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BACKGROUND: Elotuzumab plus lenalidomide and dexamethasone has shown improved progression-free and overall survival versus lenalidomide and dexamethasone in patients with relapsed or refractory multiple myeloma. We aimed to assess these regimens in patients with newly diagnosed multiple myeloma who are ineligible for haematopoietic stem-cell transplantation (HSCT). METHODS: ELOQUENT-1 is an open-label, multicentre, randomised, phase 3 trial conducted at 185 hospitals, oncology practices, and research centres in 19 countries. Eligible patients were aged 18 years or older with newly diagnosed, untreated, symptomatic myeloma and not candidates for high-dose therapy plus HSCT, and an Eastern Cooperative Oncology Group (ECOG) performance status of 2 or lower. Patients were randomly assigned (1:1) to receive elotuzumab plus lenalidomide and dexamethasone or lenalidomide and dexamethasone using an interactive voice response system, stratified by the International Staging System (ISS; stage I-II vs III), age (

Aim: To determine the safe dose range and pharmacokinetics of metronomic oral vinorelbine and obtain preliminary data on biomarkers and efficacy in patients with advanced cancer.Methods: Successive cohorts of patients received escalated doses of oral vinorelbine given thrice a week until disease progression, unacceptable toxicity (UT), or consent withdrawal. UT was any grade 4 toxicity, or grade 2 or 3 toxicity that would result to longer than 2-week break during the first 2 months of treatment. Blood samples were collected for pharmacokinetics and quantification of angiogenesis regulatory proteins.Results: Sixty-two patients (median age, 60 years) enrolled at six dose levels from 20 to 70 mg and received treatment for median 12.25 weeks (range, 2-216+). Unacceptable toxicity occurred in two of six patients treated at 60 mg (leucopenia grade 4 and epistaxis grade 2) and in one at 70 mg (leucopenia grade 2). The upper metronomic dose was 50 mg. Objective antitumor response documented in eight cases and 32% of patients experienced disease stability for minimum 6 months. Three responders (renal cancer, medullary thyroid carcinoma, and Kaposi sarcoma) received nonstop treatment for over 3 years without overt toxicity. Low pretreatment levels of circulating interleukin-8, vascular endothelial growth factor, and basic fibroblast growth factor were found predictors of efficacy. Steady-state concentrations of vinorelbine and its active metabolite ranged from 0.5 to 1.5 ng/mL.Conclusions: Metronomic administration of oral vinorelbine is feasible at doses up to 50 mg thrice a week and can yield sustainable antitumor activity without overt toxicity, probably through antiangiogenic mechanism. Further clinical investigation is warranted. (Clin Cancer Res 2009;15(20):6454–61)

Supplementary Data from Dose-Ranging Study of Metronomic Oral Vinorelbine in Patients with Advanced Refractory Cancer

Sunitinib is a multi-targeted tyrosine kinase inhibitor approved for the treatment of renal cell carcinoma and imatinib-resistant gastrointestinal stromal tumor and is currently being investigated against other forms of malignant tumors. Recently great interest has emerged for the application of sunitinib to glioblastoma treatment. In order to have a method with broad applicability it will be of importance to have access to a method that could be applied both in human plasma and cell uptake studies. No method has been reported thus far for the estimation of sunitinib uptake in glioma cells. We therefore set out to develop a method that could be applied for quantifying sunitinib in human plasma and in cell uptake studies. The method was validated and accredited according to ISO 17025:2005 guideline in human plasma and successfully applied to cancer patient plasma. Also, the method was effectively recruited to establish a protocol for the evaluation of sunitinib accumulation into M095K glioma cells. This method could significantly contribute to developmental phases in repurposing this drug in different cancer types.

Background Can learning outcomes be transformed in useful tools revealing strong and weak learning outcomes, learners, teachers; reporting student self-assessment overestimation; informing formative feedback and summative examinations? Methods Based on the ESMO / ASCO global curriculum, 66 level-two learning outcomes were identified and transformed in the iCAN!-Oncology and theyCAN!-Oncology questionnaires, anonymously completed online, before and after teaching, by trainees and trainers respectively, in a five-day fulltime undergraduate oncology course. Results In total, students assessed themselves (iCAN!) with 55% before and 70% after the course (27% improvement); teachers assessed students (theyCAN!) with 43% before and 69% after (60% improvement). Twenty level-two learning outcomes (30%) were scored below the pass / fail cut-point by students while 46 (70%) by teachers, before the course; none after the course. Students assessed themselves the highest in “TNM system” before (81%) and after (82%), while the teachers assessed students so in “Normal cell biology” before (72%) and “Moral / ethical issues in clinical research” after (83%). The lowest assessed outcome was the “Research protocol” by students (28%) and teachers (18%) before, and the “Anticancer agents” after (54% by both). Individual students self-assessed themselves from 31% to 88% before, and from 54% to 88% after; individual teachers assessed students from 29% to 66% before, and from 55% to 94% after. The iCAN! / theyCAN! provided detailed individual student or teacher profile, tightfisted or generous. Conclusions The iCAN! / theyCAN! differentiate strong and weak learning outcomes, learners, teachers; reveal no student self-assessment overestimation; inform formative feedback and summative exams at a metacognitive level; generalize to any course and assessor; support evidence-based teaching and learning SWOT policy.

Background/aim The aim of the study was to investigate whether E-cadherin and syndecan-1 are molecular markers of advanced non-small cell lung cancer (NSCLC). Patients and methods The expression of E-cadherin and syndecan-1 (SDC1) was examined immunohistochemically on tissue specimens of 64 patients, with stage III disease at presentation. The obtained expression data were correlated with clinical parameters. Results Negative expression of SDC1 was correlated with squamous histology (p=0.002). E-cadherin positive expression was significantly associated with increased 2-year overall survival (OS) rate (p=0.032). In the multivariate Cox analysis, performance status 0-1 was an independent predictor of OS (p=0.001) and disease-free survival (DFS) (p=0.001). E-cadherin expression was an independent predictor of OS (p=0.007) and DFS (p=0.029). Conclusion E-cadherin might be a prognostic factor for OS and DFS in advanced stage NSCLC patients. Further investigations are needed for the establishment of E-cadherin and syndecan-1 as molecular markers, affecting treatment response and survival.

