In Vivo Eradication of Human BCR/ABL-Positive Leukemia Cells With an ABLKinase Inhibitor

BACKGROUND: The leukemia cells of approximately 95% of patients with chronic myeloid leukemia and 30%-50% of adult patients with acute lymphoblastic leukemia express the Bcr/Abl oncoprotein, which is the product of a fusion gene created by a chromosomal translocation [(9:22) (q34;q11)]. This oncoprotein expresses a constitutive tyrosine kinase activity that is crucial for its cellular transforming activity. In this study, we evaluated the antineoplastic activity of CGP57148B, which is a competitive inhibitor of the Bcr/Abl tyrosine kinase. METHODS: Nude mice were given an injection of the Bcr/Abl-positive human leukemia cell lines KU812 or MC3. Tumor-bearing mice were treated intraperitoneally or orally with CGP57148B according to three different schedules. In vitro drug wash-out experiments and in vivo molecular pharmacokinetic experiments were performed to optimize the in vivo treatment schedule. RESULTS: Treatment schedules administering CGP57148B once or twice per day produced some inhibition of tumor growth, but no tumor-bearing mouse was cured. A single administration of CGP57148B caused substantial (>50%) but short-lived (2-5 hours) inhibition of Bcr/Abl kinase activity. On the basis of the results from in vitro wash-out experiments, 20-21 hours was defined as the duration of continuous exposure needed to block cell proliferation and to induce apoptosis in these two leukemia cell lines. A treatment regimen assuring the continuous block of the Bcr/Abl phosphorylating activity that was administered over an 11-day period cured 87%-100% of treated mice. CONCLUSION: These data indicate that the continuous block of the oncogenic tyrosine kinase of Bcr/Abl protein is needed to produce important biologic effects in viv

Characterization of compound 584, an Abl kinase inhibitor with lasting effects

Background: Resistance to imatinib is an important clinical issue in the treatment of Philadelphia chromosomepositive leukemias which is being tackled by the development of new, more potent drugs, such as the dual Src/Abl tyrosine kinase inhibitors dasatinib and bosutinib and the imatinib analog nilotinib. In the current study we describe the design, synthesis and biological properties of an imatinib analog with a chlorine-substituted benzamide, namely compound 584 (cmp-584). Design and Methods: To increase the potency, we rationally designed cmp-584, a compound with enhanced shape complementarity with the kinase domain of Abl. cmp-584 was synthesized and characterized in vitro against a panel of 67 serine/threonine and tyrosine kinases using radioactive and enzyme-linked immunosorbent kinase assays. We studied inhibitory cellular activity using Bcr/Abl-positive human cell lines, murine transfectants in proliferation experiments, and a murine xenotransplanted model. Kinase assays on isolated Bcr/Abl protein were also performed. Finally, we used a wash-out approach on whole cells to study the binding kinetics of the inhibitor. Results: cmp-584 showed potent anti-Abl activity both on recombinant protein (IC50: 8 nM) and in cell-based assays (IC50: 0.1-10 nM). The drug maintained inhibitory activity against platelet-derived growth factor receptors and c-KIT and was also active against Lyn (IC50: 301 nM). No other kinase of the panel was inhibited at nanomolar doses. cmp-584 was 20- to 300-fold more active than imatinib in cells. This superior activity was evident in intact cells, in which full-length Bcr-Abl is present. In vivo experiments confirmed the activity of cmp-584. Wash-out experiments showed that short exposure to the drug impaired cell proliferation and Bcr-Abl phosphorylation for a substantially longer period of time than imatinib. Conclusions: The present results suggest a slower off-rate (dissociation rate) of cmp-584 compared to imatinib as an explanation for the increased cellular activity of the former. ©2008 Ferrata Storti Foundation

A functional gene expression analysis in epithelial sinonasal cancer: Biology and clinical relevance behind three histological subtypes

Epithelial sinonasal cancers (SNCs) are rare diseases with overlapping morphological features and a dismal prognosis. We aimed to investigate the expression differences among the histological subtypes for discerning their molecular characteristics. We selected 47 SNCs: (i) 21 nonkeratinizing squamous cell carcinomas (NKSCCs), (ii) 13 sinonasal neuroendocrine cancers (SNECs), and (iii) 13 sinonasal undifferentiated cancers (SNUCs). Gene expression profiling was performed by DASL (cDNA-mediated annealing, selection, extension, and ligation) microarray analysis with internal validation by quantitative RT-PCR (RT-qPCR). Relevant molecular patterns were uncovered by sparse partial-least squares discriminant analysis (sPLS-DA), microenvironment cell type (xCell), CIBERSORT, and gene set enrichment (GSEA) analyses. The first two sPLS-DA components stratified samples by histological subtypes. xCell highlighted increased expression of immune components (CD8 + effector memory cells, in SNUC) and \u201cother cells\u201d: keratinocytes and neurons in NKSCC and SNEC, respectively. Pathway enrichment was observed in NKSCC (six gene sets, proliferation related), SNEC (one gene set, pancreatic \u3b2-cells), and SNUC (twenty gene sets, some of them immune-system related). Major neuroendocrine involvement was observed in all the SNEC samples. Our high-throughput analysis revealed a good diagnostic ability to differentiate NKSCC, SNEC, and SNUC, but indicated that the neuroendocrine pathway, typical and pathognomonic of SNEC is also present at lower expression levels in the other two histological subtypes. The different and specific profiles may be exploited for elucidating their biology and could help to identify prognostic and therapeutic opportunities

A microRNA prognostic signature in patients with diffuse intrinsic pontine gliomas through non-Invasive liquid biopsy

