bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A47: Inhibition of Rho kinase signaling in angiosarcomas leads to decreased tumor growth and invasiveness, but enhanced resistance to standard chemotherapeutics
Authors: Mitchell Dianne Courtenay , Bryan Brad Allen
Published: March 2013

The Rho-associated coiled-coil protein kinases (ROCK 1 & 2) are serine/threonine kinases serving as key modulators of processes involving cytoskeletal rearrangement, focal adhesion formation, cell motility, and tumor cell invasion. Deregulation of ROCK signaling has been reported across diverse tumors types and overexpression of ROCK proteins in metastatic tumors directly contributes to their aggressive behavior through stimulating cell migration and epithelial-to-mesenchymal transition. Indeed, a number of published reports from our lab and others indicate that inhibiting ROCK signaling effectively disrupts solid tumor formation, progression, and metastasis. Interestingly, over the past decade more than 20 publications from different groups have reported diverse and often conflicting roles for the ROCK proteins in cell survival in differing tumor types, bringing into question what the overall response would be of a cancer patient who is taking standard chemotherapy combined with ROCK inhibitors. Despite the widespread interest in targeting ROCK signaling for cancer therapy, the molecular mechanisms explaining these conflicting pleiotropic roles for ROCK signaling in cell fate decisions is almost exclusively unknown. In this study, we examined the effects of ROCK inhibition combined with standard chemotherapy regimens on angiosarcoma tumor progression. Pharmacological inhibition of ROCK's kinase activity or shRNA knockdown of ROCK1 or 2 gene expression resulted in decreased angiosarcoma tumor formation and invasion, however led to significantly increased tumor cell survival following cisplatin or busulfan treatment. To determine the molecular mechanisms controlling the pro-survival response of inhibiting ROCK signaling following cytotoxic injury, we performed whole genome microarray analysis revealing that pharmacological inhibition or gene expression knockdown of ROCK proteins led to a statistically significant ablation in the expression of a number of cisplatin and busulfan responsive genes known to be downstream targets of p53 induction, DNA damage response and repair, and cell cycle control. This study provides one of the most detailed explanations at the molecular level as to how ROCK signaling modulates cell survival by regulating gene expression at a global level—a paradox that must be solved before inhibition of ROCK activity can be clinically used as an effective anticancer therapeutic, lest we develop an excellent antimetastatic drug that unfortunately (for some tumor types) enhances tumor resistance to chemotherapies.
Abstract A54: Potential mechanisms of resistance to targeted agents in human clear cell renal cell carcinoma
Authors: Grozav Adrian G. , Vaziri Susan A.J. , Eric Koegle R. , Grabowski Dale R. , Bukowski Ronald M. , Ganapathi Mahrukh K. , Ganapathi Ram N.
Published: March 2013

Targeted therapy with multiple receptor tyrosine kinase inhibitors (RTKI) has led to a substantial improvement in the standard of care for patients with advanced or metastatic clear cell renal cell carcinoma (CCRCC). We have previously reported that using a phosphoproteomics approach, Ras and Rab interactor 1 (RIN1) was identified as a potential target modulating the antiproliferative effects of sorafenib and sunitinib in a panel of human CCRCC (Proc. AACR 51: Abst. #1626, 2010). While both 10 μM sorafenib and 7.5 μM sunitinib were effective in down-regulating RIN1 protein levels in CCRCC following treatment for 48h, during subsequent recovery for 48h in drug free medium, recovery of RIN1 protein levels was observed in sorafenib but not sunitinib treated cells. mRNA expression profiling revealed that: (a) VEGF-A and HIF-1α levels were unaffected during 48h treatment with sorafenib or sunitinib or subsequent recovery; and (b) while HIF-2α levels were increased >2-fold following 48h exposure to sorafenib or sunitinib, during subsequent recovery in drug-free medium, the increased levels of HIF-2α persisted in cells treated with sorafenib but not sunitinib. Compared to CCRCC cells that were growth inhibited in vitro at concentrations of 5–10 μM sorafenib or sunitinib, in 27 patient CCRCC samples evaluated, the median mRNA levels of RIN1 was 7.2-fold lower in 22 samples (Range 1.6–22.3) and 2.2-fold higher in 5 samples (Range 1.2–27.1). Stable over-expression and down-regulation of RIN1 in CCRCC led to resistance or sensitivity respectively, to the anti-proliferative effects of sorafenib and sunitinib but not the mTOR inhibitor temsirolimus. In contrast, CCRCC cells adapted to proliferate in 4 μM everolimus exhibited a >3-fold increase in IC50, and cross-resistance to temsirolimus but not sorafenib or sunitinib. Evaluation in vitro of other targeted agents, demonstrated dose-dependent anti-proliferative effects with the PI3K/mTOR dual inhibitor BEZ235 (>80% growth inhibition at 10nM) but not the pan-PI3K inhibitor BKM120 (<30% growth inhibition at 10–500 nM) with CCRCC cells. The observed anti-proliferative effects of BEZ235 or BKM120 were comparable in CCRCC cells harboring the wild type or mutant von Hippel-Lindau gene. While BEZ235 (1–5 nM) or everolimus (100 nM) alone led to <40% growth inhibition, the combination significantly enhanced anti-proliferative effects (>85% growth inhibition) in CCRCC cells expressing resistance to everolimus. These results suggest the PI3K/mTOR dual inhibitor BEZ235 may possibly improve the treatment efficacy of mTOR inhibitors for CCRCC. The findings suggest RTKI and mTOR inhibitors have distinct mechanisms of resistance, and combinations may be useful to improve disease control of metastatic CCRCC.
Abstract A64: NFκB-IκBα Interaction: A Mechanism of Resistance to Cisplatin in Non Small Cell Lung Cancer (NSCLC)
Authors: Heavey Susan , Barr Martin , Godwin Peter , Edwards Connla , O'Byrne Ken , Gately Kathy
Published: March 2013