Loss of a section of the long arm of chromosome 5, as a sole cytogenetic abnormality, characterizes a rare type of myelodysplastic syndrome [del(5q) MDS] and the co-existence of the JAK2 V617F mutation occurs in a small subset of these cases. Patients with isolated del(5q) MDS have a relatively favorable prognosis, with transformation to acute myeloid leukemia occurring in 5 years despite treatment discontinuation. The present case highlights the coexistence of the JAK2 V617F mutation in del(5q) MDS and suggests that lenalidomide treatment is beneficial and effective for these patients, leading to complete hematological, cytogenetic and molecular response. Hematological response may be sustained for long periods of time, even following the discontinuation of the treatment.

We evaluated progression-free survival (PFS) rate of patients treated with lenalidomide/dexamethasone (Len/Dex), the efficacy of the combination, and the prognostic significance of treatment at biochemical vs. clinical relapse on PFS in 207 consecutive myeloma patients treated with Len/Dex in second line, according to routine clinical practice in Greece. First-line treatment included bortezomib-based (63.3%) or immunomodulatory drug-based (34.8%) therapies; 25% of patients underwent autologous stem cell transplantation. Overall response rate was 73.4% (17.8% complete response and 23.7% very good partial response); median time to best response was 6.7 months. Overall, median PFS and 12-month PFS rate was 19.2 months and 67.6%, respectively. 67.5% of patients had biochemical relapse and 32.5% had clinical relapse prior to initiation of Len/Dex. Median PFS was 24 months for patients treated at biochemical relapse vs. 13.2 months for those treated at clinical relapse (HR:0.63, p = 0.006) and the difference remained significant after adjustment for other prognostic factors. Type of relapse was the strongest prognostic factor for PFS in multivariate analysis. These real-world data confirm the efficacy of Len/Dex combination at first relapse; more importantly, it is demonstrated for the first time outside a clinical trial setting that starting therapy with Len/Dex at biochemical, rather than at clinical relapse, is a significant prognostic factor for PFS, inducing a 37% reduction of the probability of disease progression or death.

We report 2 rare variants in the spectrum of human herpesvirus 8 (HHV8)/Epstein-Barr virus (EBV)-copositive lymphoproliferations arising in HIV-seronegative patients, including a large B-cell lymphoma arising in the setting of multicentric Castleman disease and a germinotropic lymphoproliferative disorder. In the first case, histology revealed features of multicentric Castleman disease and a proliferation of large lymphoid cells forming clusters or arcs or rings replacing the periphery of follicles or sheets of frank lymphoma outside the follicles. In the second case, a proliferation of large lymphoid cells totally or partially invaded follicle germinal centers. In both cases, the large cells were positive for EBV-encoded small RNA, HHV8 (LANA-1), MUM1/IRF4, and CD38 and negative for CD45, CD79a, CD10, BCL6, and CD138. In the large B-cell lymphoma, the large cells did not express detectable cytoplasmic immunoglobulin light- and heavy-chains, whereas in the germinotropic lymphoproliferative disorder, the large cells expressed μ heavy chain. The present cases broaden the spectrum of HHV-EBV--copositive lymphoproliferations.

Hydroxycarbamide (HC) is a ribonucleotide reductase inhibitor which promotes fetal hemoglobin (HbF) induction and has proven efficacy in sickle cell disease (SCD) patients. Given its mechanism of action and pr ior reports of genotoxicity in animal models, concern exists regarding long - term safety in relation to its cytotoxic effects. The purpose of this study was to retrospectively analyze the long - term (range 3 - 20 years, median 11) HC - derived clinical and biological effects, in 30 SCD patients (age range 20 - 68 years) from one referral center. HC treatment resulted in significant reduction of painful crises and transfusions, increase of HbF and hemoglobin as well as drop of white blood cell count and lactate dehydrogenase values. During the long term follow up time the following disea se complications were observed: pulmonary hypertension (2 patients), leg ulcers (1 patient) and renal impairment (1 patient). Seven patients discontinued HC therapy because of sc heduled pregnancy (3), severe neutropenia (2) and non - compliance (2). One poor HC compliant

Surgery still remains the cornerstone of breast cancer treatment but in some cases it has been implicated for stimulating early locoregional recurrence or even distant metastatic spread. There is evidence that angiogenesis, initiated by the surgical wounding itself during the process of healing, could offer valuable answers in such matters as well as enable a more profound understanding of the mechanisms of carcinogenesis in general. In our study, we investigated the impact of breast surgery on circulating angiogenesis-related factors in the peripheral blood. We used plasma samples from ten female patients with biopsy-proven breast cancer (Ca group) and six other females with fibroadenoma (Control group) before surgery (PRO) and post-operatively on days 3 (D3) and 7 (D7, only for breast cancer patients). These samples were analyzed for circulating levels of vascular endothelial growth factor A (VEGF-A), Interleukin-8 (IL-8) and fibroblast growth factor 2 (FGF2/bFGF) using ELISA. Moreover, 84 angiogenesis-associated mRNA transcripts were also assessed using quantitative RT-PCR arrays at the same time points. We were able to find a different angiogenic profile between breast cancer patients and those with benign breast disease (fibroadenoma). The effect of surgery on the mechanism of new blood vessel formation was found to be more robust and far more prolonged in women suffering from malignancy of the breast, than those operated for fibroadenoma. Mastectomy provoked a transient increase in VEGF-A plasma levels and altered the expression of many angiogenesis-related circulating gene transcripts.

Bcl-2 family proteins are important regulators of apoptosis and its antiapoptotic members, which are overexpressed in many types of cancer, are of high prognostic significance, establishing them as attractive therapeutic targets. Quercetin, a natural flavonoid, has drawn much attention because it exerts anticancer effects, while sparing normal cells. A multidisciplinary approach has been employed herein, in an effort to reveal its mode of action including dose-response antiproliferative activity and induced apoptosis effect, biochemical and physicochemical assays, and computational calculations. It may be concluded that, quercetin binds directly to the BH3 domain of Bcl-2 and Bcl-xL proteins, thereby inhibiting their activity and promoting cancer cell apoptosis.

