Feasibility of structuring electronic health record data to facilitate real-world data research: ICAREdata methods applied to multicenter cancer clinical trials.

BACKGROUND: The use of electronic health record (EHR) data for research is limited by a lack of structure and a standard data model. The objective of the ICAREdata (Integrating Clinical Trials and Real-World Endpoints Data) project was to structure key research data elements in EHRs using a minimal Common Oncology Data Elements (mCODE) data model to extract and transmit data. METHODS: The ICAREdata project captured two EHR data elements essential to clinical trials: cancer disease status and treatment plan change. The project was implemented in clinical sites participating in Alliance for Clinical Trials in Oncology trials. Data were extracted from EHRs and sent by secure Fast Healthcare Interoperability Resource messaging (a standard for exchanging EHRs) to a database. Selected elements were compared with corresponding data from the trial's electronic data capture (EDC) system, Medidata Rave. RESULTS: By December 2023, data were extracted and transmitted from 10 sites for 35 patients, involving 367 clinical encounters across 15 clinical trials. Data through March 2023 demonstrated that concordance for the elements treatment plan change and cancer disease status was 79% and 34%, respectively. When disease evaluation was reported by both EHR and EDC (n = 15), there was 87% agreement on cancer disease status. CONCLUSIONS: Documentation, extraction, and aggregation of structured data elements in EHRs using mCODE and ICAREdata methods is feasible in multi-institutional cancer clinical trials. EDC as a reference data set allowed assessment of the completeness of EHR data capture. Future initiatives will focus on elements with shared definitions in clinical and research environments and efficient workflows. PLAIN LANGUAGE SUMMARY: Clinical trials use electronic case report forms to report data, and data must be manually entered on these forms, which is costly and time consuming. ICAREdata methods use structured, organized data from clinical trials that can be more easily shared instead having to enter free text into electronic health records.

Breast cancer cell-secreted miR-199b-5p hijacks neurometabolic coupling to promote brain metastasis.

Breast cancer metastasis to the brain is a clinical challenge rising in prevalence. However, the underlying mechanisms, especially how cancer cells adapt a distant brain niche to facilitate colonization, remain poorly understood. A unique metabolic feature of the brain is the coupling between neurons and astrocytes through glutamate, glutamine, and lactate. Here we show that extracellular vesicles from breast cancer cells with a high potential to develop brain metastases carry high levels of miR-199b-5p, which shows higher levels in the blood of breast cancer patients with brain metastases comparing to those with metastatic cancer in other organs. miR-199b-5p targets solute carrier transporters (SLC1A2/EAAT2 in astrocytes and SLC38A2/SNAT2 and SLC16A7/MCT2 in neurons) to hijack the neuron-astrocyte metabolic coupling, leading to extracellular retention of these metabolites and promoting cancer cell growth. Our findings reveal a mechanism through which cancer cells of a non-brain origin reprogram neural metabolism to fuel brain metastases.

Regulation of Helicobacter pylori Urease and Acetone Carboxylase Genes by Nitric Oxide and the CrdRS Two-Component System.

Helicobacter pylori colonizes the human gastric mucosa and causes various gastroduodenal diseases, including peptic ulceration and gastric cancer. Colonization requires the actions of two-component systems (TCSs) to sense and respond to changes in the host environment. In this study, we evaluated gene regulation mediated by the CrdRS TCS. Few studies have evaluated this TCS, leaving the signal(s) yet to be exhaustively determined and a need for a more complete regulon to be delineated. We performed RNA sequencing (RNA-Seq) on three isogenic H. pylori 26695 mutants: a control, a mutant with deletion of the sensory histidine kinase, ΔcrdS, and a mutant with deletion of the response regulator, ΔcrdR. Comparison of the RNA-Seq results from these mutants established a 40-gene regulon putatively controlled by the CrdRS TCS. Quantitative reverse transcriptase PCR (RT-qPCR) was used to validate 7 of 11 putative regulon members selected for analysis. We further investigated 6 confirmed CrdRS regulon genes by using phospho-incompetent H. pylori 26695 CrdR D53A and CrdS H173A mutants. These experiments further confirmed the role of CrdRS in regulation of urease, acetone carboxylase, hofD, and HP1440. Expression of these CrdRS regulon genes was also evaluated under 10 µM nitric oxide (NO) conditions. This revealed that ureA, acxA, hofD, and HP1440 expression is affected by NO in a CrdRS-dependent manner. Importantly, three of these genes (ureA, acxA, and hofD) are known to play important roles in H. pylori colonization of the stomach. IMPORTANCE The molecular strategies used by Helicobacter pylori to colonize and persist in the harsh environment of the human stomach are a critical area of study. Our study identified several genes in this gastric pathogen, including ureA, a gene encoding a protein essential to the survival of H. pylori, that are regulated via the CrdRS two-component system (TCS) in response to nitric oxide (NO). NO is a product of the innate immune system of the human host. The identification of these genes whose expression is regulated by this molecule may give insights to novel therapeutics. Two genes (ureA and acxA) determined in this study to be regulated by NO via CrdRS have been previously determined to be regulated by other TCSs, indicating that the expression of these genes may be of critical importance to H. pylori.

