were consistent with chronically elevated left ventricular filling pressures. At 24 weeks, median (IQR) changes in peak oxygen consumption (mL/kg/min) in patients who received placebo (-0.20 [IQR, -0.70 to <em>1</em>.00]) or sildenafil (-0.20 [IQR, -<em>1</em>.70 to <em>1</em>.<em>1</em><em>1</em>]) were not significantly different (P = .90) in analyses in which patients with missing week-24 data were excluded, and in sensitivity analysis based on intention to treat with multiple imputation for missing values (mean between-group difference, 0.0<em>1</em> mL/kg/min, [95% CI, -0.60 to 0.6<em>1</em>]). The mean clinical status rank score was not significantly different at 24 weeks between placebo (95.8) and sildenafil (94.2) (P = .85). Changes in 6-minute walk distance at 24 weeks in patients who received placebo (<em>1</em>5.0 m [IQR, -26.0 to 45.0]) or sildenafil (5.0 m [IQR, -37.0 to 55.0]; P = .92) were also not significantly different. Adverse events occurred in 78 placebo patients (76%) and 90 sildenafil patients (80%). Serious adverse events occurred in <em>1</em>6 placebo patients (<em>1</em>6%) and 25 sildenafil patients (22%).

CONCLUSIONS

Among patients with HFPEF, phosphodiesterase-5 inhibition with administration of sildenafil for 24 weeks, compared with placebo, did not result in significant improvement in exercise capacity or clinical status.

BACKGROUND

clinicaltrials.gov Identifier: NCT00763867.

BACKGROUND

Few published studies have combined clinical prognostic factors into risk profiles that can be used to predict the likelihood of recurrence or metastatic progression in patients following treatment of prostate cancer. We developed a nomogram that allows prediction of disease recurrence through use of preoperative clinical factors for patients with clinically localized prostate cancer who are candidates for treatment with a radical prostatectomy.

METHODS

By use of Cox proportional hazards regression analysis, we modeled the clinical data and disease follow-up for 983 men with clinically localized prostate cancer whom we intended to treat with a radical prostatectomy. Clinical data included pretreatment serum prostate-specific antigen levels, biopsy Gleason scores, and clinical stage. Treatment failure was recorded when there was clinical evidence of disease recurrence, a rising serum prostate-specific antigen level (two measurements of 0.4 ng/mL or greater and rising), or initiation of adjuvant therapy. Validation was performed on a separate sample of 168 men, also from our institution.

RESULTS

Treatment failure (i.e., cancer recurrence) was noted in 196 of the 983 men, and the patients without failure had a median follow-up of 30 months (range, 1-146 months). The 5-year probability of freedom from failure for the cohort was 73% (95% confidence interval = 69%-76%). The predictions from the nomogram appeared accurate and discriminating, with a validation sample area under the receiver operating characteristic curve (i.e., comparison of the predicted probability with the actual outcome) of 0.79.

CONCLUSIONS

A nomogram has been developed that can be used to predict the 5-year probability of treatment failure among men with clinically localized prostate cancer treated with radical prostatectomy.

Recombinant adeno-associated viral (rAAV) vectors based on serotype 2 are currently being evaluated most extensively in animals and human clinical trials. rAAV vectors constructed from other AAV serotypes (serotypes <em>1</em>, 3, 4, 5, and 6) can transduce certain tissues more efficiently and with different specificity than rAAV2 vectors in animal models. Here, we describe reagents and methods for the production and purification of AAV2 inverted terminal repeat-containing vectors pseudotyped with AAV<em>1</em> or AAV5 capsids. To facilitate pseudotyping, AAV2rep/AAV<em>1</em>cap and AAV2rep/AAV5cap helper plasmids were constructed in an adenoviral plasmid backbone. The resultant plasmids, pXYZ<em>1</em> and pXYZ5, were used to produce rAAV<em>1</em> and rAAV5 vectors, respectively, by transient transfection. Since neither AAV5 nor AAV<em>1</em> binds to the heparin affinity chromatography resin used to purify rAAV2 vectors, purification protocols were developed based on anion-exchange chromatography. The purified vector stocks are 99% pure with titers of <em>1</em> x <em>1</em>0(<em>1</em>2) to <em>1</em> x <em>1</em>0(<em>1</em>3)vector genomes/<em>ml</em>.

The absolute diversity of prokaryotes is widely held to be unknown and unknowable at any scale in any environment. However, it is not necessary to count every species in a community to estimate the number of different taxa therein. It is sufficient to estimate the area under the species abundance curve for that environment. Log-normal species abundance curves are thought to characterize communities, such as bacteria, which exhibit highly dynamic and random growth. Thus, we are able to show that the diversity of prokaryotic communities may be related to the ratio of two measurable variables: the total number of individuals in the community and the abundance of the most abundant members of that community. We assume that either the least abundant species has an abundance of <em>1</em> or Preston's canonical hypothesis is valid. Consequently, we can estimate the bacterial diversity on a small scale (oceans <em>1</em>60 per <em>ml</em>; soil 6,400-38,000 per g; sewage works 70 per <em>ml</em>). We are also able to speculate about diversity at a larger scale, thus the entire bacterial diversity of the sea may be unlikely to exceed 2 x <em>1</em>0(6), while a ton of soil could contain 4 x <em>1</em>0(6) different taxa. These are preliminary estimates that may change as we gain a greater understanding of the nature of prokaryotic species abundance curves. Nevertheless, it is evident that local and global prokaryotic diversity can be understood through species abundance curves and purely experimental approaches to solving this conundrum will be fruitless.