Purpose IL-13, TNF-α and IL-1β have various effects on lung cancer growth and death, but the signaling pathways mediating these effects have not been extensively analyzed. Therefore, the effects of IL-13, TNF-α and IL-1β alone, and in combination with Fas, on cell viability and death as well as major signaling pathways involved in these effects were investigated in A549 lung carcinoma cells. Results Using MTT and flow cytometry, IL-13, TNF-α and IL-1β pretreatment decreased Fas-induced cell death. These anti-cell death effects were attenuated by pretreatment with inhibitors of Nuclear factor-κB [NF-κB], Phoshatidylinositole-3 kinase [PI3-K], JNK, p38 and ERK1/2 pathways. Using Western blot, IL-13, TNF-α and IL-1β treated cells showed time-dependent expression of p-ERK1/2, p-p38, p-JNK, p-Akt and p-IκBα proteins, decreased IκBα protein expression, no cleavage of Caspase-3 and PARP1 proteins and no notable alterations of Fas protein. IL-13 and TNF-α treated cells showed time-dependent increase of FLIP L expression. Conclusion IL-13, TNF-α and IL-1β attenuate the pro-cell death effects of Fas on A549 cells, at least partially, by pathways involving the NF-κB, PI3-K and MAP kinases, but not by alterations of Fas protein expression. The IL-13 and TNF-α cell survival effects may also be due to increased expression of FLIP L protein.

// Nikolaos Kapodistrias 1 , Konstantinos Mavridis 2 , Anna Batistatou 3 , Penelope Gogou 4 , Vasilios Karavasilis 5 , Ioannis Sainis 1 , Evangelos Briasoulis 1 , Andreas Scorilas 2 1 Cancer Biobank Center, University of Ioannina, University Campus, Ioannina, Greece 2 Department of Biochemistry and Molecular Biology, National and Kapodistrian University of Athens, Panepistimiopolis, Athens, Greece 3 Department of Pathology, School of Medicine, University of Ioannina, Greece 4 Clinical Oncology Department, Norwich University Hospital, UK 5 London Sarcoma Service, Department of Oncology, UCLH, London, UK Correspondence to: Andreas Scorilas, email: ascorilas@biol.uoa.gr Keywords: liposarcoma, miRNAs, miR-155, miRNA expression normalization, prognostic biomarker Received: July 05, 2016 Accepted: December 01, 2016 Published: December 28, 2016 ABSTRACT Liposarcoma (LPS) is a malignancy with extreme heterogeneity and thus optimization towards personalizing patient prognosis and treatment is essential. Here, we evaluated miR-155, miR-21, miR-143, miR-145 and miR-451 that are implicated in LPS, as novel FFPE tissue biomarkers. A total of 83 FFPE tissue specimens from primary LPS and lipomas (LPM) were analyzed. A proteinase K incubation-Trizol treatment coupled protocol was used for RNA isolation. After polyadenylation of total RNA and reverse transcription, expression analysis of 9 candidate reference and 5 target miRNAs was performed by qPCR. Genorm and NormFinder were used for finding the most suitable molecules for normalization. Survival analyses were performed in order to evaluate the prognostic potential of miRNAs. MiR-103 and miR-191 are most suitable for normalization of miRNA expression in LPS. MiR-155 and miR-21 are clearly overexpressed (P

Background There seems to be a significant arrhythmia burden in β-thalassemia major (TM) patients without overt cardiomyopathy. Apart from conventional electrocardiographic (ECG) and arrhythmic risk markers we studied novel markers of ventricular repolarization and autonomic imbalance both at rest and after exercise testing. Methods We studied 47 adult TM patients without systolic heart failure and 47 age and sex-matched healthy control subjects. The median age of the studied population was 36 [32–43] years, 57% men. Baseline demographic and clinical characteristics were recorded while 12-lead electrocardiograms, 24-hour ECG Holter recordings, and treadmill exercise stress tests were analyzed. Results TM patients exhibited increased QTc intervals in both 12-lead ECG recordings and in 24-hour Holter recordings. In addition, they had increased indexes of ventricular repolarization heterogeneity such as QT dispersion, and T peak-to-end/QT ratios. Furthermore, TM patients had decreased indexes of heart rate variability while the heart rate recovery after exercise was significantly attenuated compared to controls. Also, they had increased P wave and QRS duration while the QRS fragmentation was very prevalent. Finally, premature atrial extrasystoles and paroxysmal atrial fibrillation episodes were more frequent in TM patients. Conclusions TM patients with preserved left ventricular systolic function have several ECG abnormalities including alterations in ventricular depolarization and repolarization. Also, cardiac autonomic dysfunction is evident in 24-hour ECG monitoring as well as in the recovery phase after exercise testing. The prognostic value of specific arrhythmic risk indexes in this setting remains to be elucidated.

Prolyl hydroxylation is a post-translational modification that affects the structure, stability and function of proteins including collagen by catalysing hydroxylation of proline to hydroxyproline through action of collagen prolyl hydroxylases3 (C-P3H) and 4 (C-P4H). Three C-P3Hs (nomenclature was amended according to approval by the HGNC symbols and names at http://www.genenames.org/ and Entrez database at http://www.ncbi.nlm.nih.gov/gene ) leucineproline-enriched proteoglycan (leprecan) 1 (Lepre1), leprecan-like 1 (Leprel1), leprecan-like 2 (Leprel2) and two paralogs Cartilage-Related Protein (CRTAP) and leprecan-like 4 (Leprel4) are found in humans. The C-P4Hs are tetrameric proteins comprising a variable α subunit, encoded by the P4HA1, P4HA2 and P4HA3 genes and a constant β subunit encoded by P4HB. We used RT–PCR, qPCR, pyrosequencing, methylation-specific PCR, western blotting and immunohistochemistry to investigate expression and regulation of the C-P3H and C-P4H genes in B lymphomas and normal bone marrow. C-P3H and C-P4H are downregulated in lymphoma. Down-regulation is associated with methylation in the CpG islands and is detected in almost all common types of B-cell lymphoma, but the CpG islands are unmethylated or methylated at lower levels in DNA isolated from normal bone marrow and lymphoblastoid cell lines. Methylation of multiple C-P3H and C-P4H genes is present in some lymphomas, particularly Burkitt’s lymphoma. Methylation of C-P3H and C-P4H is common in B lymphomas and may have utility in differentiating disease subtypes.

