Efficacy and safety of pembrolizumab in metastatic urothelial carcinoma: results from KEYNOTE-045 and KEYNOTE-052 after up to 5 years of follow-up.

BACKGROUND: Immune checkpoint inhibitors are a standard therapy in metastatic urothelial carcinoma (UC). Long-term follow-up is necessary to confirm durability of response and identify further safety concerns. PATIENTS AND METHODS: In KEYNOTE-045, patients with metastatic UC that progressed on platinum-containing chemotherapy were randomly assigned 1:1 to receive pembrolizumab or investigator's choice of paclitaxel, docetaxel, or vinflunine. Primary endpoints were progression-free survival per RECIST version 1.1 by blinded independent central review (BICR) and overall survival. In KEYNOTE-052, cisplatin-ineligible patients with metastatic UC received first-line pembrolizumab. The primary endpoint was objective response rate per RECIST version 1.1 by BICR. RESULTS: A total of 542 patients (pembrolizumab, n = 270; chemotherapy, n = 272) were randomly assigned in KEYNOTE-045. The median follow-up was 62.9 months (range 58.6-70.9 months; data cut-off 1 October 2020). At 48 months, overall survival rates were 16.7% for pembrolizumab and 10.1% for chemotherapy; progression-free survival rates were 9.5% and 2.7%, respectively. The median duration of response (DOR) was 29.7 months (range 1.6+ to 60.5+ months) for pembrolizumab and 4.4 months (range 1.4+ to 63.1+ months) for chemotherapy; 36-month DOR rates were 44.4% and 28.3%, respectively. A total of 370 patients were enrolled in KEYNOTE-052. The median follow-up was 56.3 months (range 51.2-65.3 months; data cut-off 26 September 2020). The confirmed objective response rate was 28.9% (95% confidence interval 24.3-33.8), and the median DOR was 33.4 months (range 1.4+ to 60.7+ months); the 36-month DOR rate was 44.8%. Most treatment-related adverse events for pembrolizumab in either study were grade 1 or 2 and manageable, which is consistent with prior reports. CONCLUSION: With ∼5 years of follow-up, pembrolizumab monotherapy continued to demonstrate durable efficacy with no new safety signals in patients with platinum-resistant metastatic UC and as first-line therapy in cisplatin-ineligible patients. CLINICAL TRIAL REGISTRY AND ID: With ClinicalTrials.gov NCT02256436 (KEYNOTE-045); https://clinicaltrials.gov/ct2/show/NCT02256436 and NCT02335424 (KEYNOTE-052); https://clinicaltrials.gov/ct2/show/NCT02335424.

Consensus on the management of complications of cirrhosis of the liver in pediatrics.

The Asociación Mexicana de Hepatología A.C. carried out the Consensus on the Management of Complications of Cirrhosis of the Liver in Pediatrics to provide physicians with useful information for treating said complications. A group of pediatric gastroenterologists and experts in nutrition, nephrology, and infectious diseases participated and reviewed the medical literature. The Delphi method was applied to obtain the level of agreement on the statements that were formulated. The statements were sent to the participants to be analyzed and voted upon, after which they were discussed in virtual sessions, and the final versions were produced. The aim of the consensus results was to issue indications for the management of pediatric patients with liver cirrhosis, to prevent or control complications.

Factors associated with oropharyngeal dysphagia diagnosed by videofluoroscopy in children with cerebral palsy.