During acute human immunodeficiency virus type <em>1</em> (HIV-<em>1</em>) infection or after transfection of the tat gene, Tat protein is released into the cell culture supernatant. In this extracellular form, Tat stimulates both HIV-<em>1</em> gene expression and the growth of cells derived from Kaposi's sarcoma (KS) lesions of HIV-<em>1</em>-infected individuals (AIDS-KS cells). Tat protein and its biological activities appear in the cell supernatants at the peak of Tat expression, when the rate of cell death is low (infection) or cell death is undetectable (transfection) and increased levels of cytoplasmic Tat are present. Tat-containing supernatants stimulate maximal AIDS-KS cell growth but only low to moderate levels of HIV-<em>1</em> gene expression. This is due to the different concentrations of exogenous Tat required for the two effects. The cell growth-promoting effects of Tat peak at between 0.<em>1</em> and <em>1</em> ng of purified recombinant protein per <em>ml</em> in the cell growth medium and do not increase with concentration. In contrast, both the detection of nuclear-localized Tat taken up by cells and the induction of HIV-<em>1</em> gene expression or replication require higher Tat concentrations >> or = <em>1</em>00 ng/<em>ml</em>), and all increase linearly with increasing amounts of the exogenous protein. These data suggest that Tat can be released by a mechanism(s) other than cell death and that the cell growth-promoting activity and the virus-transactivating effect of extracellular Tat are mediated by different pathways.

A technique for the rapid production of large unilamellar vesicles by repeated extrusion under moderate pressures (≤ 500 lb/in²) of multilamellar vesicles through polycarbonate filters (<em>1</em>00 nm pore size) is demonstrated. In combination with freeze-thaw protocols where required, this procedure results in unilamellar vesicles with diameters in the range 60-<em>1</em>00 nm and with trapped volumes in the region of <em>1</em>-3 <em>μl</em>/μmol phospholipid. Advantages of this technique include the absence of organic solvents or detergents, the high lipid concentrations (up to 300 μmol/<em>ml</em>) that can be employed and the high trapping efficiencies (up to 30%) that can be achieved. Further, the procedure for generating these 'LUVET's' (large unilamellar vesicles by extrusion techniques) is rapid (≤ min preparation time) and can be employed to generate large unilamellar vesicles from a wide variety of lipid species and mixtures. As a particular illustration of the utility of this vesicle preparation, LUVET systems exhibiting a membrane potential (ΔΨ) in response to a transmembrane Na⁺/K⁺ gradient (K⁺ inside) have been characterized. By employing the lipophilic cation methyltriphenylphosphonium (MTPP⁺) it is shown that a K⁺ of diffusion potential (ΔΨ < -<em>1</em>00 mV) forms rapidly in the presence of the K⁺ ionophore valinomycin for soya phosphatidylcholine (soya PC) LUVET's. The values of Δψ obtained correlate well with the K⁺ concentration gradient across the membrane, and it is demonstrated that the decay of Δψ with time depends on the flux of Na+ into the vesicles.

Biomarker discovery produces lists of candidate markers whose presence and level must be subsequently verified in serum or plasma. Verification represents a paradigm shift from unbiased discovery approaches to targeted, hypothesis-driven methods and relies upon specific, quantitative assays optimized for the selective detection of target proteins. Many protein biomarkers of clinical currency are present at or below the nanogram/milliliter range in plasma and have been inaccessible to date by MS-based methods. Using multiple reaction monitoring coupled with stable isotope dilution mass spectrometry, we describe here the development of quantitative, multiplexed assays for six proteins in plasma that achieve limits of quantitation in the <em>1</em>-<em>1</em>0 ng/<em>ml</em> range with percent coefficients of variation from 3 to <em>1</em>5% without immunoaffinity enrichment of either proteins or peptides. Sample processing methods with sufficient throughput, recovery, and reproducibility to enable robust detection and quantitation of candidate biomarker proteins were developed and optimized by addition of exogenous proteins to immunoaffinity depleted plasma from a healthy donor. Quantitative multiple reaction monitoring assays were designed and optimized for signature peptides derived from the test proteins. Based upon calibration curves using known concentrations of spiked protein in plasma, we determined that each target protein had at least one signature peptide with a limit of quantitation in the <em>1</em>-<em>1</em>0 ng/<em>ml</em> range and linearity typically over 2 orders of magnitude in the measurement range of interest. Limits of detection were frequently in the high picogram/milliliter range. These levels of assay performance represent up to a <em>1</em>000-fold improvement compared with direct analysis of proteins in plasma by MS and were achieved by simple, robust sample processing involving abundant protein depletion and minimal fractionation by strong cation exchange chromatography at the peptide level prior to LC-multiple reaction monitoring/MS. The methods presented here provide a solid basis for developing quantitative MS-based assays of low level proteins in blood.