Introduction: The purpose of this study was to investigate the role of the PI3K-NFκB pathway in resistance to cisplatin in Non-small cell lung cancer (NSCLC). This form of malignancy is the leading cause of cancer morbidity and mortality in the western world with a poor overall 5 year survival of <15%. The most effective systemic chemotherapy for NSCLC is cisplatin-based combination treatment. However, chemoresistance is a major therapeutic problem and understanding the mechanisms involved is critical to the development of new therapeutic intervention strategies. The PI3K/AKT/mTOR pathway signals through NFκB and plays an important role in NSCLC, representing a novel therapeutic target for overcoming cisplatin resistance.

Methods: A panel of cisplatin resistant NSCLC cell lines (H460, SKMES1, A549) were developed in our laboratory. H460 parent & resistant cell lines were screened for changes in mRNA expression using the PI3K Profiler array (SABiosciences). Changes in gene expression were validated by QPCR and protein expression by Western blot and HCA. IκBα exons 3–5 were screened for mutations by sequencing. DHMEQ is an inhibitor of NFκB translocation. The effects of DHMEQ on NFκB translocation as well as on cell apoptosis and proliferation were assessed by HCA and BrdU assays.

Results: An 11.99 fold increase in IκBα mRNA expression was identified in the cisplatin resistant H460 cells compared to parent cells. QPCR, Western blot and HCA confirmed this overexpression of IκBα. No mutations were identified in exons 3-5 of the IκBα gene. Inhibition of NFκB by DHMEQ led to significantly increased apoptosis and significantly decreased proliferation in cisplatin resistant cells versus parent cells.

Conclusions: The transcription factor NFκB is involved in cell survival and regulates the transcription of its own inhibitor, IκBα. Both IκBα and NFκB were found to be upregulated in cisplatin resistant cells compared to cisplatin sensitive cells. Inhibiton of NFκB by the inhibitor DHMEQ succeeded in reducing resistant cell survival and proliferation. NFκB inhibition could represent a novel therapeutic strategy to overcome NSCLC resistance to cisplatin.
Abstract B21: Resistance to B-RAF inhibitor is mediated by reactivation of MAPK pathway by FGFR3/Ras signaling in B-RAF V600E mutant melanoma
Authors: Yadav Vipin , Zhang Xiaoyi , Liu Jiangang , Estrem Shawn , Li Shuyu Dan , Gong Xue-Qian , Buchanan Sean , Henry James R. , Starling James J. , Peng Sheng-Bin
Published: March 2013

Melanoma is the most aggressive form of skin cancer. The RAF/MEK/ERK MAP kinase signaling plays a key role in melanoma progression and etiology, and is considered an important target for anti-melanoma therapies. Approximately 50% of melanomas carry an activating B-RAF V600E mutation that results in constitutive activation of B-RAF and downstream MAPK signaling. Recently, vemurafenib (PLX4032), a selective B-RAF inhibitor, was recently approved by U.S. FDA for treatment of metastatic and unresectable melanomas that carry an activating B-RAF V600E mutation. However, patients eventually developed resistance to vemurafenib and relapsed within an average of 7 months. To improve the clinical benefit of B-RAF inhibitors, it is critical to identify the molecular mechanisms that confer resistance to B-RAF inhibition. Recent reports indicate that multiple mechanisms may render mutant B-RAF expressing melanoma cells resistant to B-RAF inhibition. In this study, we generated vemurafenib-resistant cell lines by chronic treatment of the human B-RAF V600E melanoma cell lines, and investigated the underlying mechanism(s) of resistance. Our analysis revealed that MAPK pathway reactivation through Ras is the key resistance mechanism in these cells. Furthermore, microarray-based gene expression profiling confirmed a significant elevation of RTK and Ras gene signatures in the vemurafenib-resistant cells. Mechanistically, we found that the enhanced activation of FGFR3 is linked to elevated Ras and MAPK activation in the resistant cells, therefore conferring resistance to vemurafenib. Constitutive or inducible activation of FGFR3 sufficiently reactivated Ras/MAPK signaling and conferred resistance to vemurafenib in the parental B-RAF V600E melanoma cells. Additionally, genetic or pharmacological inhibition of the FGFR3/Ras axis restored the sensitivity of the resistant cells to vemurafenib. Finally, we demonstrated that vemurafenib-resistant cells are sensitive to MEK or pan-RAF inhibition and maintain their addiction to the MAPK pathway. Therefore, inhibition of MEK or pan-RAF activities is an effective therapeutic strategy to overcome acquired-resistance to vemurafenib. Together, we describe a novel FGFR3/Ras mediated mechanism for acquired-resistance to B-RAF inhibition. Our results shed new light on the complexity of the resistance mechanisms and have implications for the development of new therapeutic strategies to improve the outcome of patients with B-RAF V600E melanoma.
Abstract B39: Mechanisms of multidrug resistance in AMLs and selective targeting via small molecules
Authors: Xing Chengguo , Hermanson David , Das Sonia
Published: March 2013

