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// Shuya Yano 1, 2, 3 , Kiyoto Takehara 1, 2, 3 , Shinji Miwa 1, 2 , Hiroyuki Kishimoto 3 , Hiroshi Tazawa 4 , Yasuo Urata 5 , Shunsuke Kagawa 3 , Michael Bouvet 2 , Toshiyoshi Fujiwara 3 , Robert M. Hoffman 1, 2 1 AntiCancer, Inc., San Diego, CA, USA 2 Department of Surgery, University of California San Diego, CA, USA 3 Department of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan 4 Center for Innovative Clinical Medicine, Okayama University Hospital, Okayama, Japan 5 Oncolys BioPharm Inc., Tokyo, Japan Correspondence to: Robert M. Hoffman, email: all@anticancer.com Toshiyoshi Fujiwara, email: toshi_ f@md.okayama-u.ac.jp Keywords: fluorescence-guided surgery (FGS), telomerase dependent, adenovirus, GFP/RFP, survival Received: July 07, 2016 Accepted: September 06, 2016 Published: September 28, 2016 ABSTRACT We have previously developed a genetically-engineered GFP-expressing telomerase-dependent adenovirus, OBP-401, which can selectively illuminate cancer cells. In the present report, we demonstrate that targeting a triple-negative high-invasive human breast cancer, orthotopically-growing in nude mice, with OBP-401 enables curative fluorescence-guided surgery (FGS). OBP-401 enabled complete resection and prevented local recurrence and greatly inhibited lymph-node metastasis due to the ability of the virus to selectively label and subsequently kill cancer cells. In contrast, residual breast cancer cells become more aggressive after bright (white)-light surgery (BLS). OBP-401-based FGS also improved the overall survival compared with conventional BLS. Thus, metastasis from a highly-aggressive triple-negative breast cancer can be prevented by FGS in a clinically-relevant mouse model.

// Sun-Hyuk La 1, 2 , Seok-Jun Kim 1, 3 , Hyeok-Gu Kang 1, 3 , Han-Woong Lee 2 , Kyung-Hee Chun 1, 3 1 Department of Biochemistry and Molecular Biology, Yonsei University College of Medicine, Seodaemun-gu, Seoul 03722, Republic of Korea 2 Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seodaemun-gu, Seoul 03722, Republic of Korea 3 Brain Korea 21 PLUS Project for Medical Science, Yonsei University, Seodaemun-gu, Seoul 03722, Republic of Korea Correspondence to: Kyung-Hee Chun, email: khchun@yuhs.ac Keywords : telomerase, hTERT, galectin-3, cellular senescence, gastric cancer Received: May 23, 2016 Accepted: July 19, 2016 Published: August 1, 2016 ABSTRACT The human Telomerase Reverse Transcriptase (hTERT) gene encodes a rate-limiting catalytic subunit of telomerase that maintains genomic integrity. Suppression of hTERT expression could induce cellular senescence and is considered a potent approach for gastric cancer therapy. However, control of hTERT expression and function remains poorly understood in gastric cancer. In this study, we demonstrated that high expression levels of hTERT in malignant tissues are correlated with poor survival probability in gastric cancer patients. Knockdown of hTERT expression retarded cell proliferation and cellular senescence, which was confirmed by increased protein expression levels of p21 cip1 and p27 kip1 , and decreased phosphorylation of Rb. In contrast, overexpression of hTERT increased cell proliferation and decreased cellular senescence. Remarkably, the down-regulation of hTERT expression was detected in lgals3 -/- mouse embryo fibroblasts (MEFs). Knockdown of galectin-3 decreased the expression of hTERT in gastric cancer cells. Galectin-3 ablation-induced cellular senescence was rescued by concomitant overexpression of hTERT. hTERT ablation-induced cellular senescence and p21 cip1 and p27 kip1 expression was rescued by concomitant overexpression of galectin-3. The size of tumor burdens was increased in hTERT-overexpressed gastric cancer cells xenografted mice, whereas it was repressed by concomitant depletion of galectin-3. Additionally, we determined that the N-terminal domain of galectin-3 directly interacted with hTERT. The telomeric activity of hTERT was also decreased by galectin-3 ablation. Taken together, ablation of hTERT induces cellular senescence and inhibits the growth of gastric cancer cells, suggesting that it could be a potent target in gastric cancer therapy. We also propose that galectin-3 is an important regulator of hTERT expression and telomeric activity in gastric tumorigenesis.

// Yun-Jung Choi 1,* , Kiho Lee 2,3,* , Woo-Jin Park 1 , Deug-Nam Kwon 1 , Chankyu Park 1 , Jeong Tae Do 1 , Hyuk Song 1 , Seong-Keun Cho 5 , Kwang-Wook Park 6 , Alana N. Brown 3,4 , Melissa S. Samuel 3,4 , Clifton N. Murphy 3,4 , Randall S. Prather 3,4 and Jin-Hoi Kim 1 1 Animal Biotechnology to Stem Cell and Regenerative Biotechnology, Humanized Pig Research Center (SRC), Konkuk University, Seoul, Republic of Korea 2 Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, USA 3 Division of Animal Science, National Swine Resource and Research Center, University of Missouri, Columbia, MO, USA 4 National Swine Resource and Research Center, University of Missouri, Columbia, MO, USA 5 Department of Animal Science, Pusan National University, Miryang, Gyeongnam, Republic of Korea 6 Department of Animal Science and Technology, Sunchon National University, Suncheon, Jeonnam, Republic of Korea * These authors have contributed equally to this work Correspondence to: Randall S. Prather, email: // Jin-Hoi Kim, email: // Keywords : IL2RG, X-SCID, immunodeficiency, somatic cell nuclear transfer, TALEN, Pathology Section Received : May 08, 2016 Accepted : July 13, 2016 Published : July 24, 2016 Abstract In this study, we described the phenotype of monoallelic interleukin 2 receptor gamma knockout ( mIL2RG +/Δ69-368 KO) pigs. Approximately 80% of mIL2RG +/Δ69-368 KO pigs (8/10) were athymic, whereas 20% (2/10) presented a rudimentary thymus. The body weight of IL2RG +/Δ69-368 KO pigs developed normally. Immunological analysis showed that mIL2RG +/Δ69-368 KO pigs possessed CD25 + CD44- or CD25-CD44 + cells, whereas single (CD4 or CD8) or double (CD4/8) positive cells were lacking in mIL2RG +/Δ69-368 KO pigs. CD3 + cells in the thymus of mIL2RG +/Δ69-368 KO pigs contained mainly CD44 + cells and/or CD25 + cells, which included FOXP3 + cells. These observations demonstrated that T cells from mIL2RG +/Δ69-368 KO pigs were able to develop to the DN3 stage, but failed to transition toward the DN4 stage. Whole-transcriptome analysis of thymus and spleen, and subsequent pathway analysis revealed that a subset of genes differentially expressed following the loss of IL2RG might be responsible for both impaired T-cell receptor and cytokine-mediated signalling. However, comparative analysis of two mIL2RG +/Δ69-368 KO pigs revealed little variability in the down- and up-regulated gene sets. In conclusion, mIL2RG +/Δ69-368 KO pigs presented a T-B+NK- SCID phenotype, suggesting that pigs can be used as a valuable and suitable biomedical model for human SCID research.