INTRODUCTION AND AIMS: Oropharyngeal dysphagia (OD) occurs in children with cerebral palsy. It is important to investigate its relationship with some variables, and the objective of this study was to identify factors associated with OD. MATERIALS AND METHODS: Case-control study in patients with cerebral palsy from 8months to 15years of age, from November 2018 to November 2019, approved by the Ethics Committee. The diagnosis of OD was made by videofluoroscopy when there was nasopharyngeal reflux, stagnation in the vallecular sinuses, in the piriformis sinuses, penetration, and aspiration. The independent variables were type of cerebral palsy, gross motor impairment classified into five levels, nutritional status and comorbidities. One case with OD was included and the next one without alterations in videofluoroscopy was control. The variables were compared with Chi square and Student's t. The association was measured with odds ratio. The confidence interval was 95%. RESULTS: Thirty patients with OD and 30without OD were studied. Sex, age, birth weight, and gestational age had a similar distribution in the two groups. From the data perceived by the mothers at the time of feeding, the greater frequency of the difficulty in the transfer of the food bolus in the group with OD showed a statistically significant difference (P<.001) and of the studied factors, the levelV of the gross motor involvement was associated with a higher frequency of OD. CONCLUSIONS: OD was associated with level V of gross motor involvement.

Induction of periodontal disease via retentive ligature, lipopolysaccharide injection, and their combination in a rat model.

Periodontitis is a highly prevalent, chronic immune-inflammatory disease of the periodontium that results in the periodontium and alveolar bone loss's progressive destruction. In this study, the induction of periodontal disease via retentive ligature, lipopolysaccharide, and their combination at three different times were compared in a rat model. Seventy-two Sprague Dawley rats were distributed into four treatment groups: 1) control group with no treatment; 2) application of 4/0 nylon ligature around second maxillary molars; 3) combination of ligature and LPS injection (ligature-LPS); 4) intragingival injection of Porphyromonas gingivalis lipopolysaccharide (Pg-LPS) to the palatal mucosa of the second maxillary molars. Six rats were sacrificed from each group after 7, 14, and 30 days of periodontal disease induction. Alveolar bone loss, attachment loss, number of inflammatory cells, and blood vessels were evaluated histologically. A micro-CT scan was used as a parameter to know the rate of alveolar bone loss. Parametric data were analyzed using two-way ANOVA followed by Bonferroni correction with a significance set at 5%. Non-parametric data were analyzed using Kruskal-Wallis, followed by multiple comparisons with Bonferroni correction. The histological results revealed significant destructive changes in the periodontal tissues and alveolar bone following the ligature and ligature-LPS induction techniques. These changes were evident as early as seven days, maintained until 14 days post-treatment, and declined with time. The ligature technique was effective in inducing acute periodontal disease. The LPS injection technique did not induce alveolar bone loss, and its combination to ligature added insignificant effects.

Numerical optimization of microfluidic vortex shedding for genome editing T cells with Cas9.

Microfluidic vortex shedding (µVS) can rapidly deliver mRNA to T cells with high yield and minimal perturbation of the cell state. The mechanistic underpinning of µVS intracellular delivery remains undefined and µVS-Cas9 genome editing requires further studies. Herein, we evaluated a series of µVS devices containing splitter plates to attenuate vortex shedding and understand the contribution of computed force and frequency on efficiency and viability. We then selected a µVS design to knockout the expression of the endogenous T cell receptor in primary human T cells via delivery of Cas9 ribonucleoprotein (RNP) with and without brief exposure to an electric field (eµVS). µVS alone resulted in an equivalent yield of genome-edited T cells relative to electroporation with improved cell quality. A 1.8-fold increase in editing efficiency was demonstrated with eµVS with negligible impact on cell viability. Herein, we demonstrate efficient processing of 5 × 106 cells suspend in 100 µl of cGMP OptiMEM in under 5 s, with the capacity of a single device to process between 106 to 108 in 1 to 30 s. Cumulatively, these results demonstrate the rapid and robust utility of µVS and eµVS for genome editing human primary T cells with Cas9 RNPs.

Ibrutinib in combination with nab-paclitaxel and gemcitabine for first-line treatment of patients with metastatic pancreatic adenocarcinoma: phase III RESOLVE study.

