PO-040 Characterisation of cdk12 knocked out ovarian cancer cell lines

Introduction While cyclin-dependent kinases (CDKs) have a key role in promoting/controlling transition between the different phases of the cell cycle, transcriptional kinases, like CDK12, are mainly involved in gene transcription. CDK12 has been shown to regulate the expression of genes involved in DNA damage and to maintain genomic stability. Impairment of CDK12 activity is synergic with PARP inhibitor and cisplatin treatments in different cellular systems. We here aimed to generate ovarian cancer cell lines knocked out (KO) for CDK12 to understand its role in ovarian cancer and in response to chemotherapy. Material and methods A2780 and SKOV3 CDK12 KO clones were generated with CRISPR/Cas9 technology. Cell cycle analysis was evaluated by standard flow cytometric methods and DNA repair genes levels by Real Time PCR. Caspase 3 activity was measured to detect apoptosis with a luminescence-based assay. Cytotoxicity experiments were performed treating cells with different drug concentrations and evaluating cell survival after 72 hours by MTS assay. For in vivo studies 7.5 millions of cells were transplanted subcutaneously in nude mice and animals were monitored for tumour appearance and growth. Results and discussions We obtained 2 CDK12 KO ovarian cancer clones, A2780 KO and SKOV3 KO, out of more than 300 clones screened. The cell growth of both A2780 KO and SKOV3 KO cells is slower than the wild type (WT) cells, they have a less clonogenic ability and a tetraploid DNA content. Both CDK12 KO clones have a higher basal caspase activity than the WT cell lines, indicative of higher basal induction of apoptosis, while no increase in autophagy or senescence is observed. Both CDK12 KO clones show a decreased expression in BRCA1 and FANCD2 DNA repair genes than the WT cells. Cytotoxic experiments with anticancer agents with different mechanism of action show that both KO clones are less sensitive to ATM, CHK1 and WEE1 inhibitors treatment as compared to WT cells, while platinum and PARP inhibitors show similar cytotoxic activity in KO and WT cells. Interestingly enough, when KO clones were transplanted in nude mice, no tumour take was observed. Conclusion We were able to obtain CDK12 KO cells. We think that these models could help in disclosing new roles of CDK12 in ovarian carcinoma and may represent a useful tool to study new combination therapies for tumours with CDK12 mutations

Unique features of the mode of action of ET-743

This paper describes the current knowledge of the primary mode of action of a natural product, ecteinascidin 743 (ET-743), derived from the marine tunicate Ecteinascidia turbinata. ET-743 was initially selected for preclinical development because of its potent antitumor activity observed against several human solid tumor types. In vitro, the drug is cytotoxic in the nanomolar range, and in the case of some very sensitive cell lines, in the picomolar range. The large potency differences observed among several solid tumor types indicate that this compound possesses some tumor selectivity, but the molecular basis of these differential effects remains to be elucidated. The present studies were undertaken to evaluate the mechanism of action of ET-743 in this context. The available information on ET-743 binding to DNA and its effects on transcriptional regulation point to a unique behavior of this drug, as it independently affects specific gene transcription in a promoter-dependent way. In addition, ET-743 shows a peculiar pattern of selectivity in cells with different defects in their DNA-repair pathways. These results highlight a unique property of ET-743, possibly explaining why it possesses antitumor activity against tumors that are refractory to standard anticancer drugs, all of which certainly act by mechanisms that are different from that of ET-743

Flow regulation increases food-chain length through omnivory mechanisms in a Mediterranean river network

[EN] Dams fragment river systems worldwide, and Mediterranean-climate rivers, characterised by highly seasonal hydrographs and adapted biotas, are particularly impacted by flow regulation. Whereas the effects of flow regulation on hydrology, sediment transport and biodiversity have long been examined, responses at the food-web level remain understudied. Environmental variation is a key control of food-web structure. Thus, we predicted that flow regulation would impact food-chain length (FCL) via changes in the flow variation regime, and we tested this prediction in a set of flow unregulated to completely regulated reaches in a Mediterranean river basin. In each reach, we characterised flow variation, together with two other putative controls of FCL (productivity and habitat size). We combined community data with carbon and nitrogen stable isotopes to estimate food-chain length, and Bayesian mixing models allowed estimates of dietary proportions of consumers. Flow variation was paramount in controlling FCL in the studied river network, and this same control largely explained the degree of omnivory among top predators. Thus, omnivory mechanisms were the main proximate structural mechanism allowing shifts in food-web structure and linking disturbance regimes to FCL. Our results suggest that flow regulation in Mediterranean rivers may impact food-web structure even when no significant changes in community composition are observed. If highly variable Mediterranean streams become increasingly affected by flow regulation, the resulting more stable conditions could enhance intraguild predation and thus lengthen riverine food chains.We thank Roberto Merciai, Jose Andres Lopez and Joan Font for their help in the field, Lina Ramirez-Solano and Emili Garcia-Berthou for their help with analyses and Marc Montenegro for the illustrations in Fig. 1. The Sabo Lab at Arizona State University and anonymous reviewers provided suggestions that improved the quality of the article. This study was funded by the Spanish Ministry of Economy and Competitiveness through the SCARCE project (CSD2009-00065). Authors acknowledge the support from the Catalan Government through the Consolidated Research Groups 'Fluvial Dynamics Research Group (2014 SGR 645)' and the 'Catalan Institute for Water Research (2014 SGR 291)'.Ruhí, A.; Muñoz, I.; Tornés, E.; Batalla, R.; Vericat, D.; Ponsati, L.; Acuña, V.... (2016). Flow regulation increases food-chain length through omnivory mechanisms in a Mediterranean river network. Freshwater Biology. 61(9):1536-1549. https://doi.org/10.1111/fwb.12794S1536154961

Valutazione al SEM dell’assorbimento di fluoro nei tessuti duri dentari in seguito ad irraggiamento con laser ad Erbio e a Diodi.

Gli incoraggianti risultati ottenuti nel presente lavoro portano a indicare come il trattamento dell’ipersensibilità dentinale con gel al fluoro e sorgenti laser, utilizzate secondo i protocolli indicati, rappresenti oggi il miglior standard clinico disponibile per taleindicazione. L’interpretazione dei risultati ottenuti porta ad ipotizzare il trattamento con gel di fluoro e laser a Diodi comedi elezione per il trattamento dell’ipersensibilità dentinaleai colletti per il maggior assorbimento di fluoro ottenuto. Iltrattamento con gel di fluoro e laser ad Erbio comunque nonè trascurabile, e lascia spazio per ulteriori analisi a diversiparametri d’applicazione

Electroconductive Hydrogel Based on Functional Poly(Ethylenedioxy Thiophene).

Poly(ethylene dioxythiophene) with functional pendant groups bearing double bonds is synthesized and employed for the fabrication of electroactive hydrogels with advantageous characteristics: covalently cross-linked porous 3D scaffolds with notable swelling ratio, appropriate mechanical properties, electroactivity in physiological conditions, and suitability for proliferation and differentiation of C2C12 cells. This is a new approach for the fabrication of conductive engineered constructs