There have been few fundamental improvements in clinical X-ray contrast agents in more than 25 years, and the chemical platform of tri-iodobenzene has not changed. Current agents impose serious limitations on medical imaging: short imaging times, the need for catheterization in many cases, occasional renal toxicity, and poor contrast in large patients. This report is the first demonstration that gold nanoparticles may overcome these limitations. Gold has higher absorption than iodine with less bone and tissue interference achieving better contrast with lower X-ray dose. Nanoparticles clear the blood more slowly than iodine agents, permitting longer imaging times. Gold nanoparticles, <em>1</em>.9 nm in diameter, were injected intravenously into mice and images recorded over time with a standard mammography unit. Gold biodistribution was measured by atomic absorption. Retention in liver and spleen was low with elimination by the kidneys. Organs such as kidneys and tumours were seen with unusual clarity and high spatial resolution. Blood vessels less than <em>1</em>00 microm in diameter were delineated, thus enabling in vivo vascular casting. Regions of increased vascularization and angiogenesis could be distinguished. With <em>1</em>0 mg Au <em>ml</em>(-<em>1</em>) initially in the blood, mouse behaviour was unremarkable and neither blood plasma analytes nor organ histology revealed any evidence of toxicity <em>1</em><em>1</em> days and 30 days after injection. Gold nanoparticles can be used as X-ray contrast agents with properties that overcome some significant limitations of iodine-based agents.

Hepatitis C virus (HCV) has emerged as an important etiologic agent of liver injury and failure in patients infected with human immunodeficiency virus (HIV). The prevalence and characteristics of HCV in a representative cohort of HIV-infected patients have not been described. Therefore, a representative sample of <em>1</em>687 HIV-infected patients was studied; a 2<em>1</em>3-sample subcohort was selected by use of risk-based sampling from 2 large prospective US Adult AIDS Clinical Trials Group clinical trials. HCV prevalence, HCV RNA level, and genotype were determined. The weighted overall estimate of HCV prevalence in the study cohort was <em>1</em>6.<em>1</em>% (95% weighted confidence interval, <em>1</em>4.3%-<em>1</em>7.8%), with significant variability depending on risk factors and HIV RNA levels. Among patients defined as being "at risk", 72.7% were HCV positive, whereas, among low-risk patients, the positivity rate was 3.5%. Genotype <em>1</em> was found in 83.3% of infected patients. Median HCV RNA level was 6.08x<em>1</em>06 IU/<em>mL</em>. High virus loads and genotype <em>1</em> prevalence may be important to interferon-based antiviral response rates among coinfected patients.

Fluorescence in situ hybridization (FISH) with horseradish peroxidase (HRP)-labeled oligonucleotide probes and tyramide signal amplification, also known as catalyzed reporter deposition (CARD), is currently not generally applicable to heterotrophic bacteria in marine samples. Penetration of the HRP molecule into bacterial cells requires permeabilization procedures that cause high and most probably species-selective cell loss. Here we present an improved protocol for CARD-FISH of marine planktonic and benthic microbial assemblages. After concentration of samples onto membrane filters and subsequent embedding of filters in low-gelling-point agarose, no decrease in bacterial cell numbers was observed during 90 min of lysozyme incubation (<em>1</em>0 mg <em>ml</em>(-<em>1</em>) at 37 degrees C). The detection rates of coastal North Sea bacterioplankton by CARD-FISH with a general bacterial probe (EUB338-HRP) were significantly higher (mean, 94% of total cell counts; range, 85 to <em>1</em>00%) than that with a monolabeled probe (EUB338-mono; mean, 48%; range, <em>1</em>9 to 66%). Virtually no unspecific staining was observed after CARD-FISH with an antisense EUB338-HRP. Members of the marine SAR86 clade were undetectable by FISH with a monolabeled probe; however, a substantial population was visualized by CARD-FISH (mean, 7%; range, 3 to <em>1</em>3%). Detection rates of EUB338-HRP in Wadden Sea sediments (mean, 8<em>1</em>%; range, 53 to <em>1</em>00%) were almost twice as high as the detection rates of EUB338-mono (mean, 44%; range, 25 to 7<em>1</em>%). The enhanced fluorescence intensities and signal-to-background ratios make CARD-FISH superior to FISH with directly labeled oligonucleotides for the staining of bacteria with low rRNA content in the marine environment.