SIMPLE SUMMARY: Diffuse intrinsic pontine glioma (DIPG) is a neuro-radiologically defined tumor of the brainstem, primarily affecting children, with most diagnoses occurring between 5 and 7 years of age. Surgical removal in DIPGs is not feasible. Subsequent tumor progression is almost universal and no biomarker for predicting the course of the disease has entered into clinical practice so far. Under these premises, it is essential to develop reliable biomarkers that are able to improve outcomes and stratify patients using non-invasive methods to determine tumor profiles. We designed a study assessing circulating miRNA expression by a high-throughput platform and divided patients into training and validation phases in order to disclose a potential signature with clinical impact. Our results for the first time have proved the usefulness of blood-circulating nucleic acids as powerful, easy-to-assay molecular markers of disease status in DIPG. ABSTRACT: Diffuse midline gliomas (DMGs) originate in the thalamus, brainstem, cerebellum and spine. This entity includes tumors that infiltrate the pons, called diffuse intrinsic pontine gliomas (DIPGs), with a rapid onset and devastating neurological symptoms. Since surgical removal in DIPGs is not feasible, the purpose of this study was to profile circulating miRNA expression in DIPG patients in an effort to identify a non-invasive prognostic signature with clinical impact. Using a high-throughput platform, miRNA expression was profiled in serum samples collected at the time of MRI diagnosis and prior to radiation and/or systemic therapy from 47 patients enrolled in clinical studies, combining nimotuzumab and vinorelbine with concomitant radiation. With progression-free survival as the primary endpoint, a semi-supervised learning approach was used to identify a signature that was also tested taking overall survival as the clinical endpoint. A signature comprising 13 circulating miRNAs was identified in the training set (n = 23) as being able to stratify patients by risk of disease progression (log-rank p = 0.00014; HR = 7.99, 95% CI 2.38–26.87). When challenged in a separate validation set (n = 24), it confirmed its ability to predict progression (log-rank p = 0.00026; HR = 5.51, 95% CI 2.03–14.9). The value of our signature was also confirmed when overall survival was considered (log-rank p = 0.0021, HR = 4.12, 95% CI 1.57–10.8). We have identified and validated a prognostic marker based on the expression of 13 circulating miRNAs that can shed light on a patient’s risk of progression. This is the first demonstration of the usefulness of nucleic acids circulating in the blood as powerful, easy-to-assay molecular markers of disease status in DIPG. This study provides Class II evidence that a signature based on 13 circulating miRNAs is associated with the risk of disease progression

Butyrate Attenuates Lipopolysaccharide-Induced Inflammation in Intestinal Cells and Crohn's Mucosa through Modulation of Antioxidant Defense Machinery

Oxidative stress plays an important role in the pathogenesis of inflammatory bowel disease (IBD), including Crohn's disease (CrD). High levels of Reactive Oxygen Species (ROS) induce the activation of the redox-sensitive nuclear transcription factor kappa-B (NF-κB), which in turn triggers the inflammatory mediators. Butyrate decreases pro-inflammatory cytokine expression by the lamina propria mononuclear cells in CrD patients via inhibition of NF-κB activation, but how it reduces inflammation is still unclear. We suggest that butyrate controls ROS mediated NF-κB activation and thus mucosal inflammation in intestinal epithelial cells and in CrD colonic mucosa by triggering intracellular antioxidant defense systems. Intestinal epithelial Caco-2 cells and colonic mucosa from 14 patients with CrD and 12 controls were challenged with or without lipopolysaccaride from Escherichia Coli (EC-LPS) in presence or absence of butyrate for 4 and 24 h. The effects of butyrate on oxidative stress, p42/44 MAP kinase phosphorylation, p65-NF-κB activation and mucosal inflammation were investigated by real time PCR, western blot and confocal microscopy. Our results suggest that EC-LPS challenge induces a decrease in Gluthation-S-Transferase-alpha (GSTA1/A2) mRNA levels, protein expression and catalytic activity; enhanced levels of ROS induced by EC-LPS challenge mediates p65-NF-κB activation and inflammatory response in Caco-2 cells and in CrD colonic mucosa. Furthermore butyrate treatment was seen to restore GSTA1/A2 mRNA levels, protein expression and catalytic activity and to control NF-κB activation, COX-2, ICAM-1 and the release of pro-inflammatory cytokine. In conclusion, butyrate rescues the redox machinery and controls the intracellular ROS balance thus switching off EC-LPS induced inflammatory response in intestinal epithelial cells and in CrD colonic mucosa

Quality of life in post-bariatric surgery patients undergoing aesthetic abdomi-noplasty: Our experience

Background/aim of the study: Aesthetic post-bariatric surgery abdominoplasty aims to reshape the abdominal wall in order to improve the patient's appearance and self-image. Patients who have undergone this surgery report significant improvements in self-esteem and quality of life. Material and Methods: The authors evaluated the quality of life in 30 patients (22 women and 8 men) aged 24-79 years (mean age: 50.5 years) undergoing aesthetic post-bariatric surgery abdominoplasty between January 2012 and September 2015. Quality of life and body image were measured with two self-report questionnaires: a basic questionnaire (BQ) and the Body Image As-sessment-Obesity (BIA-O). Results: At the end of the study, 58% of patients reported complete satisfaction with the aesthetic results, while 23% of patients expressed dissatisfaction with the surgical scars. Analysis of the responses to the BIA-O, showed that patients had a better and slimmer body image perception after surgery than they had preoperatively. Conclusion:Although our study sample was small, we could demonstrate that most patients undergoing aesthetic post-bariatric surgery abdominoplasty experience improvement in body image perception and quality of life