FoxO3a as a positive prognostic marker and a therapeutic target in Tamoxifen-resistant breast cancer

Background: Resistance to endocrine treatments is a major clinical challenge in the management of estrogen receptor positive breast cancers. Although multiple mechanisms leading to endocrine resistance have been proposed, the poor outcome of this subgroup of patients demands additional studies. Methods: FoxO3a involvement in the acquisition and reversion of tamoxifen resistance was assessed in vitro in three parental ER+ breast cancer cells, MCF-7, T47D and ZR-75-1, in the deriving Tamoxifen resistant models (TamR) and in Tet-inducible TamR/FoxO3a stable cell lines, by growth curves, PLA, siRNA, RT-PCR, Western blot, Immunofluorescence, Transmission Electron Microscopy, TUNEL, cell cycle, proteomics analyses and animal models. FoxO3a clinical relevance was validated in silico by Kaplan−Meier survival curves. Results: Here, we show that tamoxifen resistant breast cancer cells (TamR) express low FoxO3a levels. The hyperactive growth factors signaling, characterizing these cells, leads to FoxO3a hyper-phosphorylation and subsequent proteasomal degradation. FoxO3a re-expression by using TamR tetracycline inducible cells or by treating TamR with the anticonvulsant lamotrigine (LTG), restored the sensitivity to the antiestrogen and strongly reduced tumor mass in TamR-derived mouse xenografts. Proteomics data unveiled novel potential mediators of FoxO3a anti-proliferative and pro-apoptotic activity, while the Kaplan−Meier analysis showed that FoxO3a is predictive of a positive response to tamoxifen therapy in Luminal A breast cancer patients. Conclusions: Altogether, our data indicate that FoxO3a is a key target to be exploited in endocrine-resistant tumors. In this context, LTG, being able to induce FoxO3a, might represent a valid candidate in combination therapy to prevent resistance to tamoxifen in patients at risk

Probiotics for prevention of necrotizing enterocolitis in preterm infants: systematic review and meta-analysis.

Necrotizing enterocolitis (NEC) affects predominantly preterm infants, who have specific risk factors leading to intestinal dysbiosis. Manipulations of gut microbiota through probiotics have the potential to prevent NEC. The aim of this systematic review and meta-analysis was to evaluate the effect of probiotics for NEC prevention in preterm infants, with a focus on specific strains, microbiological strength of currently available studies, and high-risk populations. PubMed and the Cochrane Library were searched for trials published within 4th February 2015. Randomized-controlled trials reporting on NEC and involving preterm infants who were given probiotics in the first month of life were included in the systematic review. Twenty-six studies were suitable for inclusion in the meta-analysis. Data about study design, population, intervention and outcome were extracted and summarized independently by two observers. Study quality and quality of evidence were also evaluated. Fixed-effects models were used and random-effects models where significant heterogeneity was present. Subgroup analyses were performed to explore sources of heterogeneity among studies. Results were expresses as risk ratio (RR) with 95 % confidence interval (CI). The main outcome was incidence of NEC stage 652 according to Bell\u2019s criteria. Probiotics prevented NEC in preterm infants (RR 0.47 [95 % CI 0.36\u20130.60], p\u2009<\u20090.00001). Strain-specific sub-meta-analyses showed a significant effect for Bifidobacteria (RR 0.24 [95 % CI 0.10\u20130.54], p\u2009=\u20090.0006) and for probiotic mixtures (RR 0.39 [95 % CI 0.27\u20130.56], p\u2009<\u20090.00001). Probiotics prevented NEC in very-low-birth-weight infants (RR 0.48 [95 % CI 0.37\u20130.62], p\u2009<\u20090.00001); there were insufficient data for extremely-low-birth-weight infants. The majority of studies presented severe or moderate microbiological flaws. Probiotics had an overall preventive effect on NEC in preterm infants. However, there are still insufficient data on the specific probiotic strain to be used and on the effect of probiotics in high-risk populations such as extremely-low-birth-weight infants, before a widespread use of these products can be recommended

A Clinical and Biological Guide for Understanding Chemotherapy-Induced Alopecia and its Prevention

