Deciphering The Role of miR-506 in Ovarian Cancer Drug Sensitivity

Although the incidence is quite low, epithelial ovarian cancer (EOC) is the most lethal gynaecologic malignancy. The unfavourable prognosis and the high mortality rate associated with EOC are mainly owed to late diagnosis, frequent relapse and development of chemoresistance. Indeed, most of the patients who achieve a complete response to first-line platinum-based treatment eventually relapse often developing platinum-resistant disease. Due to their master regulatory role, miRNAs are considered powerful tools to obtain representative molecular portraits of specific tumour characteristics and behaviours. My laboratory performed microRNA expression profiles on advanced stage EOC patients and a cluster of miRNAs, including miR-506, located on ChrXq27.3 was identified as down-regulated in EOC early relapsing patients. Since I observed that expression of miR-506 was associated with EOC patients’ sensitivity to platinum treatment, the overall aim of this thesis was to better characterize the role of this miRNA in regulating response to chemotherapy. Among the miR-506 predicted targets, I identified several genes involved in DNA damage repair (DDR) pathway, like RAD51, RAD17, CHEK1 and WEE1 and I concentrated on genes not previously studied in EOC. I validated RAD17 as a direct target of miR-506 and identified the miR-506-RAD17 axis as relevant in chemosensitising EOC cells to different treatments. I demonstrated that miR-506 expression, by targeting RAD17, was able to mediate sensitisation to platinum treatment and accordingly RAD17 silencing exerted the same effect. MiR-506 expression in EOC cells led to a reduced ability to properly sense DNA damage following platinum treatment causing mitotic defects, micronuclei formation, and impairing the signalling cascade responsible for G2/M checkpoint activation upon DNA damage insults. This behaviour, recapitulating a BRCAness phenotype, would allow propagation of cells with unrepaired DNA damage with the subsequent sensitisation to DNA damaging drugs. Furthermore, miR-506 expression, by regulating RAD17, impairs ATM signalling pathway, sensitising EOC cells to PARP inhibitor olaparib. Acting in the same way, miR-506 expression was synthetic lethal with Chk1 and Wee1 checkpoint kinases inhibitors in agreement with recent data reporting RAD17 depletion to be synthetically lethal with these small molecules. Accordingly, RAD17 down-modulation phenocopied the effect of miR-506 expression. Also combination treatments of Checkpoint kinases inhibitors with platinum resulted to be synergistic. Together the findings presented in this thesis support miR-506 as a key node in regulating DDR pathway in response to drug treatments and provide the rationale for its use to select EOC patients with BRCAness phenotype for efficient personalized therapeutic treatments

Abnormal hippocampal melatoninergic system: a potential link between absence epilepsy and depression-like behavior in WAG/Rij rats?

Absence epilepsy and depression are comorbid disorders, but the molecular link between the two disorders is unknown. Here, we examined the role of the melatoninergic system in the pathophysiology of spike and wave discharges (SWDs) and depression-like behaviour in the Wistar Albino Glaxo from Rijswijk (WAG/Rij) rat model of absence epilepsy. In WAG/Rij rats, SWD incidence was higher during the dark period of the light-dark cycle, in agreement with previous findings. However, neither pinealectomy nor melatonin administration had any effect on SWD incidence, suggesting that the melatoninergic system was not involved in the pathophysiology of absence-like seizures. Endogenous melatonin levels were lower in the hippocampus of WAG/Rij rats as compared to non-epileptic control rats, and this was associated with higher levels of melatonin receptors in the hippocampus, but not in the thalamus. In line with the reduced melatonin levels, cell density was lower in the hippocampus ofWAG/Rij rats and was further reduced by pinealectomy. As expected, WAG/Rij rats showed an increased depression-like behaviour in the sucrose preference and forced swim tests, as compared to non-epileptic controls. Pinealectomy abolished the difference between the two strains of rats by enhancing depression-like behaviour in non-epileptic controls. Melatonin replacement displayed a significant antidepressant-like effect in bothWAG/Rij and control rats. These findings suggest that a defect of hippocampal melatoninergic system may be one of the mechanisms underlying the depression-like phenotype inWAG/Rij rats and that activation of melatonin receptors might represent a valuable strategy in the treatment of depression associated with absence epilepsy

N-acetyl-cysteine, a drug that enhances the endogenous activation of group-II metabotropic glutamate receptors, inhibits nociceptive transmission in humans.