Cancer-cell-secreted extracellular vesicles suppress insulin secretion through miR-122 to impair systemic glucose homeostasis and contribute to tumour growth.

Epidemiological studies demonstrate an association between breast cancer (BC) and systemic dysregulation of glucose metabolism. However, how BC influences glucose homeostasis remains unknown. We show that BC-derived extracellular vesicles (EVs) suppress pancreatic insulin secretion to impair glucose homeostasis. EV-encapsulated miR-122 targets PKM in ß-cells to suppress glycolysis and ATP-dependent insulin exocytosis. Mice receiving high-miR-122 EVs or bearing BC tumours exhibit suppressed insulin secretion, enhanced endogenous glucose production, impaired glucose tolerance and fasting hyperglycaemia. These effects contribute to tumour growth and are abolished by inhibiting EV secretion or miR-122, restoring PKM in ß-cells or supplementing insulin. Compared with non-cancer controls, patients with BC have higher levels of circulating EV-encapsulated miR-122 and fasting glucose concentrations but lower fasting insulin; miR-122 levels are positively associated with glucose and negatively associated with insulin. Therefore, EV-mediated impairment of whole-body glycaemic control may contribute to tumour progression and incidence of type 2 diabetes in some patients with BC.

Satisfação do espectador de futebol com jogos e serviços no Brasil / Satisfacción de los espectadores de fútbol con los juegos y servicios en Brasil / Football spectator satisfaction with games and services in Brazil

O objetivo desta investigação foi comparar a satisfação com os jogos e serviços dos torcedores de clubes de futebol do Estado de Pernambuco em relação ao Campeonato Brasileiro nos anos de 2016 a 2019. A amostra foi composta por 937 torcedores, coletados entre os anos de 2016 e 2019. O procedimento de coleta adotado foi o E-survey, onde os indivíduos responderam ao questionário online disponível na plataforma Google Forms. O instrumento utilizado foi um questionário composto por três dimensões: 1. Satisfação com o jogo; 2. Satisfação com os serviços; 3. Caracterização sociodemográfica. Para a análise dos dados foi realizada uma análise fatorial confirmatória a fim de confirmar o ajustamento do modelo proposto, conduzida no software AMOS, seguida de uma ANOVA a dois fatores e uma correlação de Pearson, realizadas no software SPSS Statistics versão 24. O modelo refinado apresentou um bom ajustamento aos dados [χ² (8) = 24.75 (p < .001), χ²/gl = 3.09, TLI = .99, CFI = .99, GFI = .99, RMSEA = .05, MECVI = .05]. Na satisfação com o jogo, evidenciou-se que há diferenças na interação entre clube e ano, assim como na satisfação com o serviço. Demostrou-se em sua maioria correlações significativas entre a satisfação com o jogo e com o serviço, com destaque para o clube 3 (r=.860) e o clube 2 (r=.718) em 2018. Portanto, é fundamental que os clubes de futebol passem a qualificar a gestão dos seus serviços, não só os elementos ligados ao resultado esportivo. (AU)