BACKGROUND: First-line treatment of metastatic pancreatic ductal adenocarcinoma (PDAC) includes nab-paclitaxel/gemcitabine. Ibrutinib, a Bruton's tyrosine kinase inhibitor, exhibits antitumor activity through tumor microenvironment modulation. The safety and efficacy of first-line ibrutinib plus nab-paclitaxel/gemcitabine treatment in patients with PDAC were evaluated. PATIENTS AND METHODS: RESOLVE (NCT02436668) was a phase III, randomized, double-blind, placebo-controlled study. Patients (histologically-confirmed PDAC; stage IV diagnosis ≥6 weeks of randomization; Karnofsky performance score ≥70) were randomized to once-daily oral ibrutinib (560 mg) or placebo plus nab-paclitaxel (125 mg/m2) and gemcitabine (1000 mg/m2). Primary endpoints were overall survival (OS) and investigator-assessed progression-free survival (PFS); overall response rate and safety were assessed. RESULTS: In total, 424 patients were randomized (ibrutinib arm, n = 211; placebo arm, n = 213). Baseline characteristics were balanced across arms. After a median follow-up of 25 months, there was no significant difference in OS between ibrutinib plus nab-paclitaxel/gemcitabine versus placebo plus nab-paclitaxel/gemcitabine (median of 9.7 versus 10.8 months; P = 0.3225). PFS was shorter for ibrutinib plus nab-paclitaxel/gemcitabine compared with placebo plus nab-paclitaxel/gemcitabine (median 5.3 versus 6.0 months; P < 0.0001). Overall response rates were 29% and 42%, respectively (P = 0.0058). Patients in the ibrutinib arm had less time on treatment and received lower cumulative doses for all agents compared with the placebo arm. The most common grade ≥3 adverse events for ibrutinib versus placebo arms included neutropenia (24% versus 35%), peripheral sensory neuropathy (17% versus 8%), and anemia (16% versus 17%). Primary reasons for any treatment discontinuation were disease progression and adverse events. CONCLUSIONS: Ibrutinib plus nab-paclitaxel/gemcitabine did not improve OS or PFS for patients with PDAC. Safety was consistent with known profiles for these agents.

Factors associated with oropharyngeal dysphagia diagnosed by videofluoroscopy in children with cerebral palsy. / Factores asociados a disfagia orofaríngea diagnosticada por videofluoroscopia en niños con parálisis cerebral.

INTRODUCTION AND AIMS: Oropharyngeal dysphagia (OD) occurs in children with cerebral palsy. It is important to investigate its relationship with some variables, and the objective of this study was to identify factors associated with OD. MATERIALS AND METHODS: Case-control study in patients with cerebral palsy from 8months to 15years of age, from November 2018 to November 2019, approved by the Ethics Committee. The diagnosis of OD was made by videofluoroscopy when there was nasopharyngeal reflux, stagnation in the vallecular sinuses, in the piriformis sinuses, penetration, and aspiration. The independent variables were type of cerebral palsy, gross motor impairment classified into five levels, nutritional status and comorbidities. One case with OD was included and the next one without alterations in videofluoroscopy was control. The variables were compared with Chi square and Student's t. The association was measured with odds ratio. The confidence interval was 95%. RESULTS: Thirty patients with OD and 30without OD were studied. Sex, age, birth weight, and gestational age had a similar distribution in the two groups. From the data perceived by the mothers at the time of feeding, the greater frequency of the difficulty in the transfer of the food bolus in the group with OD showed a statistically significant difference (P<.001) and of the studied factors, the levelV of the gross motor involvement was associated with a higher frequency of OD. CONCLUSIONS: OD was associated with level V of gross motor involvement.

Zinc Gluconate Induces Potentially Cancer Chemopreventive Activity in Barrett's Esophagus: A Phase 1 Pilot Study.

