PO-473 Quantification of ERCC1-XPF complexes in ovarian cancer xenografts with different sensitivity to cisplatin

Introduction Epithelial ovarian cancer is the most lethal gynaecological cancer due to the development of resistance to a platinum based therapy. As DNA repair capacity is a key determinant for the cellular response to platinum (DDP) agents, DNA repair functional assays are required to study its relevance in DDP resistance. We set up a proximity ligation assay (PLA) to study the activity of nucleotide excision repair (NER) in patient derived ovarian carcinoma xenografts (PDXs) sensitive (S) and resistant (R) to DDP. Material and methods Patient derived xenografts from fresh ovarian carcinomas were recently established in our laboratory. DDP antitumour activity was evaluated in most of the PDXs. Mice were sacrificed when tumours reached 1,5–2 gr. Tumours were fixed in formalin and paraffin embedded (FFPE). PLA was performed on tumour slides, using DuolinkII reagents (Sigma-Aldrich) and following the manufacturer instructions. PLA detects the presence of the protein complexes ERCC1-XPF, that are quantified as foci per nucleus and represent a biomarker of NER activity. Images were acquired by Olympus Virtual Slider (Olympus) and analysed with ImageJ software. Statistical analysis was performed with GraphPad Prism7. Results and discussions Our xenobank comprises PDXs with different response to DDP: MNHOC266 and MNHOC230 are very sensitive to the drug, while MNHOC315 is resistant. We also obtained three sublines resistant to DDP (MNHOC124R, MNHOC124LPR and MNHOC239R) starting from sensitive PDXs (MNHOC124S, MNHOC124LPS and MNHOC239S), after several in vivo drug treatments. Statistically significant higher level of ERCC1-XPF foci could be observed in MNHOC124R and MNHOC124LPR as compared to their sensitive counterparts. No differences were observed between MNHOC239S and R PDXs, even if the number of ERCC1-XPF foci in MNHOC239S were statistically higher than the ones observed in MNHOC124S and in MNHOC124LPS. MNHOC266 and MNHOC230 showed levels of foci comparable to those of MNHOC124S and MNHOC124LPS. mRNA and protein levels of the different isoforms of ERCC1 and of XPF were not different among the PDXs studied. Conclusion PLA for the detection of ERCC1-XPF complexes was set up in FFPE xenograft tumour slides. These preliminary results highlight a possible link between DDP resistance and higher NER activity that need to be confirmed in a wider panel of PDXs. In addition, these data confirm the importance to develop functional assays to directly evaluate the activity of different DNA repair pathways to predict DDP activity

Recent Insights into Mucinous Ovarian Carcinoma

Ovarian mucinous tumors represent a group of rare neoplasms with a still undefined cell of origin but with an apparent progression from benign to borderline to carcinoma. Even though these tumors are different from the other histological subtypes of epithelial ovarian neoplasms, they are still treated with a similar chemotherapeutic approach. Here, we review its pathogenesis, molecular alterations, (differential) diagnosis, clinical presentation and current treatment, and how recent molecular and biological information on this tumor might lead to better and more specific clinical management of patients with mucinous ovarian carcinoma

Glyphosate ingestion causing multiple organ failure: a near-fatal case report

A 55 years old man self-presented to our Emergency Department (ED) reporting an attempted suicide by cutting the left forearm veins and ingesting approximately 200 mL of an herbicide (Myrtos\uae, containing 36% of glyphosate as isopropylamine salt). Laboratory tests showed metabolic acidosis. Hydration with normal saline and alkalinization with sodium bicarbonate was started according to suggestion of the poison control center, since an antidote was unavailable. Cardiorespiratory condition gradually worsened, so that non-invasive positive pressure ventilation (NIPPV) was applied and infusion of fluids was established. Nevertheless, the patient deteriorated and he needed to be transferred to the Intensive Care Unit (ICU), where he underwent orotracheal intubation and invasive mechanical ventilation. Noradrenaline and adrenaline were infused and fluid resuscitation with crystalloids was incremented. An esophagogastroduodenoscopy (EGD) showed diffuse mucosal erosions of upper digestive tract. No signs of visceral perforation were found during ICU stay. In the following days, the clinical conditions improved and a new EGD showed marked improvement of erosive lesions. After 12 days of ICU stay, the patient was extubated and then transferred to the Psychiatric Unit, in good clinical conditions. Gliphosate ingestion is associated with rapid development of multiple organ failure (MOF). Since an effective antidote is unavailable, major attention should be placed to aggressive life-support care and careful monitoring of complications

Yeti, an essential Drosophila melanogaster gene, encodes a protein required for chromatin organization

The evolutionarily conserved family of Bucentaur (BCNT) proteins exhibits a widespread distribution in animal and plants, yet its biological role remains largely unknown. Using Drosophila melanogaster as a model organism, we investigated the in vivo role of the Drosophila BCNT member called YETI. We report that loss of YETI causes lethality before pupation and defects in higher-order chromatin organization, as evidenced by severe impairment in the association of histone H2A.V, nucleosomal histones and epigenetic marks with polytene chromosomes. We also find that YETI binds to polytene chromosomes through its conserved BCNT domain and interacts with the histone variant H2A.V, HP1a and Domino-A (DOM-A), the ATPase subunit of the DOM/Tip60 chromatin remodeling complex. Furthermore, we identify YETI as a downstream target of the Drosophila DOM-A. On the basis of these results, we propose that YETI interacts with H2A.V-exchanging machinery, as a chaperone or as a new subunit of the DOM/Tip60 remodeling complex, and acts to regulate the accumulation of H2A.V at chromatin sites. Overall, our findings suggest an unanticipated role of YETI protein in chromatin organization and provide, for the first time, mechanistic clues on how BCNT proteins control development in multicellular organisms