// Moshe Reichenstein 1 , Gat Rauner 1, 2 , Shenhav Kfir 1, 2 , Tatiana Kisliouk 1 , Itamar Barash 1 1 Institute of Animal Science, ARO, The Volcani Center, Bet-Dagan, Israel 2 The Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot, Israel Correspondence to: Itamar Barash, email: itamar.barash@mail.huji.ac.il Keywords: STAT5, H2AX, mammary gland, methylation, paracrine Received: March 15, 2016 Accepted: May 20, 2016 Published: May 30, 2016 ABSTRACT Deregulated STAT5 activity in the mammary gland causes parity-dependent tumorigenesis. Epithelial cell cultures transfected with constitutively active STAT5 express higher levels of the histone H2AX than their non-transfected counterparts. Higher H2AX expression may be involved in tumorigenesis. Here, we aimed to link high STAT5 activity to H2AX–GFP expression by looking for distinct types of mammary cells that express these proteins. In vitro and in transgenic mice, only 0.2 and 0.02%, respectively, of the cells expressed the H2AX–GFP hybrid gene. Its expression correlated with that of the endogenous H2AX gene, suggesting that detectable H2AX–GFP expression marks high levels of H2AX transcript. Methylation of the H2AX promoter characterized non-GFP-expressing H2AX–GFP cells and was inversely correlated with promoter activity. Administration of 5-azacytidine increased H2AX promoter activity in an activated STAT5-dependent manner. In transgenic mice, H2AX–GFP expression peaked at pregnancy. The number of H2AX–GFP-expressing cells and GFP expression decreased in a Stat5a-null background and increased in mice expressing the hyperactivated STAT5. Importantly, H2AX–GFP activity was allocated to basal mammary cells lacking stem-cell properties, whereas STAT5 hyperactivity was detected in the adjacent luminal cells. Knockdown of RANKL by siRNA suggested its involvement in signaling between the two layers. These results suggest paracrine activation of H2AX via promoter demethylation in specific populations of basal mammary cells that is induced by a signal from neighboring luminal cells with hyper STAT5 activity. This pathway provides an alternative route for the luminally confined STAT5 to affect basal mammary cell activity.

Adamantinomatous craniopharyngiomas (ACPs) are clinically challenging tumours, the majority of which have activating mutations in CTNNB1. They are histologically complex, showing cystic and solid components, the latter comprised of different morphological cell types (e.g. β-catenin-accumulating cluster cells and palisading epithelium), surrounded by a florid glial reaction with immune cells. Here, we have carried out RNA sequencing on 18 ACP samples and integrated these data with an existing ACP transcriptomic dataset. No studies so far have examined the patterns of gene expression within the different cellular compartments of the tumour. To achieve this goal, we have combined laser capture microdissection with computational analyses to reveal groups of genes that are associated with either epithelial tumour cells (clusters and palisading epithelium), glial tissue or immune infiltrate. We use these human ACP molecular signatures and RNA-Seq data from two ACP mouse models to reveal that cell clusters are molecularly analogous to the enamel knot, a critical signalling centre controlling normal tooth morphogenesis. Supporting this finding, we show that human cluster cells express high levels of several members of the FGF, TGFB and BMP families of secreted factors, which signal to neighbouring cells as evidenced by immunostaining against the phosphorylated proteins pERK1/2, pSMAD3 and pSMAD1/5/9 in both human and mouse ACP. We reveal that inhibiting the MAPK/ERK pathway with trametinib, a clinically approved MEK inhibitor, results in reduced proliferation and increased apoptosis in explant cultures of human and mouse ACP. Finally, we analyse a prominent molecular signature in the glial reactive tissue to characterise the inflammatory microenvironment and uncover the activation of inflammasomes in human ACP. We validate these results by immunostaining against immune cell markers, cytokine ELISA and proteome analysis in both solid tumour and cystic fluid from ACP patients. Our data support a new molecular paradigm for understanding ACP tumorigenesis as an aberrant mimic of natural tooth development and opens new therapeutic opportunities by revealing the activation of the MAPK/ERK and inflammasome pathways in human ACP. Electronic supplementary material The online version of this article (10.1007/s00401-018-1830-2) contains supplementary material, which is available to authorized users.

Probes are essential to visualize proteins in their cellular environment, both using light microscopy as well as electron microscopy (EM). Correlated light microscopy and electron microscopy (CLEM) requires probes that can be imaged simultaneously by both optical and electron-dense signals. Existing combinatorial probes often have impaired efficiency, need ectopic expression as a fusion protein, or do not target endogenous proteins. Here, we present FLIPPER-bodies to label endogenous proteins for CLEM. Fluorescent Indicator and Peroxidase for Precipitation with EM Resolution (FLIPPER), the combination of a fluorescent protein and a peroxidase, is fused to a nanobody against a target of interest. The modular nature of these probes allows an easy exchange of components to change its target or color. A general FLIPPER-body targeting GFP highlights histone2B-GFP both in fluorescence and in EM. Similarly, endogenous EGF receptors and HER2 are visualized at nm-scale resolution in ultrastructural context. The small and flexible FLIPPER-body outperforms IgG-based immuno-labeling, likely by better reaching the epitopes. Given the modular domains and possibilities of nanobody generation for other targets, FLIPPER-bodies have high potential to become a universal tool to identify proteins in immuno-CLEM with increased sensitivity compared to current approaches. Electronic supplementary material The online version of this article (10.1007/s00418-018-1632-6) contains supplementary material, which is available to authorized users.

Repair of DNA double-strand breaks (DSBs) must be orchestrated properly within diverse chromatin domains in order to maintain genetic stability. Euchromatin and heterochromatin domains display major differences in histone modifications, biophysical properties, and spatiotemporal dynamics of DSB repair. However, it is unclear whether differential histone-modifying activities are required for DSB repair in these distinct domains. We showed previously that the Drosophila melanogaster KDM4A (dKDM4A) histone demethylase is required for heterochromatic DSB mobility. Here we used locus-specific DSB induction in Drosophila animal tissues and cultured cells to more deeply interrogate the impact of dKDM4A on chromatin changes, temporal progression, and pathway utilization during DSB repair. We found that dKDM4A promotes the demethylation of heterochromatin-associated histone marks at DSBs in heterochromatin but not euchromatin. Most importantly, we demonstrate that dKDM4A is required to complete DSB repair in a timely manner and regulate the relative utilization of homologous recombination (HR) and nonhomologous end-joining (NHEJ) repair pathways but exclusively for heterochromatic DSBs. We conclude that the temporal kinetics and pathway utilization during heterochromatic DSB repair depend on dKDM4A-dependent demethylation of heterochromatic histone marks. Thus, distinct pre-existing chromatin states require specialized epigenetic alterations to ensure proper DSB repair.