The mammalian genome is transcribed in a developmentally regulated manner, generating RNA strands ranging from long to short non-coding RNA (ncRNAs). NcRNAs generated by intergenic sequences and protein-coding loci, represent up to 98 % of the human transcriptome. Non-coding transcripts comprise short ncRNAs such as microRNAs, piwi-interacting RNAs, small nucleolar RNAs and long intergenic RNAs, most of which exercise a strictly controlled negative regulation of expression of protein-coding genes. In humans, the DLK1-DIO3 genomic region, located on human chromosome 14 (14q32) contains the paternally expressed imprinted genes DLK1, RTL1, and DIO3 and the maternally expressed imprinted genes MEG3 (Gtl2), MEG8 (RIAN), and antisense RTL1 (asRTL1). This region hosts, in addition to two long intergenic RNAs, the MEG3 and MEG8, one of the largest microRNA clusters in the genome, with 53 miRNAs in the forward strand and one (mir-1247) in the reverse strand. Many of these miRNAs are differentially expressed in several pathologic processes and various cancers. A better understanding of the pathophysiologic importance of the DLK1-DIO3 domain-containing microRNA cluster may contribute to innovative therapeutic strategies in a range of diseases. Here we present an in-depth review of this vital genomic region, and examine the role the microRNAs of this region may play in controlling tissue homeostasis and in the pathogenesis of some human diseases, mostly cancer, when aberrantly expressed. The potential clinical implications of this data are also discussed.

Various cyanobacterial species have the capacity to produce different types of toxins. Microcystins, the most prominent cyanotoxins are considered health hazards because of their potential hepatotoxic effects. They are well known to contaminate freshwater ecosystems but their presence in marine ecosystems has been reported only occasionally. We investigated seasonal changes of microcystin concentrations both in water and in the edible species of mussels Mytilus galloprovincialis collected from Amvrakikos Gulf (salinity ranging from 30 parts per thousand to 34 parts per thousand), the biggest semi-enclosed basin in Greece. The microcystin concentrations in the water ranging from 0.003 to 19.8 ng l(-1), were below the World Health Organization (WHO) upper limit for recreational activities. In contrast, we found that microcystin concentrations in M. galloprovincialis mussels (ranging from 45 +/- 2 to 141.5 +/- 13.5 ng g(-1) ww) exceeded the upper limit of the tolerable daily intake (TDI) of microcystin as determined by WHO. Genotype composition of the total cyanobacterial community of the Gulf was analyzed by using denaturing gradient gel electrophoresis (DGGE) profiling of the rRNA internal transcribed spacer region (rRNA-ITS). The cyanobacterial community was found to be dominated almost exclusively by the cosmopolitan species Synechococcus - Synechocystis. In order to determine genes involved in the production of microcystins, a range of both specific and degenerate molecular primers against microcystin synthetase gene cluster (mcyS) was used. To our knowledge this is the first report of the presence of the hepatotoxic microcystins in the Mediterranean Sea, the first study on the accumulation of these toxins in mussels from a Mediterranean marine ecosystem and one of the few published works suggesting a potential association of microcystins with Synechococcus and/or Synechocystis cyanobacteria. The importance of our study is strengthened by the fact that Amvrakikos Gulf is among the most productive Greek "seafood" areas and a Mediterranean wetland of international significance according to Ramsar Convention. (C) 2011 Elsevier B.V. All rights reserved. Harmful Algae

Supplemental Digital Content is available in the text., Metronomic oral vinorelbine (VRL; Navelbine) was shown in clinical trials to yield sustainable antitumor activity possibly through antiangiogenic mechanisms. We investigated the effects of protracted low-dose VRL on human umbilical vein endothelial cells, compared with a conventional chemotherapy model. Human umbilical vein endothelial cell cultures were treated with different concentrations of VRL (0.001 nmol/l to 1 mmol/l) for 4, 24 and 96 h. The effects of different drug concentrations on cell growth, cell cycle, apoptosis and expression of the angiogenesis-modulating genes interleukin-8, cyclooxygenase-2, CD36 and peroxisome proliferator-activated receptor γ were assessed using the metronomic or conventional chemotherapy model. Apoptosis and cell-cycle effects were assessed by flow cytometry. Gene expression was measured at the transcript level by quantitative reverse transcriptase-PCR, protein expression by immunoblotting and levels of proteins secreted in the cell medium by enzyme-linked immunosorbent assay. Activation of the nuclear factor-κB pathway was investigated by immunoblot analysis of cytosolic and nuclear protein extracts. The half-maximal inhibitory concentrations (IC50) of VRL at 96 h were four orders lower compared with those after a 24-h exposure (1.23 nmol/l vs. 32 mmol/l for VRL). Drug concentrations at high nanomolar levels and above, which are relevant to conventional pulsatile dosing of VRL, induced a dose-dependent and nuclear factor-κB-related increase in proangiogenic interleukin-8 and cyclooxygenase-2 and a decrease in the thrombospondin-1 receptor CD36 and peroxisome proliferator-activated receptor γ at mRNA and protein levels. In contrast, the opposite was evident with protracted picomolar to low nanomolar concentrations (metronomic dosing). Our data provide experimental support for metronomic VRL by showing that a protracted low dose outperforms pulsed high-dose administration in inducing antiangiogenic effects in proliferating human endothelial cells.