Multidrug resistance (MDR) is a major hurdle in the treatment of cancer and there is a pressing need for new therapies. We have recently developed a small molecule, CXL017, with selective cytotoxicity towards MDR cancer cells. In this study, we offer new mechanistic insights about multidrug resistance in acute myeloid leukemias and present evidence for the selective therapeutic potential of CXL017 in the treatment of MDR AMLs.

CXL017 significantly and selectively suppresses the growth of tumors derived from the MDR AML cells. In addition, drug resistant AML cells failed to develop stable resistance to CXL017 upon prolonged exposure. Remarkably, instead of acquiring cross-resistance, drug resistant AML cells exposed to CXL017 were re-sensitized to standard therapies (10 to 100-fold). Through gene profiling, protein characterization, and genetic up/down-regulation, we have identified several key signaling processes both at the transcription, such as Topo IIβ, and post-transcription levels, such as Mcl-1, that contribute to the multidrug resistance. Some of these changes also contribute to the selective sensitivity of multidrug resistant AMLs to CXL017. These results overall demonstrate the possibility of developing small-molecule therapies selective against multidrug resistance in cancer therapy upon gaining the mechanistic insights of drug resistance.
Abstract B52: Design and synthesis of GADD45β agonists for sorefenib-resistance hepatocellular carcinoma cells
Authors: Ou Da-Liang , Fan Hsiang-Hsuan , Hsu Chiun , Cheng Ann-Lii , Hsu Cheng-Yi , Shiau Chung-Wai
Published: March 2013

Background: Growth arrest DNA damage-inducible gene 45β (GADD45β) have been reported in regulating cellular stress response, survival, senescence, and apoptosis in cancer cells. Our previous data showed that GADD45β was induced in sorafenib-treated hepatocellular carcinoma (HCC) cell line and xenograft models. Sorafenib induction was not induced after the treatment ERK kinase inhibitor (U1206) and Raf inhibitor (ZM336372), suggesting that Raf/MEK/ERK kinase signaling pathway did not involve in induction of GADD45β by sorafenib.

Methods: We have designed and synthesized a series of SC-compounds. These compounds were treated with and analyzed in terms of GADD45β expression in HCC cell lines (Huh-7, Huh7R). Also, Effects of SC-compounds and sorafenib induced cell apoptosis were evaluated.

Results: In seeking new GADD45β agonists, we have screened a series of compounds with GADD45β expression assay. SC-20, a small molecule with N-(4-(quinolin-4-yloxy)phenyl)benzamide skeleton, has shown significantly in induction of GADD45β expression. Interestingly, SC-20 has no inhibition in phosphorylation of ERK, indicating that GADD45β induction by SC-20 is independent of ERK signaling pathway. In addition, SC-20 is able to inhibit HCC cell growth with MTT assay and further induce cell apoptosis. Moreover, SC-20 is able to overcome sorafenib resistance cell line by activating GADD45β expression.

Conclusions: SC-20 provides a new concept that increasing GADD45β along will be applicable in sorafenib-sensitive and resistant HCC therapy.

Grant support: NSC-100-2325-B-002-042, DOH100-TD-B-111-001, NSC-100-2325-B-010-007
Abstract B9: Activated MAPK Pathway Mediates Resistance to Romidepsin via Bim Degradation in Romidepsin-Selected HuT 78 Cells
Authors: Chakraborty Arup , Robey Robert , Gillet Jean-Pierre , Luchenko Victoria , Piekarz Richard , Gottesman Michael , Collie Nathan , Bates Susan
Published: March 2013