The advanced glycation end products (AGEs) participate in the pathogenesis of diabetic nephropathy (DN) by promoting renal inflammation and injury. L-carnosine acts as a quencher of the AGE precursors reactive carbonyl species (RCS), but is rapidly inactivated by carnosinase. In this study, we evaluated the effect of FL-926-16, a carnosinase-resistant and bioavailable carnosine derivative, on the onset and progression of DN in db/db mice.Adult male db/db mice and coeval db/m controls were left untreated or treated with FL-926-16 (30 mg·kgIn the prevention protocol, FL-926-16 significantly attenuated increases in creatinine (-80%), albuminuria (-77%), proteinuria (-75%), mean glomerular area (-34%), fractional (-40%) and mean (-42%) mesangial area in db/db mice. This protective effect was associated with a reduction in glomerular matrix protein expression and cell apoptosis, circulating and tissue oxidative and carbonyl stress, and renal inflammatory markers, including the NLRP3 inflammasome. In the regression protocol, the progression of DN was completely blocked, although not reversed, by FL-926-16. In cultured mesangial cells, FL-926-16 prevented NLRP3 expression induced by RCS but not by the AGE NFL-926-16 is effective at preventing the onset of DN and halting its progression in db/db mice by quenching RCS, thereby reducing the accumulation of their protein adducts and the consequent inflammatory response. In a future perspective, this novel compound may represent a promising AGE-reducing approach for DN therapy.

Glucocorticoids are widely used to treat inflammatory disorders; however, prolonged use of glucocorticoids results in side effects including osteoporosis, diabetes and obesity. Compound A (CpdA), identified as a selective NR3C1/glucocorticoid receptor (nuclear receptor subfamily 3, group C, member 1) modulator, exhibits an inflammation-suppressive effect, largely in the absence of detrimental side effects. To understand the mechanistic differences between the classic glucocorticoid dexamethasone (DEX) and CpdA, we looked for proteins oppositely regulated in bone marrow-derived macrophages using an unbiased proteomics approach. We found that the autophagy receptor SQSTM1 but not NR3C1 mediates the anti-inflammatory action of CpdA. CpdA drives SQSTM1 upregulation by recruiting the NFE2L2 transcription factor to its promoter. In contrast, the classic NR3C1 ligand dexamethasone recruits NR3C1 to the Sqstm1 promoter and other NFE2L2-controlled gene promoters, resulting in gene downregulation. Both DEX and CpdA induce autophagy, with marked different autophagy characteristics and morphology. Suppression of LPS-induced Il6 and Ccl2 genes by CpdA in macrophages is hampered upon Sqstm1 silencing, confirming that SQSTM1 is essential for the anti-inflammatory capacity of CpdA, at least in this cell type. Together, these results demonstrate how off-target mechanisms of selective NR3C1 ligands may contribute to a more efficient anti-inflammatory therapy.

Colorectal cancer is a global disease with increasing incidence. Mortality is largely attributed to metastatic spread and therefore, a mechanistic dissection of the signals which influence tumor progression is needed. Cancer stroma plays a critical role in tumor proliferation, invasion and chemoresistance. Here, we sought to identify and characterize exosomal microRNAs as mediators of stromal-tumor signaling. In vitro, we demonstrated that fibroblast exosomes are transferred to colorectal cancer cells, with a resultant increase in cellular microRNA levels, impacting proliferation and chemoresistance. To probe this further, exosomal microRNAs were profiled from paired patient-derived normal and cancer-associated fibroblasts, from an ongoing prospective biomarker study. An exosomal cancer-associated fibroblast signature consisting of microRNAs 329, 181a, 199b, 382, 215 and 21 was identified. Of these, miR-21 had highest abundance and was enriched in exosomes. Orthotopic xenografts established with miR-21-overexpressing fibroblasts and CRC cells led to increased liver metastases compared to those established with control fibroblasts. Our data provide a novel stromal exosome signature in colorectal cancer, which has potential for biomarker validation. Furthermore, we confirmed the importance of stromal miR-21 in colorectal cancer progression using an orthotopic model, and propose that exosomes are a vehicle for miR-21 transfer between stromal fibroblasts and cancer cells.

Tumor growth is not solely a consequence of autonomous tumor cell properties. Rather, tumor cells act upon and are acted upon by their microenvironment. It is tumor tissue biology that ultimately determines tumor growth. Thus, we developed a compound library screen for agents that could block essential tumor-promoting effects of the glioblastoma (GBM) perivascular stem cell niche (PVN). We modeled the PVN with three-dimensional primary cultures of human brain microvascular endothelial cells in Matrigel. We previously demonstrated stimulated growth of GBM cells in this PVN model and used this to assay PVN function. We screened the Microsource Spectrum Collection library for drugs that specifically blocked PVN function, without any direct effect on GBM cells themselves. Three candidate PVN-disrupting agents, Iridin, Tigogenin and Triacetylresveratrol (TAR), were identified and evaluated in secondary in vitro screens against a panel of primary GBM isolates as well as in two different in vivo intracranial models. Iridin and TAR significantly inhibited intracranial tumor growth and prolonged survival in these mouse models. Together these data identify Iridin and TAR as drugs with novel GBM tissue disrupting effects and validate the importance of preclinical screens designed to address tumor tissue function rather than the mechanisms of autonomous tumor cell growth.

miR-146a is a NF-κB induced microRNA that serves as a feedback regulator of this critical pathway. In mice, deficiency of miR-146a results in hematolymphoid cancer at advanced ages as a consequence of constitutive NF-κB activity. In this study, we queried whether the deficiency of miR-146a contributes to B-cell oncogenesis. Combining miR-146a deficiency with transgenic expression of c-Myc led to the development of highly aggressive B-cell malignancies. Mice transgenic for c-Myc and deficient for miR-146a were characterized by significantly shortened survival, increased lymph node involvement, differential involvement of the spleen and a mature B-cell phenotype. High-throughput sequencing of the tumors revealed significant dysregulation of approximately 250 genes. Amongst these, the transcription factor Egr1 was consistently upregulated in mice deficient for miR-146a. Interestingly, transcriptional targets of Egr1 were enriched in both the high-throughput dataset and in a larger set of miR-146a-deficient tumors. miR-146a overexpression led to downregulation of Egr1 and downstream targets with concomitant decrease in cell growth. Direct targeting of the human EGR1 by miR-146a was seen by luciferase assay. Together our findings illuminate a bona fide role for miR-146a in the modulation of B-cell oncogenesis and reveal the importance of understanding microRNA function in a cell- and disease-specific context.