Rheumatoid arthritis (RA) in patients suffering from hemoglobinopathies is an important clinical problem, but the correlation between these diseases is still imperfectly known. The aim of this study was to analyze the clinical, serological and radiological characteristics of RA occurring in patients with hemoglobinopathies (thalassemia major, thalassemia intermedia and sickle-cell disease). In a single institution, in an adult cohort of 90 patients with hemoglobinopathies, we investigated retrospectively medical records of the patients. We evaluated the clinical findings, the autoantibodies and the radiological progression of patients who were diagnosed with RA according the American College of Rheumatology (ACR) criteria for RA. There were found 4 patients, with thalassemia major, who fulfilling the ACR criteria for RA. The clinical picture of the patients revealed a mild form of arthritis of the knees, shoulders, wrist and hands, while one patient had episcleritis. All patients had radiological damage compatible with RA (Larsen's score, 28.75 ± 29). All had positive rheumatoid factor, while anti-cyclic citrullinated peptide antibodies were positive in 1 patient. Three patients received steroid treatment and one immunosuppressive agent (methotrexate). True RA with low frequency of extra-articular manifestations is described. The diagnosis of RA must be suspected in patients with hemoglobinopathies picture and chronic arthritis of small joints.

Valentinos Kounnis1, Elli Ioachim2, Martin Svoboda3, Andreas Tzakos4, Ioannis Sainis1, Theresia Thalhammer3, Georg Steiner5, Evangelos Briasoulis11Cancer Biobank Center of the University of Ioannina, Greece; 2Pathology Department of Hatzikosta General Hospital, Ioannina Greece; 3Department of Pathophysiology and Allergy Research, Medical University of Vienna, Austria; 4Department of Chemistry, University of Ioannina, Greece; 5TissueGnostics GmbH, Vienna, AustriaBackground: Organic anion-transporting polypeptides (OATPs) are influx transporters that mediate intracellular uptake of selective endogenous and xenobiotic compounds. Identification of new molecular targets and discovery of novel targeted therapies is top priority for pancreatic cancer, which lacks any effective therapy.Materials and methods: We studied expression of OATP 1A2, 1B1, and 1B3 in pancreatic cancer tissue and in cell lines. Formalin-fixed paraffin-embedded biopsy material of 12 human pancreatic cancers was immunohistochemically assessed for protein expression of the three studied influx transporters. Immunohistochemistry was evaluated by experienced pathologists and quantified by use of an automated image analysis system. BxPC-3 and MIA PaCa-2 pancreatic cancer cell lines were used to quantify transcripts of OATP 1B1 and 1B3.Results: OATP 1A2, 1B1, and 1B3 proteins were found ubiquitously expressed in all studied cases. Quantification performed by HistoQuest system revealed that mean intensity was 53 for 1A2, 45 for 1B1, and 167 for OATP 1B1/1B3 on a range scale 0–250 units. At mRNA level, 1B1 and 1B3 were overexpressed in both studied cancer cell lines but not in normal pancreatic tissue.Conclusion: OATPs 1A2, 1B1, and 1B3 are highly expressed in pancreatic adenocarcinoma. We suggest that expression of these transporters in pancreatic cancer justify research efforts towards discovery of novel therapeutics targeting OATPs.Keywords: organic anion-transporting polypeptides, targeted therapy, transporter

MicroRNAs (miRNAs) are evolutionarily conserved, naturally abundant, small, regulatory non-coding RNAs that inhibit gene expression at the post-transcriptional level in a sequence-specific manner. Each miRNA represses the protein expression of several coding genes in a manner proportional to the sequence complementarity with the target transcripts. MicroRNAs play key regulatory roles in organismal development and homeostasis. They control fundamental biological processes, such as stem-cell regulation and cellular metabolism, proliferation, differentiation, stress resistance, and apoptosis. Differential miRNA expression is found in malignant tumors in comparison to normal tissue counterparts. This indicates that miRNA deregulation contributes to the initiation and progression of cancer. Currently, miRNA expression signatures are being rigorously investigated in various tumor types, with the aim of developing novel, efficient biomarkers that can improve clinical management of cancer patients. This review discusses deregulated miRNAs in solid tumors, and focuses on their emerging prognostic potential.

Cyanobacterial cyclopeptides, including microcystins and nodularins, are considered a health hazard to humans due to the possible toxic effects of high consumption. From a pharmacological standpoint, microcystins are stable hydrophilic cyclic heptapeptides with a potential to cause cellular damage following uptake via organic anion-transporting polypeptides (OATP). Their intracellular biological effects involve inhibition of catalytic subunits of protein phosphatase 1 (PP1) and PP2, glutathione depletion and generation of reactive oxygen species (ROS). Interestingly, certain OATPs are prominently expressed in cancers as compared to normal tissues, qualifying MC as potential candidates for cancer drug development. In the era of targeted cancer therapy, cyanotoxins comprise a rich source of natural cytotoxic compounds with a potential to target cancers expressing specific uptake transporters. Moreover, their structure offers opportunities for combinatorial engineering to enhance the therapeutic index and resolve organ-specific toxicity issues. In this article, we revisit cyanobacterial cyclopeptides as potential novel targets for anticancer drugs by summarizing existing biomedical evidence, presenting structure-activity data and discussing developmental perspectives. Mar Drugs

Cyanobacterial blooms occur increasingly often and raise ecological concerns worldwide. In Mediterranean freshwater ecosystems algal blooms are commonly attributed to Microcystis, Anabaena, and Aphanizomenon genera while Planktothrix is the most common bloom forming cyanobacterium in deep Northern and prealpine European oligotrophic to mesotrophic lakes. In the framework of an undertaken study of cyanobacterial species in lakes of Northwestern Greece we investigated the cyanobacterial diversity in Lake Ziros throughout a 15-month period (January 2006-March 2007) by using molecular methods. Surprisingly, a severe cyanobacterial bloom occurred during the study period, which upon microscopic examination and detailed molecular characterization found to be caused by Planktothrix rubescens species. The appearance of P. rubescens from November 2006 coincided with poor cyanobacterial diversity and resulted in a thick epilimnetic bloom in March 2007 (3.1 x 10(8) cells/l and microcystin concentration 199 mu g/l). Genotype composition of the total cyanobacterial community of the lake was analyzed by using denaturing gradient gel electrophoresis (DGGE) profiling of the intergenic transcribed spacer region of the rnn operon (rRNA-ITS). A P. rubescens strain closely related to Kpr strain from Lake Klinckenberg, The Netherlands, was found to dominate. The importance of this observation is expanded by the fact that microcystin concentrations recorded in Lake Ziros were the highest measured ever in Greek aquatic ecosystems examined so far and also found amongst the highest recorded worldwide. (C) 2008 Published by Elsevier B.V. Harmful Algae

