Preclinical Test of Dacomitinib, an Irreversible EGFR Inhibitor, Confirms Its Effectiveness for Glioblastoma.

Glioblastomas (GBM) are devastating tumors in which there has been little clinical improvement in the last decades. New molecularly directed therapies are under development. EGFR is one of the most promising targets, as this receptor is mutated and/or overexpressed in nearly half of the GBMs. However, the results obtained with first-generation tyrosine-kinase inhibitors have been disappointing with no clear predictive markers of tumor response. Here, we have tested the antitumoral efficacy of a second-generation inhibitor, dacomitinib (PF299804, Pfizer), that binds in an irreversible way to the receptor. Our results confirm that dacomitinib has an effect on cell viability, self-renewal, and proliferation in EGFR-amplified ± EGFRvIII GBM cells. Moreover, systemic administration of dacomitinib strongly impaired the in vivo tumor growth rate of these EGFR-amplified cell lines, with a decrease in the expression of stem cell-related markers. However, continuous administration of the compound was required to maintain the antitumor effect. The data presented here confirm that dacomitinib clearly affects receptor signaling in vivo and that its strong antitumoral effect is independent of the presence of mutant receptor isoforms although it could be affected by the PTEN status (as it is less effective in a PTEN-deleted GBM line). Dacomitinib is being tested in second line for EGFR-amplified GBMs. We hope that our results could help to select retrospectively molecular determinants of this response and to implement future trials with dacomitinib (alone or in combination with other inhibitors) in newly diagnosed GBMs.This work was supported by grants from the Fundación Mutua-madrileña (FMM2011/89) to J.M. Sepúlveda and from Ministerio de Economía y Competitividad, Fondo de Investigación Sanitaria (FIS): PI12/00775 to P. Sánchez-Gómez and PI13/01258 to A. Hernández-Laín, and from Ministerio de Economía y Competitividad, Red Temática de Investigación Cooperativa en Cancer (RTICC) (RD12/0036/0027) to J.M. Sepúlveda, P. Sánchez-Gómez, A. Pérez-Núñez and A. Hernández-Laín.S

Attenuation of disease phenotype through alternative translation initiation in low-penetrance retinoblastoma

Hereditary predisposition to retinoblastoma (RB) is caused by germline mutations in the retinoblastoma 1 (RB1) gene and transmits as an autosomal dominant trait. In the majority of cases disease develops in greater than 90% of carriers. However, reduced penetrance with a large portion of disease-free carrier is seen in some families. Unambiguous identification of the predisposing mutation in these families is important for accurate risk prediction in relatives and their genetic counseling but also provides conceptual information regarding the relationship between the RB1 genotype and the disease phenotype. In this study we report a novel mutation detected in 10 individuals of an extended family, only three of whom are affected by RB disease. The mutation comprises a 23-basepair (bp) duplication in the first exon of RB1 (c.43_65dup) producing a frameshift in exon 1 and premature chain termination in exon 2. Mutations resulting in premature chain termination classically are associated with high penetrance disease, as message translation may not generate functional product and nonsense mediated RNA decay (NMD) frequently eliminates the mutant transcript. However, appreciable NMD does not follow from the mutation described here and transcript expression in tissue culture cells and translation in vitro reveals that alternative in-frame translation start sites involving Met113 and possibly Met233 are used to generate truncated RB1 products (pRB94 and pRB80), known and suspected to exhibit tumor suppressor activity. These results strongly suggest that modulation of disease penetrance in this family is achieved by internal translation initiation. Our observations provide the first example for rescue of a chain-terminating mutation in RB1 through alternative translation initiation

CT2A cell viability modulated by electromagnetic fields at extremely low frequency under no thermal effects

The effects produced by electromagnetic fields (EMFs) on human beings at extremely low frequencies (ELFs) have being investigated in the past years, across in vitro studies, using different cell lines. Nevertheless, the effects produced on cells are not clarified, and the cellular mechanisms and cell-signaling processes involved are still unknown. This situation has resulted in a division among the scientific community about the adequacy of the recommended level of exposure. In this sense, we consider that it is necessary to develop long-term exposure studies and check if the recommended levels of EMFs are under thermal effects. Hence, we exposed CT2A cells to different EMFs at different ELFs at short and long times. Our results showed frequency dependence in CT2A exposed during 24 h to a small EMF of 30 ?T equal to those originated by the Earth and frequency dependence after the exposure during seven days to an EMF of 100 µT at different ELFs. Particularly, our results showed a remarkable cell viability decrease of CT2A cells exposed to EMFs of 30 Hz. Nevertheless, after analyzing the thermal effects in terms of HSP90 expression, we did not find thermal damages related to the differences in cell viability, so other crucial cellular mechanism should be involved