Acquired and intrinsic resistance to histone deacetylase inhibitors (HDIs), a new targeted group of anti-tumor agents, limits their clinical efficacy. A detailed understanding of the mechanisms of resistance to HDIs may lead to strategies designed to increase clinical efficacy. To investigate molecular mechanisms of resistance to the HDI romidepsin (Dp), we studied a cutaneous T-cell lymphoma (CTCL) cell line, HuT 78, independently selected in verapamil (Vp) or valspodar (PSC833) to prevent the emergence of P-glycoprotein (Pgp), a known resistance mechanism. The HuT 78 sublines DpVp 50 and DpP 75 display 100-200-fold resistance to romidepsin, not due to Pgp expression. A custom-made Taqman low density gene expression array detected increased expression of insulin receptor (IR) in the resistant cells. Real-time PCR analysis confirmed the results of gene array and detected more than 50-fold upregulation of IR in the romidepsin-selected cells compared to the parental cells. Increased phosphorylation (5- to 8- fold) of mitogen activated protein kinase kinase (MEK), a downstream effector of the IR pathway, was also observed in the resistant cells compared to the parental cells. While HuT 78 cells were insensitive to MEK inhibition, resistant cells were found to be exquisitely sensitive to MEK inhibition (IC50 < 10 nM) but not to phosphatidylinositol 3-kinase (PI3K) inhibition. Combined treatment of romidepsin with low concentrations (1- 3 nM) of MEK inhibitor also resulted in increased cell death in romidepsin-resistant HuT 78 cells. The exquisite sensitivity to MEK inhibition in the resistant sublines was found to correlate with restoration of the expression of Bim (BCL2L11), a Bcl-2-homology domain-3 only (BH-3) proapoptotic protein. In the resistant cells, the MAPK pathway appeared to regulate Bim expression posttranslationally as we did not detect any gene induction of BCL2L11 or FOXO3, a transcription factor known to activate BCL2L11 expression, following MEK inhibition. These findings implicate increased activation of the MAPK pathway as a mechanism of resistance to romidepsin, and suggest that combining romidepsin with MEK inhibitors may be an effective strategy to overcome resistance.
Abstract IA11: Using Molecular Profiling Strategies in Clinical Trials to Understand Resistance Mechanisms in the Era of Personalized Cancer Medicine
Authors: Siu Lillian L.
Published: March 2013

Resistance to molecularly targeted agents remains a key challenge in cancer therapeutics. Even in the setting of oncogenically addicted tumor types matched with potent and specific kinase inhibitors, the duration of drug response is limited. For example, in the phase III trial of vemurafenib versus dacarbazine in patients with advanced malignant melanoma harboring BRAF V600E mutations, 48% of patients in the vermurafenib arm achieved an objective response with a progression-free survival of 5.3 months (Chapman et al. N Engl J Med 2011). A caveat to this clearly positive clinical trial is that there are patients whose tumors have the same mutations who did not respond to vemurafenib and amongst those who did, the duration of response remains limited. In tumors that harbor driver mutations or other genetic aberrations, mechanisms of resistance are of relevance, including both primary resistance that renders them insensitive to specific inhibitors, and evolved/acquired resistance that results in tumor growth despite initial sensitivity. Comprehensive molecular profiling of pre-treatment tumor samples and at progression would be informative to offer insight into such mechanisms of primary and evolved/acquired resistance and guide clinical trial designs to counteract them. Combinatorial strategies may be considered upfront for patients who have tumor characteristics suggestive of primary resistance. For patients with sensitive disease which evolve/acquire resistance during treatment, a repeat tumor biopsy may help detect newly emerged aberrations that are potentially druggable. Additional areas of interest in defining tumor resistance include the observations that same mutation may have different degrees of functionality in different tumor types (e.g. BRAF mutations in melanoma versus colorectal cancer) and different mutations of the same gene may confer different drug sensitivities (e.g. activating and inactivating BRAF mutations), These context-dependent phenomena further complicate the planning of histology-agnostic, mutation-specific clinical trials using targeted agents. Efforts to provide detailed tumor molecular profiling information in clinical trials are relevant to help distinguish signatures of sensitivity versus resistance.
Abstract IA13: Finding biomarkers of response to targeted cancer drugs through functional genetics
Authors: Bernards Rene
Published: March 2013

Unresponsiveness to therapy remains a significant problem in the treatment of cancer, also with the new classes of cancer drugs. In my laboratory, we use functional genetic approaches to identify biomarkers that can predict responsiveness to targeted cancer therapeutics; drugs that specifically inhibit molecules or pathways that are often activated in cancer. Nevertheless, it remains poorly explained why a significant number of tumors do not respond to these therapies. We aim to elucidate the molecular pathways that contribute to unresponsiveness to targeted cancer therapeutics using a functional genetic approach. This will yield biomarkers that may be useful to predict how individual patients will respond to these drugs. Furthermore, this work may allow the development of drugs that act in synergy with the established drug to prevent or overcome drug resistance.