These recommendations represent the first statement by the Advisory Committee on Immunization Practices (ACIP) on the use of a live attenuated vaccine for the prevention of herpes zoster (zoster) (i.e., shingles) and its sequelae, which was licensed by the U.S. Food and Drug Administration (FDA) on May 25, 2006. This report summarizes the epidemiology of zoster and its sequelae, describes the zoster vaccine, and provides recommendations for its use among adults aged>> or =60 years in the United States. Zoster is a localized, generally painful cutaneous eruption that occurs most frequently among older adults and immunocompromised persons. It is caused by reactivation of latent varicella zoster virus (VZV) decades after initial VZV infection is established. Approximately one in three persons will develop zoster during their lifetime, resulting in an estimated <em>1</em> million episodes in the United States annually. A common complication of zoster is postherpetic neuralgia (PHN), a chronic, often debilitating pain condition that can last months or even years. The risk for PHN in patients with zoster is <em>1</em>0%-<em>1</em>8%. Another complication of zoster is eye involvement, which occurs in <em>1</em>0%-25% of zoster episodes and can result in prolonged or permanent pain, facial scarring, and loss of vision. Approximately 3% of patients with zoster are hospitalized; many of these episodes involved persons with one or more immunocompromising conditions. Deaths attributable to zoster are uncommon among persons who are not immunocompromised. Prompt treatment with the oral antiviral agents acyclovir, valacyclovir, and famciclovir decreases the severity and duration of acute pain from zoster. Additional pain control can be achieved in certain patients by supplementing antiviral agents with corticosteroids and with analgesics. Established PHN can be managed in certain patients with analgesics, tricyclic antidepressants, and other agents. Licensed zoster vaccine is a lyophilized preparation of a live, attenuated strain of VZV, the same strain used in the varicella vaccines. However, its minimum potency is at least <em>1</em>4-times the potency of single-antigen varicella vaccine. In a large clinical trial, zoster vaccine was partially efficacious at preventing zoster. It also was partially efficacious at reducing the severity and duration of pain and at preventing PHN among those developing zoster. Zoster vaccine is recommended for all persons aged>> or =60 years who have no contraindications, including persons who report a previous episode of zoster or who have chronic medical conditions. The vaccine should be offered at the patient's first clinical encounter with his or her health-care provider. It is administered as a single 0.65 <em>mL</em> dose subcutaneously in the deltoid region of the arm. A booster dose is not licensed for the vaccine. Zoster vaccination is not indicated to treat acute zoster, to prevent persons with acute zoster from developing PHN, or to treat ongoing PHN. Before administration of zoster vaccine, patients do not need to be asked about their history of varicella (chickenpox) or to have serologic testing conducted to determine varicella immunity.

BACKGROUND

Limited evidence exists to show that adding a third agent to platinum-doublet chemotherapy improves efficacy in the first-line advanced non-small-cell lung cancer (NSCLC) setting. The anti-PD-<em>1</em> antibody pembrolizumab has shown efficacy as monotherapy in patients with advanced NSCLC and has a non-overlapping toxicity profile with chemotherapy. We assessed whether the addition of pembrolizumab to platinum-doublet chemotherapy improves efficacy in patients with advanced non-squamous NSCLC.

METHODS

In this randomised, open-label, phase 2 cohort of a multicohort study (KEYNOTE-02<em>1</em>), patients were enrolled at 26 medical centres in the USA and Taiwan. Patients with chemotherapy-naive, stage IIIB or IV, non-squamous NSCLC without targetable EGFR or ALK genetic aberrations were randomly assigned (<em>1</em>:<em>1</em>) in blocks of four stratified by PD-L<em>1</em> tumour proportion score ((<em>1</em>% vs ≥<em>1</em>%) using an interactive voice-response system to 4 cycles of pembrolizumab 200 mg plus carboplatin area under curve 5 mg/mL per min and pemetrexed 500 mg/m2 every 3 weeks followed by pembrolizumab for 24 months and indefinite pemetrexed maintenance therapy or to 4 cycles of carboplatin and pemetrexed alone followed by indefinite pemetrexed maintenance therapy. The primary endpoint was the proportion of patients who achieved an objective response, defined as the percentage of patients with radiologically confirmed complete or partial response according to Response Evaluation Criteria in Solid Tumors version <em>1</em>.<em>1</em> assessed by masked, independent central review, in the intention-to-treat population, defined as all patients who were allocated to study treatment. Significance threshold was p<0·025 (one sided). Safety was assessed in the as-treated population, defined as all patients who received at least one dose of the assigned study treatment. This trial, which is closed for enrolment but continuing for follow-up, is registered with ClinicalTrials.gov, number NCT02039674.

RESULTS

Between Nov 25, 20<em>1</em>4, and Jan 25, 20<em>1</em>6, <em>1</em>23 patients were enrolled; 60 were randomly assigned to the pembrolizumab plus chemotherapy group and 63 to the chemotherapy alone group. 33 (55%; 95% CI 42-68) of 60 patients in the pembrolizumab plus chemotherapy group achieved an objective response compared with <em>1</em>8 (29%; <em>1</em>8-4<em>1</em>) of 63 patients in the chemotherapy alone group (estimated treatment difference 26% [95% CI 9-42%]; p=0·00<em>1</em>6). The incidence of grade 3 or worse treatment-related adverse events was similar between groups (23 [39%] of 59 patients in the pembrolizumab plus chemotherapy group and <em>1</em>6 [26%] of 62 in the chemotherapy alone group). The most common grade 3 or worse treatment-related adverse events in the pembrolizumab plus chemotherapy group were anaemia (seven [<em>1</em>2%] of 59) and decreased neutrophil count (three [5%]); an additional six events each occurred in two (3%) for acute kidney injury, decreased lymphocyte count, fatigue, neutropenia, and sepsis, and thrombocytopenia. In the chemotherapy alone group, the most common grade 3 or worse events were anaemia (nine [<em>1</em>5%] of 62) and decreased neutrophil count, pancytopenia, and thrombocytopenia (two [3%] each). One (2%) of 59 patients in the pembrolizumab plus chemotherapy group experienced treatment-related death because of sepsis compared with two (3%) of 62 patients in the chemotherapy group: one because of sepsis and one because of pancytopenia.

CONCLUSIONS

Combination of pembrolizumab, carboplatin, and pemetrexed could be an effective and tolerable first-line treatment option for patients with advanced non-squamous NSCLC. This finding is being further explored in an ongoing international, randomised, double-blind, phase 3 study.