Self-Renewal Inhibition in Breast Cancer Stem Cells: Moonlight Role of PEDF in Breast Cancer

Breast cancer is the leading cause of death among females in developed countries. Although the implementation of screening tests and the development of new therapies have increased the probability of remission, relapse rates remain high. Numerous studies have indicated the connection between cancer-initiating cells and slow cellular cycle cells, identified by their capacity to retain long labeling (LT+). In this study, we perform new assays showing how stem cell self-renewal modulating proteins, such as PEDF, can modify the properties, percentage of biomarker-expressing cells, and carcinogenicity of cancer stem cells. The PEDF signaling pathway could be a useful tool for controlling cancer stem cells’ self-renewal and therefore control patient relapse, as PEDF enhances resistance in breast cancer patient cells’ in vitro culture. We have designed a peptide consisting of the C-terminal part of this protein, which acts by blocking endogenous PEDF in cell culture assays. We demonstrate that it is possible to interfere with the self-renewal capacity of cancer stem cells, induce anoikis in vivo, and reduce resistance against docetaxel treatment in cancer patient cells in in vitro culture. We have also demonstrated that this modified PEDF protein produces a significant decrease in the percentage of expressed cancer stem cell markers

Pedf derived peptides affect colorectal cancer cell lines resistance and tumour re-growth capacity

Relapse after chemotherapy treatment depends on the cancer initiating cells (CICs). PEDF (Pigmented Epithelium Derived Factor) is an anti-angiogenic, neurotrophic and self-renewal regulator molecule, also involved in CICs biology. Acute and chronic exposition of colon cancer cell lines to CT/CTE PEDF-derived peptides decreased drug-resistance to conventional colorectal cancer treatments, such as oxaliplatin or irinotecan. We confirmed a reduction in the irinotecan and oxaliplatin IC50 doses for all tested tumour cell lines. After xenograft transplantation, CT/CTE treatments also produced a reduction in resistance to conventional chemotherapy treatments as in culture-assays. Metastatic capacity of these treated cell lines was so depleted. The PEDF signaling pathway could be a future therapeutic tool for use as an adjuvant therapy that decreases IC50 dosis, adverse effects and treatment costs. This pathway could also be involved in an increase of the time relapse in patients, decreased tumourigenicity, and decreased capacity to produce metastasis

Identification of the ligand of Pru p 3, a peach LTP

The allergen Pru p 3, a peach lipid transfer protein, has been well studied. However, its physiological function remains to be elucidated. Our results showed that Pru p 3 usually carries a lipid ligand that play an essential role in its function in plants. Using ESI-qToF, we observed that the ligand was a derivative of camptothecin binding to phytosphingosine, wich that is inserted into the hydrophobic tunnel of the protein. In addition, the described ligand displayed topoisomerase I activity inhibition and self-fluorescence, both recognized as camptothecin properties. During flower development, the highest expression of Pru p 3 was detected in the styles of pollinated flowers, in contrast to its non-expression in unpollinated pistils, where expression decreased after anthesis. During ripening, the expression of Pru p 3 were observed mainly in peel but not in pulp. In this sense, Pru p 3 protein was also localized in trichomes covering the fruit epidermis

Implication of Different Tumor Biomarkers in Drug Resistance and Invasiveness in Primary and Metastatic Colorectal Cancer Cell Lines

Protein expression profiles are directly related to the different properties of cells and are conditioned by the cellular niche. As an example, they are the cause of the characteristic cell plasticity, epithelium–mesenchymal transition (EMT), and drug resistance of cancer cells. This article characterizes ten biomarkers related to these features in three human colorectal cancer cell lines: SW-480, SW-620, and DLD-1, evaluated by flow cytometry; and in turn, resistance to oxaliplatin is studied through dose–response trials. The main biomarkers present in the three studied lines correspond to EpCAM, CD-133, and AC-133, with the latter two in low proportions in the DLD-1 line. The biomarker CD166 is present in greater amounts in SW-620 and DLD-1 compared to SW-480. Finally, DLD-1 shows high values of Trop2, which may explain the aggressiveness and resistance of these cells to oxaliplatin treatments, as EpCAM is also highly expressed. Exposure to oxaliplatin slows cell growth but also helps generate resistance to the treatment. In conclusion, the response of the cell lines is variable, due to their genetic variability, which will condition protein expression and cell growth. Further analyses in this area will provide important information for better understanding of patients’ cellular response and how to prevent resistance