BACKGROUND: Chemopreventive effects of zinc for esophageal cancer have been well documented in animal models. This prospective study explores if a similar, potentially chemopreventive action can be seen in Barrett's esophagus (BE) in humans. AIMS: To determine if molecular evidence can be obtained potentially indicating zinc's chemopreventive action in Barrett's metaplasia. METHODS: Patients with a prior BE diagnosis were placed on oral zinc gluconate (14 days of 26.4 mg zinc BID) or a sodium gluconate placebo, prior to their surveillance endoscopy procedure. Biopsies of Barrett's mucosa were then obtained for miRNA and mRNA microarrays, or protein analyses. RESULTS: Zinc-induced mRNA changes were observed for a large number of transcripts. These included downregulation of transcripts encoding proinflammatory proteins (IL32, IL1ß, IL15, IL7R, IL2R, IL15R, IL3R), upregulation of anti-inflammatory mediators (IL1RA), downregulation of transcripts mediating epithelial-to-mesenchymal transition (EMT) (LIF, MYB, LYN, MTA1, SRC, SNAIL1, and TWIST1), and upregulation of transcripts that oppose EMT (BMP7, MTSS1, TRIB3, GRHL1). miRNA arrays showed significant upregulation of seven miRs with tumor suppressor activity (-125b-5P, -132-3P, -548z, -551a, -504, -518, and -34a-5P). Of proteins analyzed by Western blot, increased expression of the pro-apoptotic protein, BAX, and the tight junctional protein, CLAUDIN-7, along with decreased expression of BCL-2 and VEGF-R2 were noteworthy. CONCLUSIONS: When these mRNA, miRNA, and protein molecular data are considered collectively, a cancer chemopreventive action by zinc in Barrett's metaplasia may be possible for this precancerous esophageal tissue. These results and the extensive prior animal model studies argue for a future prospective clinical trial for this safe, easily-administered, and inexpensive micronutrient, that could determine if a chemopreventive action truly exists.

Intervenciones psicológicas en cuidadores primarios de pacientes con trasplante de células hematopoyéticas: Revisión descriptiva / Psychological interventions on primary care healthcare providers of patients with hematopoietic cells transplantation: Descriptive revision / Intervenções psicológicas em cuidadores primários de pacientes com transplante de células hematopoiéticas: Revisão descritiva

Resumen Introducción: El ser cuidador primario informal de un paciente con indicación médica de trasplante de células progenitoras hematopoyéticas puede tener consecuencias negativas en su salud mental y calidad de vida. Objetivo: Describir las intervenciones psicológicas disponibles para el cuidador primario de pacientes sometidos a trasplante de células hematopoyéticas. Metodología: Se realizó una búsqueda sistematizada de los últimos 10 años con los términos MeSH: psychotherapy AND caregive AND stem cell transplantation en las principales bases de datos médicas y de psicología, para su análisis se empleó la estrategia: Problema, Intervención, Comparación y Outcomes (PICO). Resultados: Se identificaron 122 artículos, de ellos diez cumplieron los criterios de inclusión. Las intervenciones provenían de profesionales de enfermería o trabajo social; el 50% incluyó diadas (paciente y cuidador primario), mostraron una tendencia de duración corta, enfocada al periodo posterior al trasplante. Se basan en el entrenamiento en solución de problemas, manejo de estrés, atención plena y expresión emocional. Las intervenciones lograron la disminución de la depresión, ansiedad y estrés en el cuidador; pero no alcanzaron permanencia en la significancia estadística de dichos restablecimientos. Discusión: De acuerdo con lo observado en las publicaciones y por su impacto positivo en la salud mental, se recomienda la implementación de intervenciones psicológicas en cuidadores de pacientes con trasplante de células progenitoras hematopoyéticas. Conclusión: El apoyo psicológico brindado al cuidador generalmente es de profesionales de la salud que no pertenecen al área de la psicología, con resultados clínicos favorables en las etapas más críticas de su estado mental.