El objetivo de esta investigación fue comparar la satisfacción con los juegos y servicios de los fanáticos de los clubes de fútbol en el Estado de Pernambuco en relación con el Campeonato Brasileño en los años 2016 a 2019. La muestra consistió en 937 aficionados, recolectados entre los años 2016 a 2019. El procedimiento de recolección adoptado fue la encuesta electrónica, donde las personas respondieron el cuestionario en línea disponible en la plataforma Google Forms. El instrumento utilizado fue un cuestionario compuesto por tres dimensiones: 1. Satisfacción con el juego; 2. Satisfacción con los servicios; 3. Caracterización sociodemográfica. Para el análisis de datos, se realizó un análisis factorial confirmatorio para confirmar el ajuste del modelo propuesto, realizado en el software AMOS, seguido de un ANOVA de dos vías y una correlación de Pearson, realizado en el software SPSS Statistics versión 24. O El modelo refinado presentó un buen ajuste a los datos [χ² (8) = 24.75 (p < .001), χ²/gl = 3.09, TLI = .99, CFI = .99, GFI = .99, RMSEA = .05, MECVI = .05]. En la satisfacción con el juego, fue evidente que existen diferencias en la interacción entre el club y el año, así como en la satisfacción con el servicio. Fue demostrado correlaciones significativas entre la satisfacción con el juego y el servicio, con énfasis en el club 3 (r = .860) y el club 2 (r = .718) en 2018. Por lo tanto, es esencial que los clubes de fútbol comiencen a calificar la gestión de sus servicios, no solo los elementos vinculados al resultado deportivo. (AU)

The objective of this research was to compare the satisfaction with the games and services of soccer club fans in the State of Pernambuco in relation to the Brazilian Championship in the years 2016 to 2019. The sample consisted of 937 fans, collected between the years 2016 to 2019. The collection procedure adopted was the E-survey, where individuals answered the online questionnaire available on the Google Forms platform. The instrument used was a questionnaire composed of three dimensions: 1. Satisfaction with the game; 2. Satisfaction with services; 3. Sociodemographic characterization. For the data analysis, a confirmatory factor analysis was performed to confirm the adjustment of the proposed model, conducted in AMOS software, followed by an ANOVA two-way and a Pearson correlation, performed in the SPSS Statistics software version 24. The refined model presented a good fit to the data [χ² (8) = 24.75 (p < .001), χ²/gl = 3.09, TLI = .99, CFI = .99, GFI = .99, RMSEA = .05, MECVI = .05]. In the satisfaction with the game, it was evident that there are differences in the interaction between club and year, as well as in satisfaction with the service. Evidenced significant correlations between satisfaction with the game and with the service, with emphasis on club 3 (r=.860) and club 2 (r=.718) in 2018. Therefore, it is essential that soccer clubs start to qualify the management of their services, not only the elements linked to the sports result. (AU)

Cancer-secreted miRNAs regulate amino-acid-induced mTORC1 signaling and fibroblast protein synthesis.

Metabolic reprogramming of non-cancer cells residing in a tumor microenvironment, as a result of the adaptations to cancer-derived metabolic and non-metabolic factors, is an emerging aspect of cancer-host interaction. We show that in normal and cancer-associated fibroblasts, breast cancer-secreted extracellular vesicles suppress mTOR signaling upon amino acid stimulation to globally reduce mRNA translation. This is through delivery of cancer-derived miR-105 and miR-204, which target RAGC, a component of Rag GTPases that regulate mTORC1 signaling. Following amino acid starvation and subsequent re-feeding, 13 C-arginine labeling of de novo synthesized proteins shows selective translation of proteins that cluster to specific cellular functional pathways. The repertoire of these newly synthesized proteins is altered in fibroblasts treated with cancer-derived extracellular vesicles, in addition to the overall suppressed protein synthesis. In human breast tumors, RAGC protein levels are inversely correlated with miR-105 in the stroma. Our results suggest that through educating fibroblasts to reduce and re-prioritize mRNA translation, cancer cells rewire the metabolic fluxes of amino acid pool and dynamically regulate stroma-produced proteins during periodic nutrient fluctuations.