Purpose There are indications that a history of allergy may offer some protection against cancer. We studied the relation of three objectively determined allergy markers with cancer mortality and hospitalization risk. Methods Associations between three allergy markers (number of peripheral blood eosinophil counts, skin test positivity, and serum total IgE) with mortality and hospitalization from any type and four common types of cancer (lung, colorectal, prostate, and breast cancer) were assessed in the Vlagtwedde–Vlaardingen cohort (1965–1990), with follow-up of mortality until 31 December 2008. Hospitalization data were available since 1 January 1995. Results There were no significant associations between objective allergy markers and cancer mortality or hospitalization. We found several associations in specific subgroups. A higher number of eosinophils was associated with a decreased risk of colorectal cancer mortality in ever smokers HR (95 % CI) = 0.61 (0.45–0.83) and in males 0.59 (0.42–0.83); however, no overall association was observed 0.84 (0.64–1.09). Skin test positivity was associated with a decreased risk of any cancer mortality only among females 0.59 (0.38–0.91) and showed no overall association 0.83 (0.67–1.04). Serum total IgE levels were associated with an increased risk of lung cancer mortality among females 4.64 (1.04–20.70), but with a decreased risk of cancer hospitalization in ever smokers 0.77 (0.61–0.97) and males 0.72 (0.55–0.93); however, no overall associations were observed [mortality 0.99 (0.79–1.25), and hospitalization 0.86 (0.71–1.04)]. Conclusions We found no associations between objective allergy markers and cancer in the total population. However, skin test positivity and a high number of eosinophils were associated with a reduced risk to die of cancer in specific subgroups. Hence, it seems important to study specific subgroups defined by gender and smoking habits in order to identify allergy markers of predictive value for cancer mortality. Electronic supplementary material The online version of this article (doi:10.1007/s10552-014-0489-9) contains supplementary material, which is available to authorized users.

The uptake of nanoparticles into cells often involves their engulfment by the plasma membrane and a fission of the latter. Understanding the physical mechanisms underlying these uptake processes may be achieved by the investigation of simple model systems that can be compared to theoretical models. Here, we present experiments on a massive uptake of silica nanoparticles by giant unilamellar lipid vesicles (GUVs). We find that this uptake process depends on the size of the particles as well as on the thermodynamic state of the lipid membrane. Our findings are discussed in the light of several theoretical models and indicate that these models have to be extended in order to capture the interaction between nanomaterials and biological membranes correctly.

BACKGROUND AND PURPOSE: Cardiovascular disease associated with antiretroviral therapy (ART) has become a major clinical challenge for HIV‐positive patients. However, the role of ART in blood vessel growth is largely unknown. Here, we examined an integral component of ART, nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) and investigated their effects on key microvascular functions, including angiogenesis and lymphangiogenesis. EXPERIMENTAL APPROACH: The angiogenesis/lymphangiogenesis capability of endothelial cells (ECs) was evaluated using migration, proliferation and tube formation assays in vitro, and mouse ear and Matrigel plug assays in vivo. Expressions of signalling molecules and mitochondrial antioxidant catalases were determined using Western blotting. Receptor tyrosine kinase (RTK) internalization and endocytosis were examined using flow cytometry and confocal immunofluorescence microscopy respectively. Mitochondrial DNA copy number and ROS were determined using quantitative real‐time PCR and MitoSOX staining respectively. KEY RESULTS: Pharmaceutical doses of NRTIs [azidothymidine (AZT), tenofovir disoproxil fumarate (TDF) and lamivudine (3TC)] inhibited angiogenesis and lymphangiogenesis both in vivo and in vitro by affecting the proliferation and migration of ECs. Correspondingly, NRTIs selectively attenuated the activation and transduction of endothelial RTK signals, VEGFR2 and FGFR1 pathways, in vascular ECs and the VEGFR3 pathway in lymphatic ECs. Both TDF and 3TC restrained RTKs’ endocytosis into early endosomes but not internalization, while AZT blocked the protein maturation of RTKs. Excessive ROS levels were detected in NRTI‐treated ECs, and the MnSOD mimic MnTMPyP alleviated the angiogenic/lymphangiogenic defects induced by NRTIs. CONCLUSIONS AND IMPLICATIONS: NRTIs negatively regulate angiogenesis and lymphangiogenesis by inducing mitochondrial oxidative stress and subsequently impairing RTK signalling in ECs. LINKED ARTICLES: This article is part of a themed section on Spotlight on Small Molecules in Cardiovascular Diseases. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.8/issuetoc

Novel agents have increased survival of multiple myeloma (MM) patients, however high-risk and relapsed/refractory patients remain challenging to treat and their outcome is poor. To identify novel therapies and aid treatment selection for MM, we assessed the ex vivo sensitivity of 50 MM patient samples to 308 approved and investigational drugs. With the results we i) classified patients based on their ex vivo drug response profile; ii) identified and matched potential drug candidates to recurrent cytogenetic alterations; and iii) correlated ex vivo drug sensitivity to patient outcome. Based on their drug sensitivity profiles, MM patients were stratified into four distinct subgroups with varied survival outcomes. Patients with progressive disease and poor survival clustered in a drug response group exhibiting high sensitivity to signal transduction inhibitors. Del(17p) positive samples were resistant to most drugs tested with the exception of histone deacetylase and BCL2 inhibitors. Samples positive for t(4; 14) were highly sensitive to immunomodulatory drugs, proteasome inhibitors and several targeted drugs. Three patients treated based on the ex vivo results showed good response to the selected treatments. Our results demonstrate that ex vivo drug testing may potentially be applied to optimize treatment selection and achieve therapeutic benefit for relapsed/refractory MM.

New neurons are born throughout the life of mammals in germinal zones of the brain known as neurogenic niches: the subventricular zone of the lateral ventricles and the subgranular zone of the dentate gyrus of the hippocampus. These niches contain a subpopulation of cells known as adult neural progenitor cells (aNPCs), which self-renew and give rise to new neurons and glia. aNPCs are regulated by many factors present in the niche, including the extracellular matrix (ECM). We show that the neuropeptide PACAP (pituitary adenylate cyclase-activating polypeptide) affects subventricular zone-derived aNPCs by increasing their surface adhesion. Gene array and reconstitution assays indicate that this effect can be attributed to the regulation of ECM components and ECM-modifying enzymes in aNPCs by PACAP. Our work suggests that PACAP regulates a bidirectional interaction between the aNPCs and their niche: PACAP modifies ECM production and remodeling, in turn the ECM regulates progenitor cell adherence. We speculate that PACAP may in this manner help restrict adult neural progenitors to the stem cell niche in vivo , with potential significance for aNPC function in physiological and pathological states.