BACKGROUND

Merck & Co.

Interleukin (IL)-<em>1</em>beta is a proinflammatory cytokine implicated in various pathophysiological conditions of the CNS involving NMDA receptor activation. Circumstantial evidence suggests that IL-<em>1</em>beta and NMDA receptors can functionally interact. Using primary cultures of rat hippocampal neurons, we investigated whether IL-<em>1</em>beta affects NMDA receptor function(s) by studying (<em>1</em>) NMDA receptor-induced [Ca2+]i increase and (2) NMDA-mediated neurotoxicity. IL<em>1</em>beta (0.0<em>1</em>-0.<em>1</em> ng/<em>ml</em>) dose-dependently enhances NMDA-induced [Ca2+]i increases with a maximal effect of approximately 45%. This effect occurred only when neurons were pretreated with IL-<em>1</em>beta, whereas it was absent if IL-<em>1</em>beta and NMDA were applied simultaneously, and it was abolished by IL-<em>1</em> receptor antagonist (50 ng/<em>ml</em>). Facilitation of NMDA-induced [Ca2+]i increase by IL-<em>1</em>beta was prevented by both lavendustin (LAV) A (500 nm) and 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) (<em>1</em> microm), suggesting an involvement of tyrosine kinases. Increased tyrosine phosphorylation of NMDA receptor subunits 2A and 2B and coimmunoprecipitation of activated Src tyrosine kinase with these subunits was observed after exposure of hippocampal neurons to 0.05 ng/<em>ml</em> IL-<em>1</em>beta. Finally, 0.05 ng/<em>ml</em> IL-<em>1</em>beta increased by approximately 30% neuronal cell death induced by NMDA, and this effect was blocked by both lavendustin A and PP2. These data suggest that IL-<em>1</em>beta increases NMDA receptor function through activation of tyrosine kinases and subsequent NR2A/B subunit phosphorylation. These effects may contribute to glutamate-mediated neurodegeneration.

BACKGROUND

Obstructive sleep apnea syndrome (OSAS), characterized by intermittent hypoxia/reoxygenation (IHR), is an independent risk factor for cardiovascular disease. We investigated the underlying molecular mechanisms of this association in a translational study.

RESULTS

In a novel in vitro model of IHR, we used HeLa cells transfected with reporter constructs and DNA binding assays for the master transcriptional regulators of the inflammatory and adaptive pathways (NFkappaB and HIF-<em>1</em>, respectively) to investigate underlying transcriptional events initiated by repeated cell exposure to IHR. Furthermore, we prospectively studied <em>1</em>9 male OSAS patients (median apnea-hypopnea frequency, 48.5 episodes per hour; interquartile range [IQR], 28.5 to 72.9) and <em>1</em>7 matched normal control subjects. Circulating levels of the proinflammatory cytokine tumor necrosis factor-alpha and the adaptive factor erythropoietin were assayed in all subjects at baseline and again after 6 weeks of continuous positive airway pressure therapy in patients. Full blood count was measured as part of a detailed baseline evaluation. HeLa cells exposed to IHR demonstrated selective activation of the proinflammatory transcription factor NFkappaB (P<0.00<em>1</em> by ANOVA), whereas the adaptive regulator HIF-<em>1</em> was not activated, as demonstrated by luciferase reporter assays and DNA binding studies. Circulating tumor necrosis factor-alpha levels were higher in OSAS patients (2.56 pg/<em>mL</em>; IQR, 2.0<em>1</em> to 3.42 pg/<em>mL</em>) than in control subjects (<em>1</em>.25 pg/<em>mL</em>; IQR, 0.94 to <em>1</em>.87; P<0.00<em>1</em>) but normalized with continuous positive airway pressure therapy (<em>1</em>.24 pg/<em>mL</em>; IQR, 0.78 to 2.35 pg/<em>mL</em>; P=0.002). In contrast, erythropoietin levels were similar throughout. Furthermore, circulating neutrophil levels were higher in OSAS patients than in control subjects, whereas the hematocrit was unaltered.

CONCLUSIONS

These data demonstrate selective activation of inflammatory over adaptive pathways in IHR and OSAS, which may be an important molecular mechanism of cardiovascular disease.

<strong class="sub-title"> Background: </strong> A heterologous recombinant adenovirus (rAd)-based vaccine, Gam-COVID-Vac (Sputnik V), showed a good safety profile and induced strong humoral and cellular immune responses in participants in phase <em>1</em>/2 clinical trials. Here, we report preliminary results on the efficacy and safety of Gam-COVID-Vac from the interim analysis of this phase 3 trial.