Servicio de determinación de hormonas a instituciones hospitalarias en el Centro de Isótopos / Hormone determination service to hospital institutions the Center of Isotopes

RESUMEN En los marcos de la estrecha colaboración entre la Agencia de Energía Nuclear y Tecnologías de Avanzada y el Ministerio de Salud Pública, a inicios de este siglo se procedió a organizar, en el Centro de Isótopos, un servicio de determinación de hormonas por metodologías nucleares, atendiendo a necesidades de diversas instalaciones hospitalarias. El centro disponía de personal capacitado en técnicas de radioinmunoanálisis y seguridad radiológica, el equipamiento necesario y laboratorios licenciados para el trabajo con sustancias radiactivas. Para ello se hizo necesario incorporar al Sistema de Calidad las regulaciones vigentes para laboratorios clínicos y adecuar la infraestructura para el procesamiento de no menos de 1000 muestras por semana, procedentes de 16 hospitales. Se requería recogida y transportación de las muestras, recepción, inspección y registro, análisis, evaluación y entrega de los resultados. En función de la actividad se diseñó y puso en operación un sistema informático y se normalizaron acciones para asegurar competencia técnica avalada por: trazabilidad; incertidumbre; ejercicios de aptitud y otros requisitos exigidos por la regulación de Buenas Prácticas de Laboratorio Clínico vigentes en el país. A raíz de nuevas regulaciones, el servicio ha introducido de manera paulatina determinadas mejoras. En 15 años de trabajo este laboratorio ha resultado el principal del país en emplear métodos radiactivos para la cuantificación de hormonas, lo que favorece la atención de más de 70 000 pacientes como promedio al año, principalmente de la capital. Esto es parte de nuestra contribución en el 500 aniversario de La Habana.

ABSTRACT Within the framework of the close collaboration between the Agency of Nuclear Energy and Advanced Technologies and the Ministry of Public Health, at the beginning of this century, and considering the need of different hospitals, the Center of Isotopes (CENTIS), began providing service to determine hormones by nuclear methodologies,. CENTIS had qualified personnel in immune radioanalysis RIA/IRMA techniques and radiation safety, in addition to having the required equipment and accredited laboratories to work with radioactive substances. As a result, it was necessary to incorporate to CENTIS Quality System, the regulations in force for clinical laboratories , and to adapt its infrastructure for the processing of no less than 1000 samples per week from 16 hospitals. The Service required collection and transportation of the samples, reception, inspection and registration, test, evaluation and delivery of the test results. In order to fulfil this activity, a computer system was designed and set , and standards for actions were established to guarantee a high technical quality endorsed by traceability, uncertainty; proficiency tests and other requirements demanded by the Good Practices of Clinical Laboratory (BPLC 03 -2009) issued by CECMED. Due to new regulations, several improvements have been introduced. In 15 years, CENTIS laboratory has become the most important in the country using radioactive methods for the quantification of hormones, with an average testing of more than 70 000 patients per year, mainly in the capital. This is part of our contribution in the 500th Anniversary of Havana.

Genotypic variation in whole-plant transpiration efficiency in sorghum only partly aligns with variation in stomatal conductance.

Water scarcity can limit sorghum (Sorghum bicolor (L.) Moench) production in dryland agriculture, but increased whole-plant transpiration efficiency (TEwp, biomass production per unit of water transpired) can enhance grain yield in such conditions. The objectives of this study were to quantify variation in TEwp for 27 sorghum genotypes and explore the linkages of this variation to responses of the underpinning leaf-level processes to environmental conditions. Individual plants were grown in large lysimeters in two well-watered experiments. Whole-plant transpiration per unit of green leaf area (TGLA) was monitored continuously and stomatal conductance and maximum photosynthetic capacity were measured during sunny conditions on recently expanded leaves. Leaf chlorophyll measurements of the upper five leaves of the main shoot were conducted during early grain filling. TEwp was determined at harvest. The results showed that diurnal patterns in TGLA were determined by vapour pressure deficit (VPD) and by the response of whole-plant conductance to radiation and VPD. Significant genotypic variation in the response of TGLA to VPD occurred and was related to genotypic differences in stomatal conductance. However, variation in TGLA explained only part of the variation in TEwp, with some of the residual variation explained by leaf chlorophyll readings, which were a reflection of photosynthetic capacity. Genotypes with different genetic background often differed in TEwp, TGLA and leaf chlorophyll, indicating potential differences in photosynthetic capacity among these groups. Observed differences in TEwp and its component traits can affect adaptation to drought stress.