// Jean-Francois Denis 1, * , K.E. Ludwig Scheckenbach 1, * , Anna Pfenniger 1, 2 , Merlijn J. Meens 1 , Rob Krams 3 , Lucile Miquerol 4 , Steven Taffet 5 , Marc Chanson 6 , Mario Delmar 7 and Brenda R. Kwak 1, 2 1 Department of Pathology and Immunology, University of Geneva, Geneva, Switzerland 2 Department of Medical Specializations - Cardiology, University of Geneva, Geneva, Switzerland 3 Department of Bioengineering, Imperial College, London, UK 4 Aix-Marseille University, CNRS UMR 7288, Developmental Biology Institute of Marseille, Marseille, France 5 Department of Microbiology, SUNY Upstate Medical University, Syracuse, NY, USA 6 Departments of Pediatrics and of Cell Physiology and Metabolism, Geneva University Hospitals and University of Geneva, Geneva, Switzerland 7 The Leon H. Charney Division of Cardiology, New York University School of Medicine, New York, NY, USA * These authors have contributed equally to this work Correspondence to: Brenda R. Kwak, email: Brenda.KwakChanson@unige.ch Keywords: atherosclerosis, endothelium, Cx40, shear stress, IκBα Received: November 08, 2016 Accepted: February 27, 2017 Published: March 22, 2017 ABSTRACT Connexins are proteins forming gap junction channels for intercellular communication. Connexin40 (Cx40) is highly expressed by endothelial cells (ECs) of healthy arteries but this expression is lost in ECs overlying atherosclerotic plaques. Low/oscillatory shear stress observed in bends and bifurcations of arteries is atherogenic partly through activation of the pro-inflammatory NFκB pathway in ECs. In this study, we investigated the relation between shear stress, Cx40 and NFκB. Shear stress-modifying casts were placed around carotid arteries of mice expressing eGFP under the Cx40 promoter ( Cx40 +/eGFP ). We found that Cx40 expression is decreased in carotid regions of oscillatory shear stress but conserved in high and low laminar shear stress regions. These results were confirmed in vitro . Using phage display, we retrieved a binding motif for the intracellular regulatory Cx40 C-terminus (Cx40CT), i.e. HS[I, L, V][K, R]. One of the retrieved peptides (HSLRPEWRMPGP) showed a 58.3% homology with amino acids 5-to-16 of IκBα, a member of the protein complex inhibiting NFκB activation. Binding of IκBα (peptide) and Cx40 was confirmed by crosslinking and en face proximity ligation assay on carotid arteries. TNFα-induced nuclear translocation of NFκB in ECs was enhanced after reducing Cx40 with siRNA. Transfection of HeLa cells with either full-length Cx40 or Cx40CT demonstrated that Cx40CT was sufficient for inhibition of TNFα-induced NFκB phosphorylation. Finally, Tie2Cre Tg Cx40 fl/fl Apoe -/- mice showed exaggerated shear stress-induced atherosclerosis and enhanced NFκB nuclear translocation. Our data show a novel functional IκBα-Cx40 interaction that may be relevant for the control of NFκB activation by shear stress in atherogenesis.

Mice with the hypomorphic AIF-Harlequin mutation exhibit a highly heterogeneous mitochondriopathy that mostly affects respiratory chain complex I, causing a cerebral pathology that resembles that found in patients with AIF loss-of-function mutations. Here we describe that the antidiabetic drug pioglitazone (PIO) can improve the phenotype of a mouse Harlequin (Hq) subgroup, presumably due to an inhibition of glycolysis that causes an increase in blood glucose levels. This glycolysis-inhibitory PIO effect was observed in cultured astrocytes from Hq mice, as well as in human skin fibroblasts from patients with AIF mutation. Glycolysis inhibition by PIO resulted from direct competitive inhibition of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Moreover, GAPDH protein levels were reduced in the cerebellum and in the muscle from Hq mice that exhibited an improved phenotype upon PIO treatment. Altogether, our results suggest that excessive glycolysis participates to the pathogenesis of mitochondriopathies and that pharmacological inhibition of glycolysis may have beneficial effects in this condition., Highlights • Respiratory chain-defective Harlequin mice have low glycaemia but their phenotype yet variously improved with pioglitazone. • The phenotype improvement observed with pioglitazone in a mouse subgroup correlates with an increased glycaemia. • Pioglitazone inhibits glyceraldehyde-3-phosphate dehydrogenase reducing exaggerated glycolysis in the Harlequin mice. Human mitochondriopathies constitute a group of metabolic diseases with spectacularly heterogeneous clinical presentation and without efficient therapy so far. A similar variability of the disease presentation hallmarks the Harlequin mice suffering a mitochondriopathy. We analyzed mouse inter-individual response to drugs. Adopting such a “personalized medicine” approach, we identified a subgroup of responders to the anti-diabetic drug, pioglitazone. Paradoxically, the improvement of phenotype correlated with an increased glycaemia of the initially hypoglycemic Harlequin mouse. Studying this subgroup allowed us to identify the glyceraldehyde-3-phosphate dehydrogenase enzyme as an additional target of pioglitazone.

The phytoestrogen resveratrol has been reported to possess cancer chemo-preventive activity on the basis of its effects on tumor cell lines and xenograft or carcinogen-inducible in vivo models. Here we investigated the effects of resveratrol on spontaneous mammary carcinogenesis using Delta 16HER2 mice as HER2+/ER alpha+ breast cancer model. Instead of inhibiting tumor growth, resveratrol treatment (0.0001% in drinking water; daily intake of 4 mu g/mouse) shortened tumor latency and enhanced tumor multiplicity in Delta 16HER2 mice. This in vivo tumor-promoting effect of resveratrol was associated with up-regulation of Delta 16HER2 and down-regulation of ER alpha protein levels and was recapitulated in vitro by murine (CAM6) and human (BT474) tumor cell lines. Our results demonstrate that resveratrol, acting as a proteasome inhibitor, leads to Delta 16HER2 accumulation which favors the formation of Delta 16HER2/HER3 heterodimers. The consequential activation of downstream mTORC1/p70S6K/4EBP1 pathway triggers cancer growth and proliferation. This study provides evidence that resveratrol mechanism of action (and hence its effects) depends on the intrinsic molecular properties of the cancer model under investigation, exerting a tumor-promoting effect in luminal B breast cancer subtype models.