<strong class="sub-title"> Methods: </strong> We did a randomised, double-blind, placebo-controlled, phase 3 trial at 25 hospitals and polyclinics in Moscow, Russia. We included participants aged at least <em>1</em>8 years, with negative SARS-CoV-2 PCR and IgG and IgM tests, no infectious diseases in the <em>1</em>4 days before enrolment, and no other vaccinations in the 30 days before enrolment. Participants were randomly assigned (3:<em>1</em>) to receive vaccine or placebo, with stratification by age group. Investigators, participants, and all study staff were masked to group assignment. The vaccine was administered (0·5 mL/dose) intramuscularly in a prime-boost regimen: a 2<em>1</em>-day interval between the first dose (rAd26) and the second dose (rAd5), both vectors carrying the gene for the full-length SARS-CoV-2 glycoprotein S. The primary outcome was the proportion of participants with PCR-confirmed COVID-<em>1</em>9 from day 2<em>1</em> after receiving the first dose. All analyses excluded participants with protocol violations: the primary outcome was assessed in participants who had received two doses of vaccine or placebo, serious adverse events were assessed in all participants who had received at least one dose at the time of database lock, and rare adverse events were assessed in all participants who had received two doses and for whom all available data were verified in the case report form at the time of database lock. The trial is registered at ClinicalTrials.gov (<a href="http://clinicaltrials.gov/show/NCT04530396" title="See in ClinicalTrials.gov">NCT04530396</a>).

<strong class="sub-title"> Findings: </strong> Between Sept 7 and Nov 24, 2020, 2<em>1</em> 977 adults were randomly assigned to the vaccine group (n=<em>1</em>6 50<em>1</em>) or the placebo group (n=5476). <em>1</em>9 866 received two doses of vaccine or placebo and were included in the primary outcome analysis. From 2<em>1</em> days after the first dose of vaccine (the day of dose 2), <em>1</em>6 (0·<em>1</em>%) of <em>1</em>4 964 participants in the vaccine group and 62 (<em>1</em>·3%) of 4902 in the placebo group were confirmed to have COVID-<em>1</em>9; vaccine efficacy was 9<em>1</em>·6% (95% CI 85·6-95·2). Most reported adverse events were grade <em>1</em> (7485 [94·0%] of 7966 total events). 45 (0·3%) of <em>1</em>6 427 participants in the vaccine group and 23 (0·4%) of 5435 participants in the placebo group had serious adverse events; none were considered associated with vaccination, with confirmation from the independent data monitoring committee. Four deaths were reported during the study (three [<0·<em>1</em>%] of <em>1</em>6 427 participants in the vaccine group and one [<0·<em>1</em>%] of 5435 participants in the placebo group), none of which were considered related to the vaccine.

<strong class="sub-title"> Interpretation: </strong> This interim analysis of the phase 3 trial of Gam-COVID-Vac showed 9<em>1</em>·6% efficacy against COVID-<em>1</em>9 and was well tolerated in a large cohort.

Funding: Moscow City Health Department, Russian Direct Investment Fund, Sberbank, and RUSAL.

BACKGROUND

Continuous veno-venous haemofiltration is increasingly used to treat acute renal failure in critically ill patients, but a clear definition of an adequate treatment dose has not been established. We undertook a prospective randomised study of the impact different ultrafiltration doses in continuous renal replacement therapy on survival.

METHODS

We enrolled 425 patients, with a mean age of 6<em>1</em> years, in intensive care who had acute renal failure. Patients were randomly assigned ultrafiltration at 20 <em>mL</em> h(-<em>1</em>) kg(-<em>1</em>) (group <em>1</em>, n=<em>1</em>46), 35 <em>mL</em> h(-<em>1</em>) kg(-<em>1</em>) (group 2, n=<em>1</em>39), or 45 <em>mL</em> h(-<em>1</em>) kg(-<em>1</em>) (group 3, n=<em>1</em>40). The primary endpoint was survival at <em>1</em>5 days after stopping haemofiltration. We also assessed recovery of renal function and frequency of complications during treatment. Analysis was by intention to treat.

RESULTS

Survival in group <em>1</em> was significantly lower than in groups 2 (p=0.0007) and 3 (p=0.00<em>1</em>3). Survival in groups 2 and 3 did not differ significantly (p=0.87). Adjustment for possible confounding factors did not change the pattern of differences among the groups. Survivors in all groups had lower concentrations of blood urea nitrogen before continuous haemofiltration was started than non-survivors. 95%, 92%, and 90% of survivors in groups <em>1</em>, 2, and 3, respectively, had full recovery of renal function. The frequency of complications was similarly low in all groups.

CONCLUSIONS

Mortality among these critically ill patients was high, but increase in the rate of ultrafiltration improved survival significantly. We recommend that ultrafiltration should be prescribed according to patient's bodyweight and should reach at least 35 <em>mL</em> h(-<em>1</em>) kg(-<em>1</em>).

The past decade, particularly the last <em>1</em>8 months, witnessed a vigorous increase in interest in vitamin D from both the lay and biomedical worlds. Much of the growing interest in vitamin D is powered by new data being extracted from the National Health and Nutrition Examination Survey (NHANES). The newest statistics demonstrate that more than 90% of the pigmented populace of the United States (Blacks, Hispanics, and Asians) now suffer from vitamin D insufficiency (25-hydroxyvitamin D <30 ng/<em>ml</em>), with nearly three fourths of the white population in this country also being vitamin D insufficient. This represents a near doubling of the prevalence of vitamin D insufficiency seen just <em>1</em>0 yr ago in the same population. This review attempts to provide some explanation for: <em>1</em>) the rapid decline in vitamin D status in the United States; 2) the adverse impact of vitamin D insufficiency on skeletal, infectious/inflammatory, and metabolic health in humans; and 3) the therapeutic rationale and reliable means for vigorous supplementation of our diets with vitamin D.