Stereophotogrammetric three-dimensional photography is an accurate and precise planimetric method for the clinical visualization and quantification of human papilloma virus-induced skin lesions.

BACKGROUND: The quantification of human papilloma virus (HPV)-induced skin lesions is essential for the clinical assessment of the course of disease and the response to treatment. However, clinical assessments that measure dimensions of lesions using a caliper do not provide complete insight into three-dimensional (3D) lesions, and its inter-rater variability is often poor. OBJECTIVE: The aim of this study was to validate a stereophotogrammetric 3D camera system for the quantification of HPV-induced lesions. METHODS: The camera system was validated for accuracy, precision and interoperator and inter-rater variability. Subsequently, 3D photographs were quantified and compared to caliper measurements for clinical validation by Bland-Altman modelling, based on data from 80 patients with cutaneous warts (CW), 24 with anogenital warts (AGW) patients and 12 with high-grade squamous intraepithelial lesions of the vulva (vulvar HSIL) with a total lesion count of 220 CW, 74 AGW and 31 vulvar HSIL. RESULTS: Technical validation showed excellent accuracy [coefficients of variation (CV) ≤ 0.68%] and reproducibility (CVs ≤ 2%), a good to excellent agreement between operators (CVs ≤ 8.7%) and a good to excellent agreement between different raters for all three lesion types (ICCs ≥ 0.86). When comparing 3D with caliper measurements, excellent biases were found for diameter of AGW (long diameter 5%), good biases were found for diameter of AGW (short diameter 10%) and height of CW (8%), and acceptable biases were found for the diameter of CW (11%) and vulvar HSIL (short diameter 14%, long diameter 16%). An unfavourable difference between these methods (bias 25%) was found for the assessment of height of AGWs. CONCLUSION: Stereophotogrammetric 3D imaging is an accurate and reliable method for the clinical visualization and quantification of HPV-induced skin lesions.

Metastatic breast cancer cells overexpress and secrete miR-218 to regulate type I collagen deposition by osteoblasts.

BACKGROUND: Bone is one of the most frequent metastatic sites of advanced breast cancer. Current therapeutic agents aim to inhibit osteoclast-mediated bone resorption but only have palliative effects. During normal bone remodeling, the balance between bone resorption and osteoblast-mediated bone formation is essential for bone homeostasis. One major function of osteoblast during bone formation is to secrete type I procollagen, which will then be processed before being crosslinked and deposited into the bone matrix. METHODS: Small RNA sequencing and quantitative real-time PCR were used to detect miRNA levels in patient blood samples and in the cell lysates as well as extracellular vesicles of parental and bone-tropic MDA-MB-231 breast cancer cells. The effects of cancer cell-derived extracellular vesicles isolated by ultracentrifugation and carrying varying levels of miR-218 were examined in osteoblasts by quantitative real-time PCR, Western blot analysis, and P1NP bone formation marker analysis. Cancer cells overexpressing miR-218 were examined by transcriptome profiling through RNA sequencing to identify intrinsic genes and pathways influenced by miR-218. RESULTS: We show that circulating miR-218 is associated with breast cancer bone metastasis. Cancer-secreted miR-218 directly downregulates type I collagen in osteoblasts, whereas intracellular miR-218 in breast cancer cells regulates the expression of inhibin ß subunits. Increased cancer secretion of inhibin ßA results in elevated Timp3 expression in osteoblasts and the subsequent repression of procollagen processing during osteoblast differentiation. CONCLUSIONS: Here we identify a twofold function of cancer-derived miR-218, whose levels in the blood are associated with breast cancer metastasis to the bone, in the regulation of type I collagen deposition by osteoblasts. The adaptation of the bone niche mediated by miR-218 might further tilt the balance towards osteolysis, thereby facilitating other mechanisms to promote bone metastasis.

Cancer-cell-secreted exosomal miR-105 promotes tumour growth through the MYC-dependent metabolic reprogramming of stromal cells.