Abstract Introduction: Being an informal primary healthcare provider of a patient who undergoes hematopoietic progeny cells transplantation can have adverse consequences on mental health and the quality of life. Objective: To describe the available psychological interventions for the primary healthcare provider of patients undergoing hematopoietic cells transplantations. Methodology: A systematized search of the last 10 years using the MeSH terms psychotherapy AND caregiver AND stem cell transplantation was conducted on the main medical and psychological databases. The analysis strategy followed the PICO scheme (Problem, Intervention, Comparison, Outcomes). Results: 122 articles were identified, and 10 of them fulfilled the inclusion criteria. The interventions were related to nursing or social work professionals. 50% described patient-healthcare provider dyads with short interventions focused on the post-transplantation period. Discussion: According to what has been observed in the publications and due to its positive impact on mental health, the implementation of psychological interventions is recommended in caregivers of patients who underwent hematopoietic stem cell transplantation. Conclusion: The psychological support provided to the caregiver comes mainly from health professionals who do not belong to the area of psychology, with favorable clinical results in the most critical periods for their mental state.

Resumo Introdução: Ser cuidador primário informal de um paciente sometido a transplante de células progenitoras hematopoiéticas pode ter consequências negativas na saúde mental e na qualidade de vida. Objetivo: Descrever as intervenções psicológicas disponíveis para o cuidador primário de pacientes sometidos a transplante de células hematopoiéticas. Metodologia: Realizou-se uma busca sistematizada dos últimos 10 anos com os termos MeSH: psychotherapy AND caregive AND stem cell transplantation nas principais bases de dados médicas e de psicologia, para sua análise realizou-se a estratégia: Problema, Intervenção, Comparação e Outcomes (PICO). Resultados: Identificaram-se 122 artigos, dos quais, dez cumpriram os critérios de inclusão. As intervenções provinham de profissionais em enfermagem ou trabalho social; o 50% incluiu díades (paciente e cuidador primário), mostraram uma tendência de duração curta, focalizada no período posterior ao transplante. Baseiam-se no treinamento em solução de problemas, manejo de estresse, atenção plena e expressão emocional. As intervenções conseguiram melhoras clínicas na diminuição da depressão, ansiedade e estresse no cuidador; mas não alcançaram permanência na significância estatística destes restabelecimentos. Discussão: Conforme o observado nas publicações e por seu impacto positivo na saúde mental, recomenda-se a implementação de intervenções psicológicas em cuidadores de pacientes para quem se indicou transplante de células progenitoras hematopoiéticas. Conclusão: O apoio psicológico oferecido ao cuidador vem de principalmente profissionais da saúde que não pertencem à área da psicologia, com resultados clínicos favoráveis nos períodos mais críticos para seu estado mental.

Intratumoral delivery of tavokinogene telseplasmid yields systemic immune responses in metastatic melanoma patients.

BACKGROUND: Interleukin 12 (IL-12) is a pivotal regulator of innate and adaptive immunity. We conducted a prospective open-label, phase II clinical trial of electroporated plasmid IL-12 in advanced melanoma patients (NCT01502293). PATIENTS AND METHODS: Patients with stage III/IV melanoma were treated intratumorally with plasmid encoding IL-12 (tavokinogene telseplasmid; tavo), 0.5 mg/ml followed by electroporation (six pulses, 1500 V/cm) on days 1, 5, and 8 every 90 days in the main study and additional patients were treated in two alternative schedule exploration cohorts. Correlative analyses for programmed death-ligand 1 (PD-L1), flow cytometry to assess changes in immune cell subsets, and analysis of immune-related gene expression were carried out on pre- and post-treatment samples from study patients, as well as from additional patients treated during exploration of additional dosing schedules beyond the pre-specified protocol dosing schedule. Response was measured by study-specific criteria to maximize detection of latent and potentially transient immune responses in patients with multiple skin lesions and toxicities were graded by the Common Terminology Criteria for Adverse Events version 4.0 (CTCAE v4.0). RESULTS: The objective overall response rate was 35.7% in the main study (29.8% in all cohorts), with a complete response rate of 17.9% (10.6% in all cohorts). The median progression-free survival in the main study was 3.7 months while the median overall survival was not reached at a median follow up of 29.7 months. A total of 46% of patients in all cohorts with uninjected lesions experienced regression of at least one of these lesions and 25% had a net regression of all untreated lesions. Transcriptomic and immunohistochemistry analysis showed that immune activation and co-stimulatory transcripts were up-regulated but there was also increased adaptive immune resistance. CONCLUSIONS: Intratumoral Tavo was well tolerated and led to systemic immune responses in advanced melanoma patients. While tumor regression and increased immune infiltration were observed in treated as well as untreated/distal lesions, adaptive immune resistance limited the response.