To identify biomarkers that control tumor cell responsiveness to cancer therapeutics, we use multiple complementary approaches. First, we use genome wide loss-of-function genetic screens (with shRNA interference libraries) in cancer cells that are sensitive to the drug-of-interest to search for genes whose down-regulation confers resistance to the drug-of-interest (resistance screens). In addition, we use shRNA screens with a low dose of the drug to screen for genes whose inhibition enhances the toxicity of the cancer drug (sensitizer screens). As a third approach, we use gain of function genetic screens in which we search for genes whose over-expression modulates drug responsiveness. Once we have identified candidate drug response biomarkers in relevant cell line models, we ask if the expression of these genes is correlated with clinical response to the drug-of-interest. For this, we use tumor samples of cancer patients treated with the drug in question and whose response to therapy is documented.

In a fourth and distinct approach we perform high throughput sequencing of the “kinome” (some 600 genes) of tumor samples to identify connections between cancer genotype and drug responses.

Examples of these approaches to identify biomarkers of response to different cancer drugs will be presented.
Abstract IA15: VEGF-targeted therapies: Biomarkers, biology, and understanding resistance
Authors: Ellis Lee M.
Published: March 2013

Despite major advances in the clinical development of VEGF-targeted therapy, most patients' tumors are considered to be inherently resistant. When response is obtained, the benefit of VEGF inhibitors is transient, lasting weeks to months, in contrast to the original hypothesis that cancer would be converted to a chronic disease with this therapeutic approach. The mechanisms of action of this class of drugs vary among tumor types and therefore resistance mechanisms and biomarkers are likely to be tumor-type specific. Clinical studies have begun to uncover potential biomarkers that require validation in prospective studies. For example, data reported at the 2011 ESMO meeting suggested that circulating VEGF, as detected by a new ELISA, may be able to determine the population of patients likely to benefit from the use of bevacizumab, in some, but not all studies. In addition, several clinical studies reported this past year have shown that continued VEGF-inhibition through multiple lines of therapy may be of benefit to patients. These clinical studies suggest that resistance may be a relative term: although patients eventually progress, continued VEGF inhibition beyond initial progression may be of clinical benefit in selected patients. There is much work to be done in the field of biomarkers for VEGF-targeted therapies, but these recent clinical studies provide insights that will hopefully provide the foundation for significant improvements in patient outcomes.
Abstract IA3: PI3K inhibitors in ER+ breast cancer: Next steps
Authors: Ellis Matthew J. , Crowder Robert , Ma Cynthia X.
Published: March 2013

Gain and loss of function mutations in the PI3 kinase pathway are extremely common in breast cancer and can be seen in all five intrinsic subtypes (Luminal A, Luminal B, HER2+, Basal-like and Claudin Low). However there is a subtype-selective spectrum of activation events including gain of kinase mutations/amplifications (e.g. PIK3CA, HER2 and AKT1) and loss of negative regulators (e.g.PTEN, INPP4B and PIK3R 1). These lesions must be matched to a large number of agents in development that target PI3K, AKT and mTOR. These agents have variable specificities, toxicities, pharmacokinetics and administration routes which affect the development of combinations with standard therapies and also with each other. In keeping with very broad clinical opportunities, positive findings in preclinical models in both luminal and basal-like breast cancer will be presented. Ongoing clinical trial designs will also be discussed and biomarker approaches for patient selection and stratification will be considered.
Abstract IA6: Targeting PI3K in cancer, mechanistic insights from genetic mouse models
Authors: Zhao Jean J.
Published: March 2013

The class 1a phosphatidylinositol 3-kinase (PI3K) signaling pathway is frequently activated in human cancer by a number of mechanisms, including amplification and/or activation of RTKs (receptor tyrosine kinases), loss of PTEN (phosphatase and tensin homologue) function and somatic mutation in the PIK3CA gene that encodes the p110α catalytic subunit of PI3K. Thus PI3K represents an attractive target for cancer therapy and tremendous efforts are being devoted to developing effective PI3K inhibitors for cancer treatment. Despite the great promise of targeted therapy, drug resistance is likely to develop, causing treatment failure. To elucidate resistance mechanisms to PI3K-targeted therapy, we constructed mouse models of breast cancer and brain tumors conditionally expressing an oncogenic allele of PIK3CA. Interestingly, we found that oncogenic PIK3CA-driven tumors in both mammary gland and brain tissues frequently recur in PI3K pathway-dependent and -independent manners. Genomic and functional analyses of recurrent tumors revealed multiple lesions, providing mechanistic insights for tumor recurrence and resistance to PI3K-targeted therapy.
Abstract PR4: Incomplete inhibition of phosphorylation of 4EBP1 is a mechanism for KRAS driven resistance to the ATP-competitive mTOR inhibitor PP242 in colorectal cancer
Authors: Ducker Gregory S. , Atreya Chloe E. , Shokat Kevan M. , Warren Robert S.
Published: March 2013