Cancer and other cells residing in the same niche engage various modes of interactions to synchronize and buffer the negative effects of environmental changes. Extracellular microRNAs (miRNAs) have recently been implicated in the intercellular crosstalk. Here we show a mechanistic model involving breast-cancer-secreted, extracellular-vesicle-encapsulated miR-105, which is induced by the oncoprotein MYC in cancer cells and, in turn, activates MYC signalling in cancer-associated fibroblasts (CAFs) to induce a metabolic program. This results in the capacity of CAFs to display different metabolic features in response to changes in the metabolic environment. When nutrients are sufficient, miR-105-reprogrammed CAFs enhance glucose and glutamine metabolism to fuel adjacent cancer cells. When nutrient levels are low and metabolic by-products accumulate, these CAFs detoxify metabolic wastes, including lactic acid and ammonium, by converting them into energy-rich metabolites. Thus, the miR-105-mediated metabolic reprogramming of stromal cells contributes to sustained tumour growth by conditioning the shared metabolic environment.

Cross-kingdom inhibition of breast cancer growth by plant miR159.

MicroRNAs (miRNAs) are critical regulators of gene expression, and exert extensive impacts on development, physiology, and disease of eukaryotes. A high degree of parallelism is found in the molecular basis of miRNA biogenesis and action in plants and animals. Recent studies interestingly suggest a potential cross-kingdom action of plant-derived miRNAs, through dietary intake, in regulating mammalian gene expression. Although the source and scope of plant miRNAs detected in mammalian specimens remain controversial, these initial studies inspired us to determine whether plant miRNAs can be detected in Western human sera and whether these plant miRNAs are able to influence gene expression and cellular processes related to human diseases such as cancer. Here we found that Western donor sera contained the plant miRNA miR159, whose abundance in the serum was inversely correlated with breast cancer incidence and progression in patients. In human sera, miR159 was predominantly detected in the extracellular vesicles, and was resistant to sodium periodate oxidation suggesting the plant-originated 2'-O-methylation on the 3' terminal ribose. In breast cancer cells but not non-cancerous mammary epithelial cells, a synthetic mimic of miR159 was capable of inhibiting proliferation by targeting TCF7 that encodes a Wnt signaling transcription factor, leading to a decrease in MYC protein levels. Oral administration of miR159 mimic significantly suppressed the growth of xenograft breast tumors in mice. These results demonstrate for the first time that a plant miRNA can inhibit cancer growth in mammals.

Breast-cancer-secreted miR-122 reprograms glucose metabolism in premetastatic niche to promote metastasis.

Reprogrammed glucose metabolism as a result of increased glycolysis and glucose uptake is a hallmark of cancer. Here we show that cancer cells can suppress glucose uptake by non-tumour cells in the premetastatic niche, by secreting vesicles that carry high levels of the miR-122 microRNA. High miR-122 levels in the circulation have been associated with metastasis in breast cancer patients, and we show that cancer-cell-secreted miR-122 facilitates metastasis by increasing nutrient availability in the premetastatic niche. Mechanistically, cancer-cell-derived miR-122 suppresses glucose uptake by niche cells in vitro and in vivo by downregulating the glycolytic enzyme pyruvate kinase. In vivo inhibition of miR-122 restores glucose uptake in distant organs, including brain and lungs, and decreases the incidence of metastasis. These results demonstrate that, by modifying glucose utilization by recipient premetastatic niche cells, cancer-derived extracellular miR-122 is able to reprogram systemic energy metabolism to facilitate disease progression.

TGFß induces "BRCAness" and sensitivity to PARP inhibition in breast cancer by regulating DNA-repair genes.