Randomized phase III KEYNOTE-045 trial of pembrolizumab versus paclitaxel, docetaxel, or vinflunine in recurrent advanced urothelial cancer: results of >2 years of follow-up.

BACKGROUND: Novel second-line treatments are needed for patients with advanced urothelial cancer (UC). Interim analysis of the phase III KEYNOTE-045 study showed a superior overall survival (OS) benefit of pembrolizumab, a programmed death 1 inhibitor, versus chemotherapy in patients with advanced UC that progressed on platinum-based chemotherapy. Here we report the long-term safety and efficacy outcomes of KEYNOTE-045. PATIENTS AND METHODS: Adult patients with histologically/cytologically confirmed UC whose disease progressed after first-line, platinum-containing chemotherapy were enrolled. Patients were randomly assigned 1 : 1 to receive pembrolizumab [200 mg every 3 weeks (Q3W)] or investigator's choice of paclitaxel (175 mg/m2 Q3W), docetaxel (75 mg/m2 Q3W), or vinflunine (320 mg/m2 Q3W). Primary end points were OS and progression-free survival (PFS) per Response Evaluation Criteria in Solid Tumors, version 1.1 (RECIST v1.1) by blinded independent central radiology review (BICR). A key secondary end point was objective response rate per RECIST v1.1 by BICR. RESULTS: A total of 542 patients were enrolled (pembrolizumab, n = 270; chemotherapy, n = 272). Median follow-up as of 26 October 2017 was 27.7 months. Median 1- and 2-year OS rates were higher with pembrolizumab (44.2% and 26.9%, respectively) than chemotherapy (29.8% and 14.3%, respectively). PFS rates did not differ between treatment arms; however, 1- and 2-year PFS rates were higher with pembrolizumab. The objective response rate was also higher with pembrolizumab (21.1% versus 11.0%). Median duration of response to pembrolizumab was not reached (range 1.6+ to 30.0+ months) versus chemotherapy (4.4 months; range 1.4+ to 29.9+ months). Pembrolizumab had lower rates of any grade (62.0% versus 90.6%) and grade ≥3 (16.5% versus 50.2%) treatment-related adverse events than chemotherapy. CONCLUSIONS: Long-term results (>2 years' follow-up) were consistent with those of previously reported analyses, demonstrating continued clinical benefit of pembrolizumab over chemotherapy for efficacy and safety for treatment of locally advanced/metastatic, platinum-refractory UC. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02256436.

Historical atmospheric pollution trends in Southeast Asia inferred from lake sediment records.

Fossil fuel combustion leads to increased levels of air pollution, which negatively affects human health as well as the environment. Documented data for Southeast Asia (SEA) show a strong increase in fossil fuel consumption since 1980, but information on coal and oil combustion before 1980 is not widely available. Spheroidal carbonaceous particles (SCPs) and heavy metals, such as mercury (Hg), are emitted as by-products of fossil fuel combustion and may accumulate in sediments following atmospheric fallout. Here we use sediment SCP and Hg records from several freshwater lentic ecosystems in SEA (Malaysia, Philippines, Singapore) to reconstruct long-term, region-wide variations in levels of these two key atmospheric pollution indicators. The age-depth models of Philippine sediment cores do not reach back far enough to date first SCP presence, but single SCP occurrences are first observed between 1925 and 1950 for a Malaysian site. Increasing SCP flux is observed at our sites from 1960 onward, although individual sites show minor differences in trends. SCP fluxes show a general decline after 2000 at each of our study sites. While the records show broadly similar temporal trends across SEA, absolute SCP fluxes differ between sites, with a record from Malaysia showing SCP fluxes that are two orders of magnitude lower than records from the Philippines. Similar trends in records from China and Japan represent the emergence of atmospheric pollution as a broadly-based inter-region environmental problem during the 20th century. Hg fluxes were relatively stable from the second half of the 20th century onward. As catchment soils are also contaminated with atmospheric Hg, future soil erosion can be expected to lead to enhanced Hg flux into surface waters.