// Maria Roel 1 , Juan A. Rubiolo 1 , Jorge Guerra-Varela 2 , Siguara B. L. Silva 3, 4 , Olivier P. Thomas 3, 5 , Pablo Cabezas-Sainz 2 , Laura Sanchez 2 , Rafael Lopez 6 , Luis M. Botana 1 1 Department of Pharmacology, Universidade de Santiago de Compostela, Campus Lugo, 27002 Lugo, Spain 2 Department of Genetics, Universidade de Santiago de Compostela, Campus Lugo, 27002 Lugo, Spain 3 Geoazur, UMR Universite Nice Sophia Antipolis-CNRS-IRD-OCA, 06560 Valbonne, France 4 Laboratoire de Pharmacognosie, UMR CNRS 8076 BioCIS, LabEx LERMIT, Universite Paris-Sud, Faculte de Pharmacie, 92290 Châtenay-Malabry, France 5 School of Chemistry, Marine Biodiscovery, National University of Ireland Galway, SW4 Galway, Ireland 6 Translational Medical Oncology, Health Research Institute of Santiago (IDIS), Complexo Hospitalario Universitario de Santiago de Compostela (SERGAS), 15706 Santiago de Compostela, Spain Correspondence to: Luis M. Botana, email: luis.botana@usc.es Keywords: crambescidins, cell cycle inhibition, apoptosis, zebrafish xenograft model, cancer treatment Received: March 13, 2016 Accepted: September 27, 2016 Published: November 04, 2016 ABSTRACT The marine environment constitutes an extraordinary resource for the discovery of new therapeutic agents. In the present manuscript we studied the effect of 3 different sponge derived guanidine alkaloids, crambescidine-816, -830, and -800. We show that these compounds strongly inhibit tumor cell proliferation by down-regulating cyclin-dependent kinases 2/6 and cyclins D/A expression while up-regulating the cell cyclin-dependent kinase inhibitors -2A, -2D and -1A. We also show that these guanidine compounds disrupt tumor cell adhesion and cytoskeletal integrity promoting the activation of the intrinsic apoptotic signaling, resulting in loss of mitochondrial membrane potential and concomitant caspase-3 cleavage and activation. The crambescidin 816 anti-tumor effect was fnally assayed in a zebrafish xenotransplantation model confirming its potent antitumor activity against colorectal carcinoma in vivo . Considering these results crambescidins could represent promising natural anticancer agents and therapeutic tools.

Ageing of human skin is associated with phenotypic changes in the cutaneous cells; the major functional markers of ageing occur as consequences of dermal and epidermal cell senescence and of structural and compositional remodeling of normally long-lived dermal extracellular matrix proteins. Understanding the contribution of the dermal cells in skin ageing is a key question, since this tissue is particularly important for skin integrity and its properties can affect the epidermis. Several microRNAs have been shown to be involved in the regulation of pathways involved in cellular senescence and exerted important effects on tissues ageing. In this study, we demonstrate that the expression of miR-152 and miR-181a increased during the human dermal fibroblasts senescence and that their overexpression, is sufficient to induce cellular senescence in early-passage cells. The increase of these miRNAs during cells senescence was accompanied by a decrease in integrin α5 and collagen XVI expression at mRNA and/or protein levels resulting in reduced cellular adhesion and suggesting extracellular matrix remodeling. These findings indicate that changes in miRNAs expression, by modulating the levels of adhesion proteins and extra-cellular matrix components, such as integrin α5 and collagen XVI, could contribute to the compositional remodelling of the dermis and epidermis occurring during skin aging.

// Clementine Druelle 1,6,* , Claire Drullion 1,* , Julie Desle 1,* , Nathalie Martin 1 , Laure Saas 1 , Johanna Cormenier 1 , Nicolas Malaquin 1,7 , Ludovic Huot 2 , Christian Slomianny 3 , Fatima Bouali 1 , Chantal Vercamer 1 , David Hot 2 , Albin Pourtier 1 , Eric Chevet 4,5 , Corinne Abbadie 1 and Olivier Pluquet 1 1 Universite de Lille, Institut Pasteur de Lille, CNRS UMR8161, Mechanisms of Tumourigenesis and Targeted Therapies, Lille, France 2 Universite de Lille, Institut Pasteur de Lille, INSERM, CNRS UMR8204, Centre d’Infection et d’Immunite de Lille, Lille, France 3 Universite de Lille, INSERM U1003, Villeneuve d’Ascq, France 4 Oncogenesis, Stress, Signaling, INSERM ER-440 Universite de Rennes, Rennes, France 5 Centre de Lutte Contre le Cancer Eugene Marquis, Rennes, France 6 Department of Anatomy and Neuroscience, Biosciences Institute, University College Cork, Cork, Ireland 7 Institut du Cancer de Montreal, Montreal, QC, Canada * These authors have contributed equally to this work Correspondence to: Olivier Pluquet, email: // Keywords : senescence, unfolded protein response, ATF6α, endoplasmic reticulum, normal human dermal fibroblast, Gerotarget Received : June 22, 2016 Accepted : August 02, 2016 Published : August 22, 2016 Abstract Cellular senescence is known as an anti-tumor barrier and is characterized by a number of determinants including cell cycle arrest, senescence associated β-galactosidase activity and secretion of pro-inflammatory mediators. Senescent cells are also subjected to enlargement, cytoskeleton-mediated shape changes and organelle alterations. However, the underlying molecular mechanisms responsible for these last changes remain still uncharacterized. Herein, we have identified the Unfolded Protein Response (UPR) as a player controlling some morphological aspects of the senescent phenotype. We show that senescent fibroblasts exhibit ER expansion and mild UPR activation, but conserve an ER stress adaptive capacity similar to that of exponentially growing cells. By genetically invalidating the three UPR sensors in senescent fibroblasts, we demonstrated that ATF6α signaling dictates senescence-associated cell shape modifications. We also show that ER expansion and increased secretion of the pro-inflammatory mediator IL6 were partly reversed by silencing ATF6α in senescent cells. Moreover, ATF6α drives the increase of senescence associated-β-galactosidase activity. Collectively, these findings unveil a novel and central role for ATF6α in the establishment of morphological features of senescence in normal human primary fibroblasts.