Chemotherapy-induced alopecia (CIA) is the most visibly distressing side effect of commonly administered chemotherapeutic agents. As psychological health has huge relevance on lifestyle, diet and self-esteem, it is important for clinicians to fully appreciate the psychological burden that CIA can place on patients. Here, for the first time, we provide a comprehensive review encompassing the molecular characteristics of the human hair follicle (HF), how different anticancer agents damage the HF to cause CIA, subsequent HF pathophysiology and we assess known and emerging prevention modalities that have aimed to reduce or prevent CIA. We argue that, at present, scalp cooling is the only safe and FDA-cleared modality available, and we highlight the extensive available clinical and experimental (biological) evidence for its efficacy. The likelihood of a patient that uses scalp cooling during chemotherapy maintaining enough hair to not require a wig is approximately 50%. This is despite different types of chemotherapy regimens, patient-specific differences and possible lack of staff experience in effectively delivering scalp cooling. The increased use of scalp cooling and an understanding of how to deliver it most effectively to patients has enormous potential to ease the psychological burden of CIA, until other, more efficacious, equally safe treatments become available

ZD6474 reverses multidrug resistance by directly inhibiting the function of P-glycoprotein

P-glycoprotein (P-gp) pumps multiple types of drugs out of the cell, using energy generated from ATP, and confers multidrug resistance (MDR) on cancer cells. ZD6474 is an orally active, selective inhibitor of the vascular endothelial growth factor receptor, epidermal growth factor receptor, and rearranged during transfection tyrosine kinases. This study was designed to examine whether ZD6474 reverses P-gp-mediated MDR in cancer cells. Here, we show that clinically achievable levels of ZD6474 reverse P-gp-mediated MDR of the P-gp-overexpressing cell lines derived from breast cancer, MCF-7/adriamycin (ADR), and human oral epidermoid carcinoma, KBV200 to ADR, docetaxel, and vinorelbine. This ability to reverse the P-gp-mediated resistance is comparable to that of another frequently used reversal agent known as verapamil. ZD6474 itself moderately inhibits the proliferation of both MCF-7 and MCF-7/ADR cells with almost equal activity, but its inhibitory effect is not altered by co-incubation with verapamil, suggesting that ZD6474 may not be a substrate of P-gp. In addition, ZD6474 increases the intracellular accumulation of the P-gp substrate, rhodamine-123, and ADR, by enhancing the uptake and/or decreasing the efflux of these compounds in resistant cells. Further studies show that ZD6474 stimulates ATPase activity in a dose-dependent manner, which is required for the proper function of P-gp. In contrast, ZD6474 does not inhibit the expression level of P-gp. Our results suggest that ZD6474 is capable of reversing MDR in cancer cells by directly inhibiting the function of P-gp, a finding that may have clinical implications for ZD6474

A phase II study to determine the ability of gefitinib to reverse fluoropyrimidine resistance in metastatic colorectal cancer (the INFORM study)

There are data suggesting that inhibition of epidermal growth factor receptor (EGFR) tyrosine kinase signalling may reverse resistance to fluoropyrimidine treatment. To investigate this further, the INFORM study was an open-label, non-comparative phase II study of gefitinib (Iressa, ZD1839; AstraZeneca, Wilmington, DE, USA) 250 mg daily in combination with 5-fluorouracil (5-FU administered as an intravenous 400 mg m−2 bolus injection followed by 2800 mg m−2 infusion over 46 h and folinic acid administered as a 350 mg infusion over 2 h) every 2 weeks for up to 12 cycles in 24 patients with metastatic colorectal cancer refractory to previous fluoropyrimidine treatment. There were no objective responses. The stable disease rate was 37.5% (95% CI: 18.80, 59.41), median progression-free survival measured 116 days and overall survival was 226 days. Quality of life was unchanged compared to baseline values, and the commonest toxicities were diarrhoea, rash and fatigue with 7 out of 24 (29%) patients having a grade 3 or 4 toxicity. Gefitinib does not sensitise patients with fluoropyrimidine refractory metastatic colorectal cancer to 5-FU chemotherapy

Natural orifice surgery: initial clinical experience

Natural orifice translumenal endoscopic surgery (NOTES) has moved quickly from preclinical investigation to clinical implementation. However, several major technical problems limit clinical NOTES including safe access, retraction and dissection of the gallbladder, and clipping of key structures. This study aimed to identify challenges and develop solutions for NOTES during the initial clinical experience. Under an Institutional Review Board (IRB)-approved protocol, patients consented to a natural orifice operation for removal of either the gallbladder or the appendix via either the vagina or the stomach using a single umbilical trocar for safety and assistance. Nine transvaginal cholecystectomies, one transgastric appendectomy, and one transvaginal appendectomy have been completed to date. All but one patient were discharged on postoperative day 1 as per protocol. No complications occurred. The limited initial evidence from this study demonstrates that NOTES is feasible and safe. The addition of an umbilical trocar is a bridge allowing safe performance of NOTES procedures until better instruments become available. The addition of a flexible long grasper through the vagina and a flexible operating platform through the stomach has enabled the performance of NOTES in a safe and easily reproducible manner. The use of a uterine manipulator has facilitated visualization of the cul de sac in women with a uterus to allow for safe transvaginal access