Characterisation of a flavonoid ligand of the fungal protein Alt a 1

Spores of pathogenic fungi are virtually ubiquitous and cause human disease and severe losses in crops. The endophytic fungi Alternaria species produce host-selective phytotoxins. Alt a 1 is a strongly allergenic protein found in A. alternata that causes severe asthma. Despite the well-established pathogenicity of Alt a 1, the molecular mechanisms underlying its action and physiological function remain largely unknown. To gain insight into the role played by this protein in the pathogenicity of the fungus, we studied production of Alt a 1 and its activity in spores. We found that Alt a 1 accumulates inside spores and that its release with a ligand is pH-dependent, with optimum production in the 5.0-6.5 interval. The Alt a 1 ligand was identified as a methylated flavonoid that inhibits plant root growth and detoxifies reactive oxygen species. We also found that Alt a 1 changes its oligomerization state depending on the pH of the surrounding medium and that these changes facilitate the release of the ligand. Based on these results, we propose that release of Alt a 1 should be a pathogenic target in approaches used to block plant defenses and consequently to favor fungal entry into the plant

Biallelic MYH germline mutations as cause of Muir-Torre syndrome

Muir-Torre syndrome is a rare, inherited disease predisposing of gastrointestinal and cutaneous tumours, such as keratoacanthomas and sebaceous gland adenomas. Muir-Torre syndrome is usually inherited in an autosomal dominant fashion and associated with mutations in the mismatch repair genes, predominantly in MLH1 and MSH2 genes. This report describes a man who has multiple adenomatous colon polyps, a gastric cancer, multiple colorectal cancers and sebaceous adenomas caused by biallelic MYH germline mutations. This finding demonstrates that MYH gene analysis should be considered in Muir-Torre families where no mismatch repair gene mutations have been found. Furthermore, this report contributes to characterize the clinical phenotype caused by biallelic mutations in MYH gene, which may share with other hereditary colon cancer syndromes

Reunión científica del Programa Geomateriales: durabilidad y conservación de geomateriales del patrimonio construido, 9 de octubre 2013

El Programa Geomateriales Durabilidad y conservación del geomateriales del patrimonio construido, enmarcado en el IV Plan Regional de Investigación Científica e Innovación Tecnológica (IV PRICIT), fue concedido a través de su Convocatoria de Programas de I+D en Tecnologías 2009 (orden 679/2009). Iniciado el 1 de enero de 2010 y finalizado el 31 de mayo de 2014, el programa de investigación se plantea con una clara concepción experimental, pretendiendo dar solución y optimizando las actuaciones aplicadas para mejorar la durabilidad y conservación de los geomateriales utilizados en el patrimonio construido. El consorcio que integra el programa está constituido por grupos de investigación que desarrollan su labor en la Comunidad de Madrid, pertenecientes a la Agencia Estatal del Consejo Superior de Investigaciones Científicas (CSIC), a la Universidad Complutense de Madrid (UCM) y a la Universidad de Alcalá de Henares (UAH), otros grupos, laboratorios y/o centros tecnológicos ubicados en otras provincias españolas y en países europeos así como empresas privadas y otros organismos de colaboración. El programa ha conseguido alcanzar los objetivos científico-tecnológicos planteados, en gran parte mediante la colaboración de los diferentes grupos integrantes, jugando un papel esencial la participación también en el programa del Laboratorio de Petrofísica (RedLab 217). El presente volumen recoge las ponencias presentadas en la Reunión Científica del Programa Geomateriales, celebrada el 9 de octubre de 2013. Estas ponencias se organizaron fundamentalmente en base a los objetivos científicos del programa, incidiendo en las principales actividades realizadas y resultados obtenidos, y apostando por los jóvenes investigadores incorporados. En los resúmenes incluidos se pretende resaltar, por un lado, los resultados científicos de mayor relevancia, las investigaciones que han generado el desarrollo de patentes, las colaboraciones establecidas entre los diferentes grupos y/o empresas y las previsiones futuras. Por otro, aspectos relacionados con los objetivos de formación, como la realización de tesis doctorales y movilidad de investigadores, así como la participación conjunta en los objetivos de difusión y en actividades de gestión de los diferentes miembros del programa. El Programa Geomateriales ha supuesto un importante avance del conocimiento, permitiendo los logros científicos obtenidos posicionar al consorcio en unas condiciones muy favorables respecto a las diversas temáticas implicadas en la conservación del patrimonio. Este aspecto hace del consorcio el único a nivel nacional que aborda su estudio de forma integral y confirma, una vez más, la necesidad y conveniencia de crear equipos de trabajo que apuesten por la colaboración y conjunción de esfuerzos para alcanzar mayores y mejores logros.Peer Reviewe