JAK1 frameshift mutations may promote cancer cell immune evasion by impeding upregulation of the antigen presentation pathway in microsatellite unstable endometrial cancers (ECs). This study investigated the JAK1 mutation frequency, its functional implication in immune evasion and its prognostic significance in microsatellite unstable EC. Microsatellite instability and three microsatellite repeats within JAK1 were analyzed in 181 ECs. Sixty-two (34%) ECs showed microsatellite instability, of which 22 (35%) had a JAK1 mutation. LMP7, TAP1 and HLA class I protein expression and the presence of CD8-positive T-cells were analyzed in the microsatellite unstable ECs. JAK1 mutant microsatellite unstable ECs showed impaired upregulation of LMP7 (P=0.074) and HLA class I (P

// Massimiliano Agostini 1,2 , Margherita Annicchiarico-Petruzzelli 3 , Gerry Melino 1,2 and Alessandro Rufini 4 1 Medical Research Council, Toxicology Unit, Leicester University, Leicester, UK 2 Department of Experimental Medicine and Surgery, University of Rome “Tor Vergata”, Rome, Italy 3 Biochemistry Laboratory IDI-IRCC, Department of Experimental Medicine and Surgery, University of Rome “Tor Vergata”, Rome, Italy 4 Department of Cancer Studies, CRUK Leicester Cancer Centre, University of Leicester, Leicester, UK Correspondence to: Alessandro Rufini, email: // Keywords : p73, p53 family, oxidative stress, metabolism, ROS Received : February 22, 2016 Accepted : April 16, 2016 Published : April 22, 2016 Abstract Reactive oxygen species are involved in both physiological and pathological processes including neurodegeneration and cancer. Therefore, cells have developed scavenging mechanisms to maintain redox homeostasis under control. Tumor suppressor genes play a critical role in the regulation of antioxidant genes. Here, we investigated whether the tumor suppressor gene TAp73 is involved in the regulation of metabolic adaptations triggered in response to oxidative stress. H 2 O 2 treatment resulted in numerous biochemical changes in both control and TAp73 knockout (TAp73-/-) mouse embryonic fibroblasts, however the extent of these changes was more pronounced in TAp73-/- cells when compared to control cells. In particular, loss of TAp73 led to alterations in glucose, nucleotide and amino acid metabolism. In addition, H 2 O 2 treatment resulted in increased pentose phosphate pathway (PPP) activity in null mouse embryonic fibroblasts. Overall, our results suggest that in the absence of TAp73, H 2 O 2 treatment results in an enhanced oxidative environment, and at the same time in an increased pro-anabolic phenotype. In conclusion, the metabolic profile observed reinforces the role of TAp73 as tumor suppressor and indicates that TAp73 exerts this function, at least partially, by regulation of cellular metabolism.

Motivation: Understanding the structure and interplay of cellular signalling pathways is one of the great challenges in molecular biology. Boolean Networks can infer signalling networks from observations of protein activation. In situations where it is difficult to assess protein activation directly, Nested Effect Models are an alternative. They derive the network structure indirectly from downstream effects of pathway perturbations. To date, Nested Effect Models cannot resolve signalling details like the formation of signalling complexes or the activation of proteins by multiple alternative input signals. Here we introduce Boolean Nested Effect Models (B-NEM). B-NEMs combine the use of downstream effects with the higher resolution of signalling pathway structures in Boolean Networks. Results: We show that B-NEMs accurately reconstruct signal flows in simulated data. Using B-NEM we then resolve BCR signalling via PI3K and TAK1 kinases in BL2 lymphoma cell lines. Availability and implementation: R code is available at https://github.com/MartinFXP/B-NEM (github). The BCR signalling dataset is available at the GEO database (http://www.ncbi.nlm.nih.gov/geo/) through accession number GSE68761. Contact: martin-franz-xaver.pirkl@ukr.de, Rainer.Spang@ukr.de Supplementary information: Supplementary data are available at Bioinformatics online.

In this study we address the mechanical properties of Sb2S3 nanowires and determine their Young’s modulus using in situ electric-field-induced mechanical resonance and static bending tests on individual Sb2S3 nanowires with cross-sectional areas ranging from 1.1·104 nm2 to 7.8·104 nm2. Mutually orthogonal resonances are observed and their origin explained by asymmetric cross section of nanowires. The results obtained from the two methods are consistent and show that nanowires exhibit Young’s moduli comparable to the value for macroscopic material. An increasing trend of measured values of Young’s modulus is observed for smaller thickness samples.

Robust, long-lasting immune responses are elicited by memory T cells that possess properties of stem cells, enabling them to persist long-term and to permanently replenish the effector pools. Thus, stem cell-like memory T (TSCM) cells are of key therapeutic value and efforts are underway to characterize TSCM cells and to identify means for their targeted induction. Here, we show that inhibition of mechanistic/mammalian Target of Rapamycin (mTOR) complex 1 (mTORC1) by rapamycin or the Wnt-β-catenin signalling activator TWS119 in activated human naive T cells leads to the induction of TSCM cells. We show that these compounds switch T cell metabolism to fatty acid oxidation as favoured metabolic programme for TSCM cell generation. Of note, pharmacologically induced TSCM cells possess superior functional features as a long-term repopulation capacity after adoptive transfer. Furthermore, we provide insights into the transcriptome of TSCM cells. Our data identify a mechanism of pharmacological mTORC1 inhibitors, allowing us to confer stemness to human naive T cells which may be significantly relevant for the design of innovative T cell-based cancer immunotherapies., Highlights • Immunostimulatory effect of rapamycin on induction of TSCM cells • Previously unknown mTORC1 inhibiting drug effect of the Wnt activator TWS119 • Insights into the metabolic regulation of human CD4 + TSCM cells and into their transcriptome Stem cell-like memory T (TSCM) cells represent the newly identified memory T cell subset, which harbours key therapeutic value by its self-renewal capacity and ability to generate memory and effector T cells. However, the signalling pathways controlling TSCM cell formation remain incompletely understood. Here, we present the induction of CD4 + TSCM cells from highly purified naïve cells by pharmacological inhibition of mTORC1 by either rapamycin or TWS119. We carried out comprehensive transcriptome and metabolic analyses of induced and naturally occurring TSCM cells. Targeted induction of TSCM cells by pharmacological means is highly relevant for the design of novel immunotherapeutic approaches.

In recent years, tumor Adoptive Cell Therapy (ACT), using administration of ex vivo-enhanced T cells from the cancer patient, has become a promising therapeutic strategy. However, efficient homing of the anti-tumoral T cells to the tumor or metastatic site still remains a substantial hurdle. Yet the tumor site itself attracts both tumor-promoting and anti-tumoral immune cell populations through the secretion of chemokines. We attempted to identify these chemokines in a model of spontaneous metastasis, in order to "hijack" their function by expressing matching chemokine receptors on the cytotoxic T cells used in ACT, thus allowing us to enhance the recruitment of these therapeutic cells. Here we show that this enabled the modified T cells to preferentially home into spontaneous lymph node metastases in the TRAMP model, as well as in an inducible tumor model, E.G7-OVA. Due to the improved homing, the modified CD8+ T cells displayed an enhanced in vivo protective effect, as seen by a significant delay in E.G7-OVA tumor growth. These results offer a proof of principle for the tailored application of chemokine receptor modification as a means of improving T cell homing to the target tumor, thus enhancing ACT efficacy. Surprisingly, we also uncover that the formation of the peri-tumoral fibrotic capsule, which has been shown to impede T cell access to tumor, is partially dependent on host T cell presence. This finding, which would be impossible to observe in immunodeficient model studies, highlights possible conflicting roles that T cells may play in a therapeutic context.