The Effect of Immune Selection on the Structure of the Meningococcal Opa Protein Repertoire

The opa genes of the Gram negative bacterium Neisseria meningitidis encode Opacity-associated outer membrane proteins whose role is to promote adhesion to the human host tissue during colonisation and invasion. Each meningococcus contains 3–4 opa loci, each of which may be occupied by one of a large number of alleles. We analysed the Opa repertoire structure in a large, well-characterised collection of asymptomatically carried meningococci. Our data show an association between Opa repertoire and meningococcal lineages similar to that observed previously for meningococci isolated from cases of invasive disease. Furthermore, these Opa repertoires exhibit discrete, non-overlapping structure at a population level, and yet low within-repertoire diversity. These data are consistent with the predictions of a mathematical model of strong immune selection upon a system where identical alleles may occupy different loci

Corneal Epithelium Expresses a Variant of P2X7 Receptor in Health and Disease

Improper wound repair of the corneal epithelium can alter refraction of light resulting in impaired vision. We have shown that ATP is released after injury, activates purinergic receptor signaling pathways and plays a major role in wound closure. In many cells or tissues, ATP activates P2X7 receptors leading to cation fluxes and cytotoxicity. The corneal epithelium is an excellent model to study the expression of both the full-length P2X7 form (defined as the canonical receptor) and its truncated forms. When Ca2+ mobilization is induced by BzATP, a P2X7 agonist, it is attenuated in the presence of extracellular Mg2+ or Zn2+, negligible in the absence of extracellular Ca2+, and inhibited by the competitive P2X7 receptor inhibitor, A438079. BzATP enhanced phosphorylation of ERK. Together these responses indicate the presence of a canonical or full-length P2X7 receptor. In addition BzATP enhanced epithelial cell migration, and transfection with siRNA to the P2X7 receptor reduced cell migration. Furthermore, sustained activation did not induce dye uptake indicating the presence of truncated or variant forms that lack the ability to form large pores. Reverse transcription-polymerase chain reaction and Northern blot analysis revealed a P2X7 splice variant. Western blots identified a full-length and truncated form, and the expression pattern changed as cultures progressed from monolayer to stratified. Cross-linking gels demonstrated the presence of homo- and heterotrimers. We examined epithelium from age matched diabetic and non-diabetic corneas patients and detected a 4-fold increase in P2X7 mRNA from diabetic corneal epithelium compared to non-diabetic controls and an increased trend in expression of P2X7variant mRNA. Taken together, these data indicate that corneal epithelial cells express full-length and truncated forms of P2X7, which ultimately allows P2X7 to function as a multifaceted receptor that can mediate cell proliferation and migration or cell death

Modulation of the surface proteome through multiple ubiquitylation pathways in African Trypanosomes

Recently we identified multiple suramin-sensitivity genes with a genome wide screen in Trypanosoma brucei that includes the invariant surface glycoprotein ISG75, the adaptin-1 (AP-1) complex and two deubiquitylating enzymes (DUBs) orthologous to ScUbp15/HsHAUSP1 and pVHL-interacting DUB1 (type I), designated TbUsp7 and TbVdu1, respectively. Here we have examined the roles of these genes in trafficking of ISG75, which appears key to suramin uptake. We found that, while AP-1 does not influence ISG75 abundance, knockdown of TbUsp7 or TbVdu1 leads to reduced ISG75 abundance. Silencing TbVdu1 also reduced ISG65 abundance. TbVdu1 is a component of an evolutionarily conserved ubiquitylation switch and responsible for rapid receptor modulation, suggesting similar regulation of ISGs in T. brucei. Unexpectedly, TbUsp7 knockdown also blocked endocytosis. To integrate these observations we analysed the impact of TbUsp7 and TbVdu1 knockdown on the global proteome using SILAC. For TbVdu1, ISG65 and ISG75 are the only significantly modulated proteins, but for TbUsp7 a cohort of integral membrane proteins, including the acid phosphatase MBAP1, that is required for endocytosis, and additional ISG-related proteins are down-regulated. Furthermore, we find increased expression of the ESAG6/7 transferrin receptor and ESAG5, likely resulting from decreased endocytic activity. Therefore, multiple ubiquitylation pathways, with a complex interplay with trafficking pathways, control surface proteome expression in trypanosomes