Emerging research seeking novel analgesic drugs focuses on agents targeting group-II metabotropic glutamate receptors (mGlu2 and mGlu3 receptors). N-Acetylcysteine (NAC) enhances the endogenous activation of mGlu2/3 receptors by activating the glial glutamate:cystine membrane exchanger. Here, we examined whether NAC inhibits nociceptive responses in humans and animals. We tested the effect of oral NAC (1.2 g) on thermal-pain thresholds and laser-evoked potentials in 10 healthy volunteers, according to a crossover, double-blind, placebo-controlled design, and the effect of NAC (100 mg/kg, i.p.) on the tail-flick response evoked by radiant heat stimulation in mice.In healthy subjects, NAC treatment left thermal-pain thresholds unchanged, but significantly reduced pain ratings to laser stimuli and amplitudes of laser-evoked potentials. NAC induced significantly greater changes in these measures than placebo. In the tail-flick test, NAC strongly reduced the nocifensive reflex response to radiant heat. The action of NAC was abolished by the preferential mGlu2/3 receptor antagonist, LY341495 (1 mg/kg, i.p.).Our findings show for the first time that NAC inhibits nociceptive transmission in humans, and does the same in mice by activating mGlu2/3 receptors. These data lay the groundwork for investigating the therapeutic potential of NAC in patients with chronic pain

Requests from Companies and Requirements for Design Education in Brazil: where do they meet?

In this paper, we study what companies request from applicants for graphic design positions in Brazil. Based on a document analysis of 371 job advertisements, we uncover 35 different types of requests which we structure in terms of (1) Design deliverables, (2) Knowledge and skills and (3) Personal traits. In addition, we explore how the content of job advertisements potentially can inform educational developments by reporting on a group interview with design educators. In the interview we explore the degree to which different requests in the advertisements are covered in a regulatory educational design policy document for higher education in Brazil. Our results show that requests for skills in 2D software, an ability to deliver print and digital design outcomes and knowledge of layout and photography are frequently occurring in the studied advertisements. We also describe how the educators could locate the majority of the requests in reviewing the educational policy document. We end the paper by discussing how job advertisements could be further studied and used by design educators and practitioners.Peer reviewe

In vitro mechanical stimulation to reproduce the pathological hallmarks of human cardiac fibrosis on a beating chip and predict the efficacy of drugs and advanced therapies

Cardiac fibrosis is one of the main causes of heart failure, significantly contributing to mortality. The discovery and development of effective therapies able to heal fibrotic pathological symptoms thus remain of paramount importance. Micro-physiological systems (MPS) are recently introduced as promising platforms able to accelerate this finding. Here a 3D in vitro model of human cardiac fibrosis, named uScar, is developed by imposing a cyclic mechanical stimulation to human atrial cardiac fibroblasts (AHCFs) cultured in a 3D beating heart-on-chip and exploited to screen drugs and advanced therapeutics. The sole provision of a cyclic 10% uniaxial strain at 1 Hz to the microtissues is sufficient to trigger fibrotic traits, inducing a consistent fibroblast-to-myofibroblast transition and an enhanced expression and production of extracellular matrix (ECM) proteins. Standard of care anti-fibrotic drugs (i.e., Pirfenidone and Tranilast) are confirmed to be efficient in preventing the onset of fibrotic traits in uScar. Conversely, the mechanical stimulation applied to the microtissues limit the ability of a miRNA therapy to directly reprogram fibroblasts into cardiomyocytes (CMs), despite its proved efficacy in 2D models. Such results demonstrate the importance of incorporating in vivo-like stimulations to generate more representative 3D in vitro models able to predict the efficacy of therapies in patients

Characterization of the ear canal bacterial flora present in hearing Aids (HA) wearing subjects

The use of hearing aids (HA) is considered a predisposing factor for ear microbial infections. We undertook this study to compare the presence and nature of the microbial flora inhabiting of ears of HA and non-HA (nHA) users. Swab samples of the ears of HA and nHA users were collected from the Institute of Otolaryngology, “Cattolica del Sacro Cuore” University “Agostino Gemelli”, Rome, Italy. Swab samples were taken from the ear canal of 57 HA and 33 nHA users. The components of the microbial flora present on each swab sample were identified and characterized at the level of species. A total of 41 different bacterial species were identified. A statistically significant prevalence of polymicrobial communities was found in ears presenting signs of inflammation (2.5 ± 1.7 vs 2.1 ± 1.3; P = 0.02) and in HA users (2.3 ± 1.2 vs 1.7 ± 1.0; P = 0.002). Few putative pathogens were detected. Candida albicans spp. was not isolated in our study. A small number of swab samples presented no microbial growth. Bacterial species isolated from HA users with and without inflammation were assayed for the ability to form a biofilm. Among gram-positive and gram-negative bacteria, S. aureus, CoNS, P. aeruginosa, and K. pneumoniae were found to be strong biofilm producers. S. aureus and P. aeruginosa, isolated only from the ears of HA and nHA users presenting signs of inflammation, were further analyzed for their antibiotic-resistance profile and characterized by the Multilocus Sequence Typing (MLST) assay. The highest rates of antibacterial resistance were in S. aureus to a penicillin (75.5%) and in P. aeruginosa, to amoxicillin-clavulanic acid, cefotaxime, ertapenem, tigecycline and trimethoprim-sulfamethoxazole (100%). Moreover, three S. aureus strains (37.5%) were methicillin-resistant (MRSA). Of the eight S. aureus isolates, we identified six sequence types (ST) indicating that 75% are likely independent clones. For what it concerned P. aeruginosa, six different STs were assigned. Interestingly, two out of the six strains presented newly identified ST values. This study sheds new light on the combined effect of the presence of HA devices and signs of external ear inflammation on the composition of the ear bacterial flora. Our results reinforce the need to practice careful hygiene of HA devices to prevent serious ear canal infections