Objective: Combination of trastuzumab and anthracyclines in metastatic breast cancer (MBC) is precluded due to cardiotoxicity. Pegylated liposomal doxorubicin (PLD) is the least cardiotoxic among the anthracyclines. We performed a phase II study of trastuzumab and PLD with biomarker evaluation. Methods: Patients with MBC and HER2 overexpression, assessed as 3+ at local laboratories, received trastuzumab 8 mg/kg as loading dose followed by 6 mg/kg in combination with PLD 30 mg/m2, both given every 3 weeks. To be eligible, patients should have received first-line chemotherapy for MBC or should have relapsed within a year of adjuvant taxane. Tumor tissue blocks were collected for central review and exploratory biomarker evaluation. Left-ventricular ejection fraction (LVEF) was closely monitored by cardiac ultrasound. Results: Among 37 patients, an overall response rate of 22% was observed with a progression-free survival (PFS) of 6.5 months (0.8–31.1, 95% CI 2.7–10.3) and a survival of 18.7 months (1.6–40.8, 95% CI 3.7–33.7). No decline in LVEF was noticed. Overexpression of mTOR and TOP2A gene alterations were associated with better PFS. PTEN gene deletion was associated with resistance to treatment. Conclusion: Trastuzumab combined with PLD every 3 weeks is feasible, effective and safe in HER2-positive patients.

Introduction: The combination of Carboplatin and Paclitaxel is considered the standard of care as initial chemotherapy for Advanced Ovarian Cancer (AOC). We compared this regimen with the combination of Cisplatin, Paclitaxel and Doxorubicin. Patients and methods: Patients with AOC were randomised to either six courses of Paclitaxel 175 mg/m(2) plus Carboplatin 7AUC or Paclitaxel at the same dose plus Cisplatin 75 mg/m(2) plus Doxorubicin 40 mg/m(2). Results: Analysis was performed on 451 patients. The treatment groups were well balanced with regard to patient and disease characteristics. Performance status (PS) was better in the anthracycline arm. In terms of severe toxicity, the only significant difference between the two groups was the development of febrile neutropaenia in the anthracycline arm. Overall response rate was similar in both groups. With a median follow-up of 57.5 months, a marginal significance towards improved Progression-Free Survival (PFS) was noted in favour of the anthracycline arm, whilst there was no difference in overall survival. In multivariate analysis the hazard of disease progression at any time was significantly decreased by 25.5% for patients of the anthracycline arm. Conclusion: The combination of Cisplatin, Paclitaxel and Doxorubicin demonstrates a marginal PFS improvement, but no additional survival benefit when compared with the standard Carboplatin/Paclitaxel regimen. (c) 2008 Elsevier Ltd. All rights reserved. European Journal of Cancer

We report on a small-cell lung cancer case that was heralded by the presence of CENP-B specific anti-centromere autoantibodies (ACA) detected well before the diagnosis of cancer. The patient received chemotherapy plus radiotherapy, which resulted in complete remission of the tumor. Serum ACA levels were zeroed at the time of radiological documentation of tumor response and remained undetected at serial follow-up assessments until they rose again along with documentation of tumor recurrence which occurred 12 months later. We review in brief published research and discuss anti-centromere autoantibodies as potential biomarkers in small-cell lung cancer, a highly proliferative tumor which lacks sensitive serum markers.

Life-saving, risk-reducing medical interventions are required for women with a BRCA1/2 mutation. Interventions comprise a four-stage approach that involves risk assessment, genetic counseling, gene-mutation analysis and medical intervention strategies. Genetic counseling should be offered at specialized familial breast-cancer clinics and gene-mutation analysis should be recommended on the basis of personal and family-history-based risk criteria. Prophylactic bilateral salpingo-oophorectomy appears to offer the optimal benefit-risk ratio compared with prophylactic bilateral mastectomy, chemoprevention, or intensified surveillance. Tamoxifen is an alternative approach only for BRCA2 mutation carriers. The comprehensive, clinical decision-making Ioannina algorithm provided here can facilitate the complex preventive strategic approach. Newly diagnosed BRCA1/2 carriers might benefit from extensive surgery and a specific pharmacological treatment, but data to support this strategy are limited. Microarray gene-expression studies show that breast tumors from BRCA1 carriers are predominantly of basal subtype (i.e. triple negative) and BRCA2 carriers are of luminal subtype (i.e. estrogen-receptor-positive). Although optimum management of women with familial susceptibility to breast and ovarian cancer has not yet been prospectively validated, data indicate substantial benefits when an individualized evidence-based prevention strategy is provided by an experienced team.

Microcystin-LR (MC-LR) is a cyanobacterial cyclopeptide, known for its unique ability to cause acute liver injury. Its cellular uptake is facilitated by specific transmembrane organic anion-transporting polypeptides (OATPs) specifically OATP1B1 and 1B3. The objective of the present study was to investigate the expression of OATPs 1A2, 1B1 and 1B3 in pancreatic cancer cell lines BxPC-3 and MIA PACA-2 and assess their role in MC-LR-mediated cytotoxicity by using the novel xCELLigence system and flow cytometry. OATP1B1 and 1B3 were found to be expressed in both cell lines at both the mRNA and protein levels. The cytotoxic effects of MC-LR were proportionally related to the expression of these transporters. Moreover the cytotoxic potency of MC-LR was found superior to gemcitabine. Based on the expression of the organic anion transporting polypeptides 1B1 and 1B3 in pancreatic carcinoma tissue and cell lines and the potent cytotoxicity induced by MC-LR in vitro, we propose that this molecule could be held as structural basis for the development of novel targeted-compounds against pancreatic cancer.