// Andres F. Cardona 1, 2, 3, * , Leonardo Rojas 4, 5, * , Beatriz Wills 2, * , Oscar Arrieta 6, * , Hernan Carranza 1, 2, 3 , Carlos Vargas 1, 2, 3 , Jorge Otero 1, 2, 3 , Luis Corrales-Rodriguez 7 , Claudio Martin 8 , Noemi Reguart 9 , Pilar Archila 2 , July Rodriguez 2 , Mauricio Cuello 10 , Carlos Ortiz 1 , Sandra Franco 1 , Christian Rolfo 11 , Rafael Rosell 12 , on behalf of the CLICaP # 1 Clinical and Traslational Oncology Group, Institute of Oncology, Clinica del Country, Bogota, Colombia 2 Foundation for Clinical and Applied Cancer Research (FICMAC), Bogota, Colombia 3 Clinical and Traslational Research Department, Faculty of Medicine, Universidad el Bosque, Bogota, Colombia 4 Clinical Oncology Department, Centro Javeriano de Oncologia, Hospital Universitario San Ignacio, Bogota, Colombia 5 Faculty of Medicine, Pontificia Universidad Javeriana, Bogota, Colombia 6 Thoracic Oncology Unit, Instituto Nacional de Cancerologia (INCan), Mexico City, Mexico 7 Medical Oncology Department, Hospital San Juan de Dios, San Jose, Costa Rica 8 Thoracic Oncology Unit, Alexander Fleming Institute, Buenos Aires, Argentina 9 Medical Oncology Department, Hospital Clinic, Barcelona, Spain 10 Clinical Oncology Department, Hospital de Clinicas-UdeLAR, Montevideo, Uruguay 11 Early Clinical Trials Unit, Oncology Department, Antwerp University Hospital and Center for Oncological Research (CORE), Antwerp University, Edegem, Belgium 12 Medical Oncology Department, Catalan Institute of Oncology, Hospital Germans Trias i Pujol, Badalona, Barcelona, Spain # Latin American Consortium for Lung Cancer Investigation * These authors have contributed equally to this study Correspondence to: Andres F. Cardona, email: andres.cardona@clinicadelcountry.com , a_cardonaz@yahoo.com Keywords: non-small cell lung cancer, BIM deletion, EGFR mutation, survival Received: February 24, 2016 Accepted: August 24, 2016 Published: September 19, 2016 ABSTRACT Background: Germline alterations in the proapoptotic protein Bcl-2-like 11 (BIM) can have a crucial role in diverse tumors. To determine the clinical utility of detecting BIM deletion polymorphisms (par4226 bp/ par363 bp) in EGFR positive non-small-cell lung cancer (NSCLC) we examined the outcomes of patients with and without BIM alterations. Results: BIM deletion was present in 14 patients (15.7%). There were no significant differences between patients with and without BIM -del in clinical characteristics or EGFR mutation type; however, those with BIM -del had a worse overall response rate (ORR) to erlotinib (42.9% vs. 73.3% in patients without BIM -del ; p=0.024) as well as a significantly shorter progression-free survival (PFS) (10.8 BIM -del+ vs. 21.7 months for patients without BIM -del ; p=0.029) and overall survival (OS) (15.5 BIM -del+ vs. 34.0 months for patients without BIM -del ; p=0.035). Multivariate Cox regression analysis showed that BIM -del+ was an independent indicator of shorter PFS (HR 3.0; 95%CI 1.2-7.6; p=0.01) and OS (HR 3.4; 95%CI 1.4-8.3; p=0.006). Methods: We studied 89 NSCLC Hispanic patients with EGFR mutation who were treated with erlotinib between January 2009 and November 2014. BIM deletion polymorphisms (BIM -del ) was analyzed by PCR in formalin-fixed paraffin-embedded (FFPE) tissues of tumor biopsies. We retrospectively analyzed clinical characteristics, response rate, toxicity, and outcomes among patients with and without BIM- del . Conclusions: The incidence of BIM -del found in Hispanic patients is similar to that previously described in Asia. This alteration is associated with a poor clinical response to erlotinib and represents an independent prognostic factor for patients who had NSCLC with an EGFR mutation.

The expansion of myeloid derived suppressor cells (MDSCs), a suppressive population able to hamper the immune response against cancer, correlates with tumor progression and overall survival in several cancer types. We have previously shown that MDSCs can be induced in vitro from precursors present in the bone marrow and observed that these cells are able to actively proliferate in the presence of activated T cells, whose activation level is critical to drive the suppressive activity of MDSCs. Here we investigated at molecular level the mechanisms involved in the interplay between MDSCs and activated T cells. We found that activated T cells secrete IL-10 following interaction with MDSCs which, in turn, activates STAT3 phosphorylation on MDSCs then leading to B7-H1 expression. We also demonstrated that B7-H1+ MDSCs are responsible for immune suppression through a mechanism involving ARG-1 and IDO expression. Finally, we show that the expression of ligands B7-H1 and MHC class II both on in vitro-induced MDSCs and on MDSCs in the tumor microenvironment of cancer patients is paralleled by an increased expression of their respective receptors PD-1 and LAG-3 on T cells, two inhibitory molecules associated with T cell dysfunction. These findings highlight key molecules and interactions responsible for the extensive cross-talk between MDSCs and activated T cells that are at the basis of immune suppression.

In breast cancer, constitutive activation of NF-κB has been reported, however, the impact of genetic variation of the pathway on patient prognosis has been little studied. Furthermore, a combination of genetic variants, rather than single polymorphisms, may affect disease prognosis. Here, in an extensive dataset (n = 30,431) from the Breast Cancer Association Consortium, we investigated the association of 917 SNPs in 75 genes in the NF-κB pathway with breast cancer prognosis. We explored SNP-SNP interactions on survival using the likelihood-ratio test comparing multivariate Cox’ egression models of SNP pairs without and with an interaction term. We found two interacting pairs associating with prognosis: patients simultaneously homozygous for the rare alleles of rs5996080 and rs7973914 had worse survival (HRinteraction 6.98, 95% CI=3.3-14.4, P=1.42E-07), and patients carrying at least one rare allele for rs17243893 and rs57890595 had better survival (HRinteraction 0.51, 95% CI=0.3-0.6, P = 2.19E-05). Based on in silico functional analyses and literature, we speculate that the rs5996080 and rs7973914 loci may affect the BAFFR and TNFR1/TNFR3 receptors and breast cancer survival, possibly by disturbing both the canonical and non-canonical NF-κB pathways or their dynamics, whereas, rs17243893-rs57890595 interaction on survival may be mediated through TRAF2-TRAIL-R4 interplay. These results warrant further validation and functional analyses. ispartof: Oncotarget vol:6 issue:35 pages:37979-94 ispartof: location:United States status: published