Secondary metabolites from the endophytic fungus Talaromyces pinophilus

Endophytic fungi have a great influence on plant health and growth, and are an important source of bioactive natural compounds. Organic extracts obtained from the culture filtrate of an endophytic strain of Talaromyces pinophilus isolated from strawberry tree (Arbutus unedo) were studied. Metabolomic analysis revealed the presence of three bioactive metabolites, the siderophore ferrirubin, the platelet-aggregation inhibitor herquline B and the antibiotic 3-O-methylfunicone. The latter was the major metabolite produced by this strain and displayed toxic effects against the pea aphid Acyrthosiphon pisum (Homoptera Aphidiidae). This toxicity represents an additional indication that the widespread endophytic occurrence of T. pinophilus may be related to a possible role in defensive mutualism. Moreover, the toxic activity on aphids could promote further study on 3-O-methylfunicone, or its derivatives, as an alternative to synthetic chemicals in agriculture

Evolution of HER2-positive mammary carcinoma: HER2 loss reveals claudin-low traits in cancer progression

HER2-positive breast cancers may lose HER2 expression in recurrences and metastases. In this work, we studied cell lines derived from two transgenic mammary tumors driven by human HER2 that showed different dynamics of HER2 status. MamBo89HER2stable cell line displayed high and stable HER2 expression, which was maintained upon in vivo passages, whereas MamBo43HER2labile cell line gave rise to HER2-negative tumors from which MamBo38HER2loss cell line was derived. Both low-density seeding and in vitro trastuzumab treatment of MamBo43HER2labile cells induced the loss of HER2 expression. MamBo38HER2loss cells showed a spindle-like morphology, high stemness and acquired in vivo malignancy. A comprehensive molecular profile confirmed the loss of addiction to HER2 signaling and acquisition of an EMT signature, together with increased angiogenesis and migration ability. We identified PDGFR-B among the newly expressed determinants of MamBo38HER2loss cell tumorigenic ability. Sunitinib inhibited MamBo38HER2loss tumor growth in vivo and reduced stemness and IL6 production in vitro. In conclusion, HER2-positive mammary tumors can evolve into tumors that display distinctive traits of claudin-low tumors. Our dynamic model of HER2 status can lead to the identification of new druggable targets, such as PDGFR-B, in order to counteract the resistance to HER2-targeted therapy that is caused by HER2 loss

Oncological outcomes in fertility-sparing treatment in stage IA-G2 endometrial cancer

BACKGROUND: The gold standard treatment for early-stage endometrial cancer (EC) is hysterectomy with bilateral salpingo-oophorectomy (BSO) with lymphadenectomy. In selected patients desiring pregnancy, fertility-sparing treatment (FST) can be adopted. Our review aims to collect the most incisive studies about the possibility of conservative management for patients with grade 2, stage IA EC. Different approaches can be considered beyond demolition surgery, such as local treatment with levonorgestrel-releasing intra-uterine device (LNG-IUD) plus systemic therapy with progestins. STUDY DESIGN: Our systematic review was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. PubMed, EMBASE, and Scopus databases were consulted, and five studies were chosen based on the following criteria: patients with a histological diagnosis of EC stage IA G2 in reproductive age desiring pregnancy and at least one oncological outcome evaluated. Search imputes were “endometrial cancer” AND “fertility sparing” AND “oncologic outcomes” AND “G2 or stage IA”. RESULTS: A total of 103 patients were included and treated with a combination of LNG-IUD plus megestrol acetate (MA) or medroxyprogesterone acetate (MPA), gonadotrophin-releasing hormone (GnRH) plus MPA/MA, hysteroscopic resectoscope (HR), and dilation and curettage (D&C). There is evidence of 70% to 85% complete response after second-round therapy prolongation to 12 months. CONCLUSIONS: Conservative measures must be considered temporary to allow pregnancy and subsequently perform specific counseling to adopt surgery. Fertility-sparing management is not the current standard of care for young women with EC. It can be employed for patients with early-stage diseases motivated to maintain reproductive function. Indeed, the results are encouraging, but the sample size must be increased