The mammalian target of rapamycin (mTOR) regulates cell growth by integrating nutrient and growth factor signaling and has been strongly implicated in cancer. But mTOR is not an oncogene, and which tumors will be resistant and sensitive to new ATP-competitive mTOR inhibitors now in clinical trials remains unknown. We screened a panel of 357 human cancer cell lines to identify common markers of resistance and sensitivity to the mTOR inhibitor PP242. Analysis by tissue of origin and genetic mutation revealed colorectal origin as the most statistically significant marker for resistance to therapy, while KRAS and PI3K mutations were the most significant genetic markers for resistance and sensitivity respectively. Notably, KRAS mutations are overrepresented in colorectal cancer (CRC) cell lines. Within CRC cell lines, those with KRAS mutations were most resistant to PP242 while those with other mutations were more sensitive; cell lines with co-mutation of PI3K and KRAS had intermediate sensitivity. Immunoblot analysis of the signaling networks downstream of mTOR revealed that the degree of cellular growth inhibition induced by PP242 was directly correlated with inhibition of phosphorylation of the translational repressor eIF4E binding protein 1 (4EBP1), but not ribosomal protein S6. We were able to recapitulate these observations using a pair of KRAS mutant isogenic cell lines differing only by a PI3K mutation. PP242 treatment of the PI3K mutant cell line increased the observed inhibition of 4EBP1 phosphorylation and improved the drug response. In an efficacy trial of PP242 in primary CRC xenograft tumors, resistance to inhibition of 4EBP1 phosphorylation was again observed in KRAS mutant tumors and these tumors responded poorly compared to KRAS WT controls. We show that, in the absence of PI3K co-mutation, KRAS mutations are associated with resistance to PP242 and that this is specifically linked to changes in the 4EBP1 phosphorylation state in both CRC cell lines and primary patient xenografts.

This proffered talk is also presented as Poster A35.
Effects of Lycopene on Protein Expression in Human Primary Prostatic Epithelial Cells
Authors: Qiu Xi , Yuan Yang , Vaishnav Avani , Tessel Michael A. , Nonn Larisa , van Breemen Richard B.
Published: March 2013

Clinical trials and animal studies have suggested that lycopene, the red carotenoid found in tomatoes, might be useful for the prevention of prostate cancer in the diet or as a dietary supplement through a variety of chemoprevention mechanisms. As most mechanism of action studies have used prostate cancer cells or males with existing prostate cancer, we investigated the effects of lycopene on protein expression in human primary prostatic epithelial cells. After treatment with lycopene at a physiologically relevant concentration (2 μmol/L) or placebo for 48 hours, the primary prostatic epithelial cells were lysed and fractionated using centrifugation into cytosolic/membrane and nuclear fractions. Proteins from lycopene-treated and placebo-treated cells were trypsinized and derivatized for quantitative proteomics using isobaric tags for relative and absolute quantitation (iTRAQ) reagent. Peptides were analyzed using two-dimensional microcapillary high-performance liquid chromatography-tandem mass spectrometry to identify proteins that were significantly upregulated or downregulated following lycopene exposure. Proteins that were most affected by lycopene were those involved in antioxidant responses, cytoprotection, apoptosis, growth inhibition, androgen receptor signaling, and the Akt/mTOR cascade. These data are consistent with previous studies suggesting that lycopene can prevent cancer in human prostatic epithelial cells at the stages of cancer initiation, promotion, and/or progression. Cancer Prev Res; 6(5); 419–27. ©2013 AACR.
Is Lycopene an Effective Agent for Preventing Prostate Cancer?
Authors: Sporn Michael B. , Liby Karen T.
Published: March 2013

The role of lycopene, an open-chain carotenoid found in tomatoes and devoid of retinoid activity, as an anticarcinogenic, chemopreventive agent, especially for use in prostate cancer, is still under active investigation. In this issue, Qui and colleagues show that lycopene induces responses in human prostate epithelial cells that are antiproliferative, antioxidative, and anti-inflammatory, as well as downregulating targets in the androgen receptor signaling pathway. In this perspective, we review aspects of the molecular and cellular biology of lycopene that support its use for prevention of prostate cancer. Whether lycopene itself or its metabolites induce most of its benefits is still uncertain. At present, meta-analysis of clinical studies of lycopene for prevention of prostate cancer in men does not yet support the definitive clinical use of this carotenoid in a preventive setting. Cancer Prev Res; 6(5); 384–6. ©2013 AACR.
Morphological Changes of Bladder Mucosa in Patients Who Underwent Instillation with Combined Sodium Hyaluronic Acid-Chondroitin Sulphate (Ialuril®)
Authors: Costantini E. , Lazzeri M. , Pistolesi D. , Del Zingaro M. , Frumenzio E. , Boni A. , Pietropaolo A. , Fragalà E. , Porena M.
Published: March 2013