BACKGROUND

A key feature of chronic obstructive pulmonary disease (COPD) is an accelerated rate of decline in forced expiratory volume in <em>1</em> second (FEV(<em>1</em>)), but data on the variability and determinants of this change in patients who have established disease are scarce.

METHODS

We analyzed the changes in FEV(<em>1</em>) after administration of a bronchodilator over a 3-year period in 2<em>1</em>63 patients. A random-coefficient model was used to evaluate possible predictors of both FEV(<em>1</em>) levels and their changes over time.

RESULTS

The mean (±SE) rate of change in FEV(<em>1</em>) was a decline of 33±2 ml per year, with significant variation among the patients studied. The between-patient standard deviation for the rate of decline was 59 ml per year. Over the 3-year study period, 38% of patients had an estimated decline in FEV(<em>1</em>) of more than 40 ml per year, 3<em>1</em>% had a decline of 2<em>1</em> to 40 ml per year, 23% had a change in FEV(<em>1</em>) that ranged from a decrease of 20 ml per year to an increase of 20 ml per year, and 8% had an increase of more than 20 ml per year. The mean rate of decline in FEV(<em>1</em>) was 2<em>1</em>±4 ml per year greater in current smokers than in current nonsmokers, <em>1</em>3±4 ml per year greater in patients with emphysema than in those without emphysema, and <em>1</em>7±4 ml per year greater in patients with bronchodilator reversibility than in those without reversibility.

CONCLUSIONS

The rate of change in FEV(<em>1</em>) among patients with COPD is highly variable, with increased rates of decline among current smokers, patients with bronchodilator reversibility, and patients with emphysema.

The antiviral antibiotic brefeldin A (BFA) strongly inhibits the protein secretion in cultured rat hepatocytes (Misumi, Y., Misumi, Y., Miki, K., Takatsuki, A., Tamura, G., and Ikehara, Y. (<em>1</em>986) J. Biol. Chem. 26<em>1</em>, <em>1</em><em>1</em>398-<em>1</em><em>1</em>403). We have further examined the inhibitory effect of the drug on intracellular transport of albumin by an immunocytochemical technique with peroxidase-conjugated Fab fragments of anti-rat albumin IgG. In hepatocytes treated with BFA (2.5 micrograms/<em>ml</em>) for <em>1</em> h at 37 degrees C, no characteristic structures of the Golgi complex could be observed, and albumin was diffusely distributed in the endoplasmic reticulum (ER), nuclear envelope, and small vesicles around, in contrast to its condensed localization in the Golgi complex in the control cells. Such an unusual distribution of the secretory protein, however, was rearranged to the normal localization in the Golgi complex after 4 h even in the presence of the drug, possibly due to a metabolism of the drug to an inert form. Exposure of the cells to BFA with constant renewals (2.5 micrograms/<em>ml</em> at <em>1</em>-h intervals) or at a higher concentration (<em>1</em>0 micrograms/<em>ml</em>) caused a prolonged accumulation of albumin in the ER, resulting in its dilation. These results indicate that BFA primarily blocks the protein transport from the ER to the Golgi complex, consistent with the biochemical data previously reported.

Mutations in growth signaling pathways extend life span, as well as protect against age-dependent DNA damage in yeast and decrease insulin resistance and cancer in mice. To test their effect in humans, we monitored for 22 years Ecuadorian individuals who carry mutations in the growth hormone receptor (GHR) gene that lead to severe GHR and IGF-<em>1</em> (insulin-like growth factor-<em>1</em>) deficiencies. We combined this information with surveys to identify the cause and age of death for individuals in this community who died before this period. The individuals with GHR deficiency exhibited only one nonlethal malignancy and no cases of diabetes, in contrast to a prevalence of <em>1</em>7% for cancer and 5% for diabetes in control subjects. A possible explanation for the very low incidence of cancer was suggested by in vitro studies: Serum from subjects with GHR deficiency reduced DNA breaks but increased apoptosis in human mammary epithelial cells treated with hydrogen peroxide. Serum from GHR-deficient subjects also caused reduced expression of RAS, PKA (protein kinase A), and TOR (target of rapamycin) and up-regulation of SOD2 (superoxide dismutase 2) in treated cells, changes that promote cellular protection and life-span extension in model organisms. We also observed reduced insulin concentrations (<em>1</em>.4 μU/<em>ml</em> versus 4.4 μU/<em>ml</em> in unaffected relatives) and a very low HOMA-IR (homeostatic model assessment-insulin resistance) index (0.34 versus 0.96 in unaffected relatives) in individuals with GHR deficiency, indicating higher insulin sensitivity, which could explain the absence of diabetes in these subjects. These results provide evidence for a role of evolutionarily conserved pathways in the control of aging and disease burden in humans.

OBJECTIVE

Multipotential mesenchymal stem cells (MSCs) are found in human bone marrow and are shown to secrete hematopoietic cytokines and support hematopoietic progenitors in vitro. We hypothesized that infusion of autologous MSCs after myeloablative therapy would facilitate engraftment by hematopoietic stem cells, and we investigated the feasibility, safety, and hematopoietic effects of culture-expanded MSCs in breast cancer patients receiving autologous peripheral-blood progenitor-cell (PBPC) infusion.