Predictors of responses to immune checkpoint blockade in advanced melanoma.

Immune checkpoint blockers (ICB) have become pivotal therapies in the clinical armamentarium against metastatic melanoma (MMel). Given the frequency of immune related adverse events and increasing use of ICB, predictors of response to CTLA-4 and/or PD-1 blockade represent unmet clinical needs. Using a systems biology-based approach to an assessment of 779 paired blood and tumor markers in 37 stage III MMel patients, we analyzed association between blood immune parameters and the functional immune reactivity of tumor-infiltrating cells after ex vivo exposure to ICB. Based on this assay, we retrospectively observed, in eight cohorts enrolling 190 MMel patients treated with ipilimumab, that PD-L1 expression on peripheral T cells was prognostic on overall and progression-free survival. Moreover, detectable CD137 on circulating CD8+ T cells was associated with the disease-free status of resected stage III MMel patients after adjuvant ipilimumab + nivolumab (but not nivolumab alone). These biomarkers should be validated in prospective trials in MMel.The clinical management of metastatic melanoma requires predictors of the response to checkpoint blockade. Here, the authors use immunological assays to identify potential prognostic/predictive biomarkers in circulating blood cells and in tumor-infiltrating lymphocytes from patients with resected stage III melanoma.

Upregulation of MARCKS in kidney cancer and its potential as a therapeutic target.

Targeted therapeutics, such as those abrogating hypoxia inducible factor (HIF)/vascular endothelial growth factor signaling, are initially effective against kidney cancer (or renal cell carcinoma, RCC); however, drug resistance frequently occurs via subsequent activation of alternative pathways. Through genome-scale integrated analysis of the HIF-α network, we identified the major protein kinase C substrate MARCKS (myristoylated alanine-rich C kinase substrate) as a potential target molecule for kidney cancer. In a screen of nephrectomy samples from 56 patients with RCC, we found that MARCKS expression and its phosphorylation are increased and positively correlate with tumor grade. Genetic and pharmacologic suppression of MARCKS in high-grade RCC cell lines in vitro led to a decrease in cell proliferation and migration. We further demonstrated that higher MARCKS expression promotes growth and angiogenesis in vivo in an RCC xenograft tumor. MARCKS acted upstream of the AKT/mTOR pathway, activating HIF-target genes, notably vascular endothelial growth factor-A. Following knockdown of MARCKS in RCC cells, the IC50 of the multikinase inhibitor regorafenib was reduced. Surprisingly, attenuation of MARCKS using the MPS (MARCKS phosphorylation site domain) peptide synergistically interacted with regorafenib treatment and decreased survival of kidney cancer cells through inactivation of AKT and mTOR. Our data suggest a major contribution of MARCKS to kidney cancer growth and provide an alternative therapeutic strategy of improving the efficacy of multikinase inhibitors.

Molecular and serological prevalence of Anaplasma and Ehrlichia sp. among stray dogs in East Malaysia.