Objective: To investigate what changes are endoscopically evident after glycosaminoglycans (GAGs) therapy by hyaluronic acid (HA) and chondroitin sulphate (CS) (Ialuril®) in female patients affected by bladder pain syndrome(BPS)/ interstitial cystitis (IC) or recurrent urinary tract infections (rUTIs). Patients and Methods: 21 female patients over 18 years affected by rUTIs or BPS/IC received intravesical instillation of HA and CS (4 weekly instillations followed by 2 instillations every 2 weeks and 2 instillation monthly). Post-treatment evaluation included cystoscopy and patient assessment of improvement in symptoms and satisfaction on a visual analogue scale (VAS) from 0 to 10. Results: The post-treatment endoscopy showed a positive effect on bladder mucosa morphology. In 2 cases, treatment did not change endoscopic findings and clinical symptoms. In the other patients, when macroscopic features of the bladder mucosa normalized, the clinical picture improved. Conclusions: GAGs therapy by HA and CS (Ialuril) improves the morphology of bladder mucosa in patients with rUTI or BPS/IC.
Cross-Talk between Available Guidelines for the Management of Patients with Beta-Thalassemia Major
Authors: Musallam Khaled M. , Angastiniotis Michael , Eleftheriou Androulla , Porter John B.
Published: March 2013

Efforts to optimize the management of patients with β-thalassemia major (TM) continue to expand. Evidence from biomedical research evaluating safe and careful processing measures of blood products, the efficacy and safety of oral iron chelators, and noninvasive techniques for the assessment of iron overload are translated into better patient outcomes. The construction of TM management guidelines facilitated the incorporation of such evidence into practice. However, as several aspects of the management of TM remain controversial or governed by resource availability, a concern regarding potential variations in recommendations made by the different guidelines becomes rational, especially for physicians treating TM patients outside countries where the guidelines were constructed. In this work, we overview currently available guidelines for the management of TM and explore apparent similarities and differences between them. The evaluated guidelines included the Thalassaemia International Federation, US, Canadian, UK, Italian and Australian guidelines. We noted a general consensus for most aspects of management, although some guidelines provided more comprehensive and contemporary recommendations than others. We did not identify differences warranting concern, although minor differences in iron overload assessment strategy and more notable variations in the recommendations for iron chelation therapy were observed.
Traumatic Urethral Diverticula: A Rare Malformation of the Male Urethra
Authors: Li Xu , Sa Yinglong , Jin Chongrui , Xu Yuemin
Published: March 2013

Urethral diverticula are pouches opening into the urethral canal that may be either acquired or congenital. Diverticula of the female urethra are fairly common. However, diverticula of the male urethra are relatively uncommon abnormalities. Clinical manifestation is frequently described with irritative and/or obstructive voiding symptoms or ventral bulging, but manifestation as a scrotal mass is extremely rare. We report a giant urethral diverticulum presenting as a scrotal mass in an adult male.
Metallothioneins I/II Expression in Rat Strains with Genetically Different Susceptibility to Experimental Autoimmune Encephalomyelitis
Authors: Grubić-Kezele Tanja , Jakovac Hrvoje , Tota Marin , Čanadi-Jurešić Gordana , Barac-Latas Vesna , Milin Čedomila , Radošević-Stašić Biserka
Published: March 2013

Objectives: Compared to the Dark Agouti (DA), the Albino Oxford (AO) rat strain exhibits lower susceptibility to the induction of experimental autoimmune encephalomyelitis (EAE). Here, we investigated the potential contribution of the heavy metal-binding proteins metallothioneins (MTs) I/II to these effects. Methods: Rats were immunized with bovine brain homogenate emulsified in complete Freund's adjuvant or only with complete Freund's adjuvant. The expression patterns of MTs mRNA and proteins and tissue concentrations of Zn2+ and Cu2+ were estimated in the brain and in the liver on days 7 and 12 after immunization, by real-time PCR, immunohistochemistry and inductively coupled plasma spectrometry, respectively. Additionally, the hepatic transforming growth factor beta and nuclear factor kappa B immunoreactivities were tested. Results: Clinical signs of EAE were not induced in AO rats, but they upregulated the expression of MT I/II proteins in the brain (hippocampus and cerebellum) and in the liver, similarly as DA rats. The transcriptional activation of MT-I occurred, however, only in DA rats, which accumulated also more zinc in the brain and in the liver. In contrast, intact AO rats had greater hepatic MT-I mRNA immunoreactivity and more Cu2+ in the hippocampus. Besides, in immunized AO rats a high upregulation of transforming growth factor beta and nuclear factor kappa B immunoreactivities was found in several hepatic structures (vascular endothelium, Kupffer cells and hepatocytes). Conclusions: Our data show that AO and DA rats differ in constitutive and inductive MT-I gene expression in the brain and in the liver, as well as in the hepatic cytokine profile, suggesting that these mechanisms may contribute to the discrepancy in the susceptibility to EAE.
Time-Dependent Resistance to Erythropoiesis-Stimulating Agent and Mortality in Hemodialysis Patients in the Japan Dialysis Outcomes and Practice Patterns Study
Authors: Fujikawa Tetsuya , Ikeda Yumiko , Fukuhara Shunichi , Akiba Takashi , Akizawa Tadao , Kurokawa Kiyoshi , Saito Akira
Published: March 2013