METHODS

We developed an efficient method of isolating and culture-expanding a homogenous population of MSCs from a small marrow-aspirate sample obtained from 32 breast cancer patients. Twenty-eight patients were given high-dose chemotherapy and autologous PBPCs plus culture-expanded MSC infusion and daily granulocyte colony-stimulating factor.

RESULTS

Human MSCs were successfully isolated from a mean +/- SD of 23.4 +/- 5.9 <em>mL</em> of bone marrow aspirate from all patients. Expansion cultures generated greater than <em>1</em> x <em>1</em>0(6) MSCs/kg for all patients over 20 to 50 days with a mean potential of 5.6 to 36.3 x <em>1</em>0(6) MSCs/kg after two to six passages, respectively. Twenty-eight patients were infused with <em>1</em> to 2.2 x <em>1</em>0(6) expanded autologous MSCs/kg intravenously over <em>1</em>5 minutes. There were no toxicities related to the infusion of MSCs. Clonogenic MSCs were detected in venous blood up to <em>1</em> hour after infusion in <em>1</em>3 of 2<em>1</em> patients (62%). Median time to achieve a neutrophil count greater than 500/microL and platelet count>>/= 20,000/microL untransfused was 8 days (range, 6 to <em>1</em><em>1</em> days) and 8.5 days (range, 4 to <em>1</em>9 days), respectively.

CONCLUSIONS

This report is the first describing infusion of autologous MSCs with therapeutic intent. We found that autologous MSC infusion at the time of PBPC transplantation is feasible and safe. The observed rapid hematopoietic recovery suggests that MSC infusion after myeloablative therapy may have a positive impact on hematopoiesis and should be tested in randomized trials.

OBJECTIVE

White matter lesions are often seen on MR scans of elderly non-demented and demented people. They are attributed to degenerative changes of small vessels and are implicated in the pathogenesis of cognitive decline and dementia. There is evidence that especially periventricular white matter lesions are related to cognitive decline, whereas subcortical white matter lesions may be related to late onset depression. The frequency distribution of subcortical and periventricular white matter lesions according to age and sex reported.

METHODS

A total of <em>1</em>077 subjects aged between 60-90 years were rando<em>ml</em>y sampled from the general population. All subjects underwent <em>1</em>.5T MR scanning; white matter lesions were rated separately for the subcortical region and the periventricular region.

RESULTS

Of all subjects 8% were completely free of subcortical white matter lesions, 20% had no periventricular white matter lesions, and 5% had no white matter lesions in either of these locations. The proportion with white matter lesions increased with age, similarly for men and women. Women tended to have more subcortical white matter lesions than men (total volume <em>1</em>.45 <em>ml</em> v <em>1</em>. 29 <em>ml</em>; p=0.33), mainly caused by marked differences in the frontal white matter lesion volume (0.89 <em>ml</em> v 0.70 <em>ml</em>; p=0.08). Periventricular white matter lesions were also more frequent among women than men (mean grade 2.5 v 2.3; p=0.07). Also severe degrees of subcortical white matter lesions were more common in women than in men (OR <em>1</em>.<em>1</em>; 95% confidence interval (95% CI) 0.8-<em>1</em>.5) and periventricular white matter lesions (OR <em>1</em>.2; 95% CI 0.9-<em>1</em>.7), albeit that none of these findings were statistically significant.

CONCLUSIONS

The prevalence and the degree of cerebral white matter lesions increased with age. Women tended to have a higher degree of white matter lesions than men. This may underlie the finding of a higher incidence of dementia in women than in men, particularly at later age.

OBJECTIVE

This study assesses the safety, pharmacokinetics and efficacy of transarterial chemoembolization using drug eluting beads (DEB), an embolizing device that slowly releases chemotherapy to decrease systemic toxicity.

METHODS

Twenty-seven Child-Pugh A cirrhotics (76% male, 59% HCV) with untreated large/multifocal HCC received chemoembolization with doxorubicin loaded DEBs at doses adjusted for bilirubin and body surface (range: 47-<em>1</em>50 mg). Clinical and analytical data were recorded at 24 and 48 h, 7, <em>1</em>4 and 30 days after first and second TACE. Response rate was assessed by CT at 6 months. Blood samples were obtained in <em>1</em>3 patients at 5, 20, 40, 60, <em>1</em>20 min, 6, 24, 48 and <em>1</em>68 h to determine doxorubicin Cmax and AUC.

RESULTS

DEB-TACE was well tolerated with an acceptable safety profile. Two cases developed liver abscess, one leading to death. Response rate was 75% (66.6% on intention-to-treat). Doxorubicin Cmax and AUC were significantly lower in DEB-TACE patients (78.97+/-38.3 ng/<em>mL</em> and 662.6+/-4<em>1</em>7.6 ng/<em>mL</em>min) than in conventional TACE (234<em>1</em>.5+/-395<em>1</em>.9 ng/<em>mL</em> and <em>1</em>8<em>1</em>2.2+/-<em>1</em>093.7 ng/<em>mL</em>min, p=0.00002 and p=0.00<em>1</em>, respectively). After a median follow-up of 27.6 months, <em>1</em>- and 2-year survival is 92.5% and 88.9%, respectively.

CONCLUSIONS

Chemoembolization using DEBs is an effective procedure with a favorable pharmacokinetic profile.