Metronomic chemotherapy is the protracted, dense administration of low sub-toxic doses of chemotherapy, to inhibit tumor angiogenesis. Vinorelbine is an orally bioavailable vinca alkaloid shown to be useable for metronomic administration. In clinical trials, metronomic vinorelbine has been demonstrated to generate sustainable antitumor efficacy at low nanomolar (nM) concentrations with negligible toxicity. We sought to determine whether the clinically relevant metronomic concentration of vinorelbine is anti-angiogenic in vitro and whether hypoxia, often induced by anti-angiogenic therapy, modifies its effectiveness. We found that the metronomic concentration of 10 nM vinorelbine inhibited human umbilical vein endothelial cell (HUVEC) proliferation, migration, tube formation and sprouting. Severe hypoxia, did not affect the inhibitory effect of metronomic vinorelbine on migration, tube formation and sprouting. However, severe hypoxia reduced its anti-proliferative effect by decreasing its ability to induce G2/M block as it shifted the cell population to the G1 phase and decreased the fraction of the cells in the DNA synthesis S phase. Furthermore, the pro-apoptotic effects of 10 nM vinorelbine were also decreased. Metronomic vinorelbine decreased the Bcl-2/Bax ratio in normoxia whereas the ratio was reduced in severe hypoxia but unaltered by vinorelbine treatment. Akt signals to an anti-apoptotic pathway and we demonstrated that the Akt inhibitor V reversed the protective effect of severe hypoxia. Thus, we provide evidence for the anti-angiogenic basis of metronomic vinorelbine and we show that severe hypoxia mediates resistance to its anti-proliferative effect on endothelial cells. Akt warrants further investigation as a potential target to circumvent this hypoxic resistance.

MicroRNAs are small RNA molecules that negatively regulate the expression of the majority of proteins, mainly at the post-transcriptional level. Being stable in the circulation and resistant to storage handling, they are potentially promising biomarkers.We measured RNA levels of three microRNAs with tumorigenic or angiogenic potential (miR-155, miR-195, and miR-21) in blood samples taken from patients with early breast cancer, both preoperatively and postoperatively.We found that persistently elevated postoperative levels of miR-195 were detected only in patients who developed early tumor relapse and that miR-155 levels tended to increase three days postoperatively (p=0.05) and fell below baseline one month post-surgery (p0.05). We had no major findings for miR-21.The results of this pilot study indicate a possible involvement of miR-155 in surgery-induced angiogenesis and potential prognostic significance of high postoperative levels of circulating miR-195 in patients with breast cancer.

Purpose: The aim of this study was to explore the effect of dose-dense sequential chemotherapy with or without paclitaxel primarily on disease-free survival (DFS) and secondarily on overall survival (OS) in patients with high-risk operable breast cancer. Patients and methods: From June 1997 until November 2000, 604 patients with T1–3N1M0 or T3N0M0 tumors were randomized to three cycles of epirubicin 110 mg/m2 followed by three cycles of paclitaxel 250 mg/m2 followed by three cycles of ‘intensified’ CMF (cyclophosphamide 840 mg/m2, methotrexate 47 mg/m2 and fluorouracil 840 mg/m2) (group A), or to four cycles of epirubicin followed by four cycles of CMF, as in group A (group B). All cycles were given every 2 weeks with granulocyte colony-stimulating factor support. Results: A total of 595 patients were eligible. Median follow-up was 61.7 months for group A and 62 months for group B. The 3-year DFS was 80% in group A and 77% in group B. Survival rates were 93% and 90%, respectively. The effect of treatment on the hazard of death was different according to hormonal receptor status. More specifically, in patients with negative receptor status the hazard of death was significantly higher for group B (hazard ratio 2.42). Both regimens were well tolerated and severe acute side-effects were infrequent. No cases of severe cardiotoxicity or acute leukemia were recorded. Conclusions: The present study failed to demonstrate a significant difference in DFS or OS between the two treatment groups. However, our study has shown clearly that high-dose paclitaxel can be safely incorporated to dose-dense sequential chemotherapy.

To determine the more effective dosing sequence of intermittent erlotinib and docetaxel for treating chemotherapy-naive patients with advanced Non-Small Cell Lung Cancer (NSCLC).Patients were randomized to receive daily erlotinib for 12 consecutive days prior to docetaxel (Arm A) or after docetaxel (Arm B). Progression-free survival (PFS) was the primary end-point; secondary end-points were overall survival (OS) and objective response rate (ORR).Fifty eligible patients received a total of 226 treatment cycles (median: 3). Median PFS and OS were 3.6 months and 10.5 months, respectively (differences were not statistically significant between the two arms). Neutropenia grade 3 and 4 occurred in 15 patients, while two patients developed grade 3 diarrhea. There were two treatment-related deaths (pulmonary embolism and non-neutropenic sepsis).Intermittent administration of erlotinib does not appear to improve the clinical outcome of single-agent docetaxel chemotherapy in unselected patients with NSCLC in the first-line setting.

Pacritinib (previously known as SB-1518) is an innovative selective inhibitor of Janus kinase 2 and FMS-related tyrosine kinase 3 providing potential in the treatment of hematological malignancies such as myeloproliferative neoplasias, acute myeloid leukemia, and various lymphomas. Pacritinib has potent antiproliferative activity in Janus kinase 2 and/or FMS-related tyrosine kinase 3 activity-dependent cell lines and an ability to promote apoptosis and inhibit the signal transducers and activators of transcription (STAT) pathway. Pharmacokinetic studies have indicated a good per os bioavailability and favorable kinetic parameters. To date, promising results have been produced in five completed early-phase clinical trials in which pacritinib has been studied. Pacritinib displayed interesting activity and an acceptable safety profile, with mild to moderate gastrointestinal disorders being its most common adverse effects.

Cancer cells contain multiple genetic and epigenetic changes. The relative specificity of many epigenetic changes for neoplastic cells has allowed the identification of diagnostic, prognostic and predictive biomarkers for a number of solid tumors and hematological malignancies. Moreover, epigenetically-acting drugs are already in routine use for cancer and numerous additional agents are in clinical trials. Here, we review recent progress in the development and application of epigenetic strategies for the diagnosis, risk stratification and treatment of cancer.