Anaplasma and Ehrlichia species are important bacterial pathogens of various animal species including dogs and humans. They constitute a major health problem worldwide, especially in the tropics and sub-tropics. In order to identify the prevalence of Anaplasma and Ehrlichia sp. from dogs in the selected area, a total of 100 randomly selected clinically healthy rescued dogs from animal shelters in Sarawak and Sabah, East Malaysia (50 dogs per State) were used for this study. Whole blood was collected from these dogs and screened for presence of Anaplasma and Ehrlichia spp. using polymerase chain reaction (PCR) and commercial ELISA test (SNAP 4Dx® Plus) for detection of antigen and antibody against the two pathogens respectively. 27% (A. platys) and 33% (E. canis) of the sampled population were positive using PCR; while the serological prevalence was 29% for Anaplasma sp. and 55% for Ehrlichia sp. The results of this investigation revealed that stray dogs are potential reservoir hosts of Anaplasma and Ehrlichia sp. infection with a high prevalence in the area. The high prevalence rates detected highlights the urgent need to address the growing population of possible vectors and reservoirs for transmission in the region and also to look into the public health aspect of the disease.

Nitric oxide participates in IFN-gamma-induced HUVECs hyperpermeability.

The endothelial barrier function is tightly controlled by a broad range of signaling cascades including nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway. It has been proposed that disturbances in NO and cGMP production could interfere with proper endothelial barrier function. In this study, we assessed the effect of interferon-gamma (IFN-gamma), a pro-inflammatory cytokine, on NO and cGMP levels and examined the mechanisms by which NO and cGMP regulate the IFN-gamma-mediated HUVECs hyperpermeability. The flux of fluorescein isothiocyanate-labeled dextran across cell monolayers was used to study the permeability of endothelial cells. Here, we found that IFN-gamma significantly attenuated basal NO concentration and the increased NO levels supplied by a NO donor, sodium nitroprusside (SNP). Besides, application of IFN-gamma also significantly attenuated both the basal cGMP concentration and the increased cGMP production donated by a cell permeable cGMP analogue, 8-bromo-cyclic GMP (8-Br-cGMP). In addition, exposure of the cell monolayer to IFN-gamma significantly increased HUVECs basal permeability. However, L-NAME pretreatment did not suppress IFN-gamma-induced HUVECs hyperpermeability. L-NAME pretreatment followed by SNP or SNP pretreatment partially reduced IFN-gamma-induced HUVECs hyperpermeability. Pretreatment with a guanylate cyclase inhibitor, 6-anilino-5,8-quinolinedione (LY83583), led to a further increase in IFN-gamma-induced HUVECs hyperpermeability. The findings suggest that the mechanism underlying IFN-gamma-induced increased HUVECs permeability is partly related to the inhibition of NO production.

Selection by mating competitiveness improves the performance of Anastrepha ludens males of the genetic sexing strain Tapachula-7.

The sexual performance of Anastrepha ludens males of the Tapachula-7 genetic sexing strain, produced via selection based on mating success, was compared with that of males produced without selection in competition with wild males. Mating competition, development time, survival, mass-rearing quality parameters and pheromone production were compared. The results showed that selection based on mating competitiveness significantly improved the sexual performance of offspring. Development time, survival of larvae, pupae and adults, and weights of larvae and pupae increased with each selection cycle. Differences in the relative quantity of the pheromone compounds (Z)-3-nonenol and anastrephin were observed when comparing the parental males with the F4 and wild males. The implications of this colony management method on the sterile insect technique are discussed.

Incremental value of three-dimensional over two-dimensional transthoracic echocardiography in the assessment of cor triatriatum sinister in a child.

A 2.5-year-old boy presented with frequent hospitalizations due to recurrent respiratory tract infections with dyspnea. A fibromuscular membrane dividing the left atrium with obstruction of left atrial inflow to the left ventricle was documented by two-dimensional transthoracic echocardiography (2DTTE). Live/real time three-dimensional transthoracic echocardiography (3DTTE) provided incremental value over 2DTTE by providing en face views of the 2 obstructing orifices in the membrane enabling accurate assessment of their position, shape and size. 3DTTE also showed clearly the location of the membrane superior and proximal to the left atrial appendage which was not well delineated by 2DTTE. In addition, 3DTTE demonstrated the full extent of the left atrial appendage and careful sequential cropping of the 3D dataset showed it to have 2 distinct lobes and no thrombus. These findings provided comprehensive assessment of the lesion and were helpful in surgical decision making and planning.