UNLABELLED: Transforming growth factor beta (TGFß) proteins are multitasking cytokines, in which high levels at tumor sites generally correlate with poor prognosis in human patients with cancer. Previously, it was reported that TGFß downregulates the expression of ataxia telangiectasia-mutated (ATM) and mutS homolog 2 (MSH2) in breast cancer cells through an miRNA-mediated mechanism. In this study, expression of a panel of DNA-repair genes was examined, identifying breast cancer 1, early onset (BRCA1) as a target downregulated by TGFß through the miR181 family. Correlations between the expression levels of TGFß1 and the miR181/BRCA1 axis were observed in primary breast tumor specimens. By downregulating BRCA1, ATM, and MSH2, TGFß orchestrates DNA damage response in certain breast cancer cells to induce a "BRCAness" phenotype, including impaired DNA-repair efficiency and synthetic lethality to the inhibition of poly (ADP-ribose) polymerase (PARP). Xenograft tumors with active TGFß signaling exhibited resistance to the DNA-damaging agent doxorubicin but increased sensitivity to the PARP inhibitor ABT-888. Combination of doxorubicin with ABT-888 significantly improved the treatment efficacy in TGFß-active tumors. Thus, TGFß can induce "BRCAness" in certain breast cancers carrying wild-type BRCA genes and enhance the responsiveness to PARP inhibition, and the molecular mechanism behind this is characterized. IMPLICATIONS: These findings enable better selection of patients with sporadic breast cancer for PARP interventions, which have exhibited beneficial effects in patients carrying BRCA mutations.

Macrophage immunomodulation by breast cancer-derived exosomes requires Toll-like receptor 2-mediated activation of NF-κB.

Growing evidence links tumor progression with chronic inflammatory processes and dysregulated activity of various immune cells. In this study, we demonstrate that various types of macrophages internalize microvesicles, called exosomes, secreted by breast cancer and non-cancerous cell lines. Although both types of exosomes targeted macrophages, only cancer-derived exosomes stimulated NF-κB activation in macrophages resulting in secretion of pro-inflammatory cytokines such as IL-6, TNFα, GCSF, and CCL2. In vivo mouse experiments confirmed that intravenously injected exosomes are efficiently internalized by macrophages in the lung and brain, which correlated with upregulation of inflammatory cytokines. In mice bearing xenografted human breast cancers, tumor-derived exosomes were internalized by macrophages in axillary lymph nodes thereby triggering expression of IL-6. Genetic ablation of Toll-like receptor 2 (TLR2) or MyD88, a critical signaling adaptor in the NF-κB pathway, completely abolished the effect of tumor-derived exosomes. In contrast, inhibition of TLR4 or endosomal TLRs (TLR3/7/8/9) failed to abrogate NF-κB activation by exosomes. We further found that palmitoylated proteins present on the surface of tumor-secreted exosomes contributed to NF-κB activation. Thus, our results highlight a novel mechanism used by breast cancer cells to induce pro-inflammatory activity of distant macrophages through circulating exosomal vesicles secreted during cancer progression.

Cancer-secreted miR-105 destroys vascular endothelial barriers to promote metastasis.

Cancer-secreted microRNAs (miRNAs) are emerging mediators of cancer-host crosstalk. Here we show that miR-105, which is characteristically expressed and secreted by metastatic breast cancer cells, is a potent regulator of migration through targeting the tight junction protein ZO-1. In endothelial monolayers, exosome-mediated transfer of cancer-secreted miR-105 efficiently destroys tight junctions and the integrity of these natural barriers against metastasis. Overexpression of miR-105 in nonmetastatic cancer cells induces metastasis and vascular permeability in distant organs, whereas inhibition of miR-105 in highly metastatic tumors alleviates these effects. miR-105 can be detected in the circulation at the premetastatic stage, and its levels in the blood and tumor are associated with ZO-1 expression and metastatic progression in early-stage breast cancer.

Metabolomic profiling of ovarian carcinomas using mass spectrometry.

Most of the research on tumor cell metabolism has focused on glucose utilization. However, when glucose is limited, solid tumors are forced to catabolize alternative substrates such as fatty acids and amino acids as an energy source. Measuring these alternations in tumor cell metabolism enables us to track neoplastic changes in the tissue to lead towards a more reliable diagnostic outcome. Although a very small number of elements are used in biochemistry, the metabolome is structurally diverse for the production of simple compounds such as phosphate and amino acids as well as more structurally complex compounds such as nucleotides, oligosaccharides, and complex lipids. Characterization of the metabolome, therefore, requires analytical methods that can handle a wide range of molecular structures and physicochemical properties, including solubility, polarity, and molecular weight. A further factor for consideration in the selection of technology for metabolomics is the wide range of concentrations of biochemical typically present in biological systems. MS has established itself as the high-throughput, information-rich, industrially stable approach to assess both the composition of diverse sample types as well as changes to that composition following perturbation.