BACKGROUND Cancer antigen 15-3 (CA 15-3), a circulating marker that determines secreted products of the polymorphic MUC1 gene, has been established as a convenient tool for monitoring breast carcinoma patients. METHODS The authors investigated alterations of soluble CA 15-3 in 57 postoperative breast carcinoma patients while they were receiving intensified adjuvant chemotherapy with granulocyte colony stimulating factor (G-CSF) support; 26 patients had American Joint Committee on Cancer (AJCC) Stage II, and 31 patients had AJCC Stage III breast carcinoma. Serial CA 15-3 values recorded thoughout the treatment were compared with baseline values, analyzed for correlation with hematologic and biochemical parameters, and compared with clinicopathologic characteristics and patient outcome. At a median follow-up time of 32 months, 47 of these patients remained relapse-free. RESULTS A twofold increase of CA 15-3 was detected at the end of the second week of treatment, remained significantly elevated in most patients at above the cutoff level of 30 U/mL throughout the treatment period (P < 0.0001), and subsided to pretreatment values 1–2 months after treatment cessation. CA 15-3 values were found to be associated strongly with absolute neutrophil count, serum lactate dehydrogenase, and alkaline phosphatase. The median values and the kinetics of tumor markers did not differ over time in regard to hormonal receptor status and disease recurrence. CONCLUSIONS These data provide strong evidence that G-CSF administration can induce elevation of CA 15-3 and indicate that false-positive results should be considered when evaluating CA 15-3 in patients who are receiving G-CSF. It is speculated that this phenomenon occurs through the induction of MUC1 antigen of unknown origin by G-CSF. Experimental investigation of this clinical observation is warranted. Cancer 2001;91:909–17. © 2001 American Cancer Society.

The encouraging results that have been reported for cisplatin combination chemotherapy in a minority of patients with cancer of unknown primary (CUP) together with the previously shown equal activity of carboplatin in this setting, prompted us to investigate the effectiveness of a carboplatin-containing regimen in a phase II clinical trial.Sixty-two evaluable CUP patients entered the protocol. The chemotherapy regimen consisted of carboplatin 300 mg/m2 IV D1, epirubicin 45 mg/m2 IV D1 and etoposide 120 mg/m2 IV D1–3 (CEE regimen) administered every 3 weeks. The median age of the patients was 61 years and 36 were male. Thirty-one diagnosed as poorly differentiated carcinomas (pdc) and the rest as adenocarcinomas. By clinicopathological criteria, 14 patients had a predominately nodal disease with pdc or poorly differentiated adenocarcinomas (pda), 3 women peritoneal carcinomatosis, and the remaining 46 patients had a predominately splanchnic involvement (24 pdc, 22 pda). Twenty-three patients responded to chemotherapy with 4 (6.5%) complete and 19 (30.5%) partial responders (RR 37%, 95% CI 25–49%). An equal activity of the regimen was observed between the two major histopathological types, the pdc and the adenocarcinomas. Nevertheless, significant differences were seen when the CEE regimen was assessed for its activity in the distinct clinicopathological subsets of CUP. Patients with predominately nodal disease of midline distribution with pdc or pda, and women with peritoneal carcinomatosis, achieved a response rate of 64 and 62% respectively, as compared with a 26% response rate for those with predominately splanchnic involvement. Overall median survival was 10 months and for patients with midline distribution 15 months. The regimen was well tolerated. It is concluded that CEE is a relatively nontoxic chemotherapy regimen and easily administered on an outpatient basis. This prospective phase II study confirmed the activity of carboplatin in the chemosensitive subsets of the predominately nodal disease of midline distribution and peritoneal carcinomatosis in women in the CUP syndrome.

About 3% of all cancer patients suffer from cancer of unknown primary origin. These patients present with metastatic disease for which a primary site cannot be detected at the time of diagnosis. Sophisticated diagnostic techniques and operational procedures have failed to improve the diagnostic efficacy in this group of patients. Consequently, a limited diagnostic procedure with basic laboratory tests and imaging studies is sufficient for the diagnosis of this syndrome. The use of immunohistochemistry, as well as serum tumor markers of high specificity that may help to identify other tumors, is highly suggested. Although the prognosis for the majority of these patients still remains poor, several subsets of favorable outcome to treatment have been recognized. Nevertheless, promising in vitro data and new drugs on trials, paralleled with a better knowledge of the underlying pathogenetic molecular mechanisms, offer a more optimistic look to the future therapeutic management of these patients.

Summary Background Paclitaxel has been demonstrated to have significant activity in recurrent or metastatic head and neck cancer (HNC). In addition, the combination of paclitaxel and cisplatin is active in untreated patients with inoperable HNC. Substitution of carboplatin for cisplatin allows the treatment to be delivered on an outpatient basis. Purpose of the study To evaluate the activity and toxicity of the combination of paclitaxel by three-hour infusion and carboplatin as first-line chemotherapy in patients with recurrent or metastatic HNC. Patients and Methods From March 1994 until August 1996, 49 patients with recurrent or metastatic HNC were treated with paclitaxel (200 mg/m2, by three-hour infusion) followed by carboplatin at an AUC of 7 mg.min/ml, every four weeks. G-CSF was administered prophylactically on days 2 to 12 of each cycle. There were 41 men and 8 women with a median age of 57 years (range 23–73). The majority of the patients were symptomatic and they had recurrent disease locoregionally. Fourteen patients had nasopharyngeal cancer (NPC) and 35 had squamous cell cancers of other areas of the head and neck region (non-NPC). Results At the completion of treatment, two patients with NPC demonstrated complete and six partial responses for an overall response rate of 57% (95% CI 29%–82%). Among patients with non-NPC, the response rate was 23% (95% CI 9%37%). After a median follow up period of 15 months, the median time to progression was 4.3 months in the non-NPC group and 16.5 months in the NPC group. At the time of the analysis, median survival had not been reached in NPC while it was 7.3 months in non-NPC patients. Grade 3–4 toxicities included anemia (2%) and leukopenia, thrombocytopenia, stomatitis, nausea/vomiting and diarrhea (4% each). Conclusions The combination of paclitaxel and carboplatin appears to be well tolerated but only moderately active in patients with advanced non-NPC of the head and neck region. However, its activity appears promising in NPC and deserves further investigation.