Tumorigenic potential of pituitary tumor transforming gene (PTTG) in vivo investigated using a transgenic mouse model, and effects of cross breeding with p53 (+/-) transgenic mice.

BACKGROUND: Pituitary tumor-transforming gene (PTTG) is an oncogene that is overexpressed in variety of tumors and exhibits characteristics of a transforming gene. Previous transgenic mouse models to access the tumorigenic potential in the pituitary and ovary have resulted in dysplasia without formation of visible tumors, possibly due to the insufficient expression of PTTG. PTTG expression level is critical for ovarian tumorigenesis in a xenograft model. Therefore, the tumorigenic function of PTTG in vivo remains unclear. We generated a transgenic mouse that overexpresses PTTG driven by the CMV promoter to determine whether PTTG functions as a transforming oncogene that is capable of initiating tumorigenesis. METHODS: Transgenic animals were generated by microinjection of PTTG transgene into the male pronucleus of FVB 0.5 day old embryos. Expression levels of PTTG in tissues of transgenic animals were analyzed using an immunohistochemical analysis. H&E staining and immunohistostaining were performed to examine the type of tumor in transgenic and PTTG transgenic/p53+/- animals. RESULTS: PTTG transgenic offspring (TgPTTG) were monitored for tumor development at various ages. H&E analysis was performed to identify the presence of cancer and hyperplastic conditions verified with the proliferation marker PCNA and the microvessel marker CD31. Immunohistochemistry was performed to determine transgene expression, revealing localization to the epithelium of the fallopian tube, with more generalized expression in the liver, lung, kidney, and spleen. At eight months of age, 2 out of 15 TgPTTG developed ovarian cancer, 2 out of 15 developed benign tumors, 2 out of 15 developed cervical dysplasia, and 3 out of 15 developed adenomyosis of the uterus. At ten months of age, 2 out of 10 TgPTTG developed adenocarcinoma of the ovary, 1 out of 10 developed a papillary serous adenocarcinoma, and 2 out of 10 presented with atypia of ovarian epithelial cells. Tumorigenesis is a multi-step process, often requiring multiple oncogenes and/or inactivation of tumor suppressor genes. Therefore, to understand the contribution of p53 to PTTG induced tumorigenesis, we crossbred TgPTTG to p53+/- mice and maintained those 8 to 10 months. TgPTTG/p53+/- animals developed sarcomas faster than p53+/- alone as well as different tumor types in addition to cervical carcinomas in situ in 10 out of 17 females. CONCLUSIONS: We conclude that while PTTG is a functional transforming oncogene, it requires an additional partner to effectively promote tumorigenesis through the loss of p53 include or between function or modulation.

Withaferin A synergizes the therapeutic effect of doxorubicin through ROS-mediated autophagy in ovarian cancer.

Application of doxorubicin (Dox) for the treatment of cancer is restricted due to its severe side effects. We used combination strategy by combining doxorubicin (Dox) with withaferin A (WFA) to minimize the ill effects of Dox. Treatment of various epithelial ovarian cancer cell lines (A2780, A2780/CP70 and CaOV3) with combination of WFA and Dox (WFA/DOX) showed a time- and dose-dependent synergistic effect on inhibition of cell proliferation and induction of cell death, thus reducing the dosage requirement of Dox. Combination treatment resulted in a significant enhancement of ROS production resulting in immense DNA damage, induction of autophagy analyzed by transmission electron microscope and increase in expression of autophagy marker LC3B, and culminated in cell death analyzed by cleaved caspase 3. We validated combination therapy on tumor growth using an in vitro 3Dimension (3D) tumor model and the more classic in vivo xenograft model of ovarian cancer. Both tumor models showed a 70 to 80% reduction in tumor growth compared to control or animals treated with WFA or Dox alone. Immunohistochemical analysis of the tumor tissues from animals treated with WFA/Dox combination showed a significant reduction in cell proliferation and formation of microvessels accompanied by increased in LC3B level, cleaved caspase 3, and DNA damage. Taken together, our data suggest that combining WFA with Dox decreases the dosage requirement of Dox, therefore, minimizing/eliminating the severe side effects associated with high doses of DOX, suggesting the application of this combination strategy for the treatment of ovarian and other cancers with no or minimum side effects.