Background/Aims: Resistance to erythropoiesis-stimulating agent (ESA) is associated with mortality in hemodialysis (HD) patients. Time-dependent variability of ESA resistance has been not investigated adequately, although consistently high ESA resistance is expected to be a high risk for mortality. Our aim, therefore, was to investigate consistently high ESA resistance as an independent predictor of mortality in HD patients. Methods: This study evaluated 2,104 Japanese HD patients in the Dialysis Outcomes and Practice Patterns Study (DOPPS) III. ESA resistance index (ERI) was defined as the weekly weight-adjusted dose of ESA divided by hemoglobin concentration. The average ERI was calculated from ERI levels every 4 months throughout the observation period for each patient. To assess the size of the fluctuation in average ERI during the observation periods according to ERI quartiles at the enrollment periods, six patient groups were defined on the basis of patterns of ERI level fluctuation: low-low (Low), intermediate-intermediate (Intermediate), high-high (High), low-intermediate, intermediate-high, and low-high. Results: The number of deaths among the patients was 227 (10.8%), which included 113 (5.4%) cases of cardiovascular disease (CVD). In multivariate analysis after adjustment for age, albumin, C-reactive protein, comorbidities, etc., the High group was independently and significantly related to all-cause and CVD-related mortality (OR = 2.33, 95% CI: 1.33-4.07, p = 0.002, and OR = 2.09, 95% CI: 1.05-4.14, p = 0.035, respectively). Conclusion: Factoring out fluctuating ERI increases the ability of consistently high ERI levels as an independent risk factor for all-cause and CVD mortality in HD patients.
Effect of New Method of Intracoronary Adenosine Injection during Primary Percutaneous Coronary Intervention on Microvascular Reperfusion Injury - Clinical Outcome and 1-Year Follow-Up
Authors: Grygier Marek , Araszkiewicz Aleksander , Lesiak Maciej , Grajek Stefan
Published: March 2013

Objectives: The aim of this single-center, randomized placebo-controlled trial in 70 consecutive patients (64 ± 14 years) with acute myocardial infarction was to examine the role of a new protocol of adenosine administration during primary angioplasty on immediate electrocardiographic and angiographic results, clinical outcome and 1-year follow-up. Methods: Group A (n = 35) twice received intracoronary adenosine through the guiding catheter: immediately after crossing the lesion of the infarct-related artery with guidewire and then after first balloon inflation. Group B (n = 35) received placebo. Results: Resolution of ST segment elevation was more frequently observed in the adenosine than in the placebo group (p < 0.01). Percutaneous coronary intervention (PCI) resulted in borderline better TIMI 3 flow after the procedure in the adenosine group than in the placebo group. Myocardial blush grade 3 at the end of the procedure was significantly improved in the adenosine compared to the placebo group (p < 0.05). At 1-year the composite end-point of death, recurrent myocardial infarction, heart failure and clinically driven target vessel revascularization was present in 8 patients in the adenosine group and 16 patients in placebo group (p < 0.05). Conclusions: Intracoronary adenosine improved electrocardiographic and angiographic results in patients undergoing primary PCI and seemed to be associated with more favorable clinical course.
G-Protein-Coupled Receptor Kinase-5 Mediates Inflammation but Does Not Regulate Cellular Infiltration or Bacterial Load in a Polymicrobial Sepsis Model in Mice
Authors: Packiriswamy Nandakumar , Lee Taehyung , Raghavendra Pongali B. , Durairaj Haritha , Wang Hongbing , Parameswaran Narayanan
Published: March 2013

NFκB-dependent signaling is an important modulator of inflammation in several diseases including sepsis. G-protein-coupled receptor kinase-5 (GRK5) is an evolutionarily conserved regulator of the NFκB pathway. We hypothesized that GRK5 via NFκB regulation plays an important role in the pathogenesis of sepsis. To test this we utilized a clinically relevant polymicrobial sepsis model in mice that were deficient in GRK5. We subjected wild-type (WT) and GRK5 knockout (KO) mice to cecal ligation and puncture (CLP)-induced polymicrobial sepsis and assessed the various events in sepsis pathogenesis. CLP induced a significant inflammatory response in the WT and this was markedly attenuated in the KO mice. To determine the signaling mechanisms and the role of NFκB activation in sepsis-induced inflammation, we assessed the levels of IκBa phosphorylation and expression of NFκB-dependent genes in the liver in the two genotypes. Both IκBa phosphorylation and gene expression were significantly inhibited in the GRK5 KO compared to the WT mice. Interestingly, however, GRK5 did not modulate either immune cell infiltration (to the primary site of infection) or local/systemic bacterial load subsequent to sepsis induction. In contrast GRK5 deficiency significantly inhibited sepsis-induced plasma corticosterone levels and the consequent thymocyte apoptosis in vivo. Associated with these outcomes, CLP-induced mortality was significantly prevented in the GRK5 KO mice in the presence of antibiotics. Together, our studies demonstrate that GRK5 is an important regulator of inflammation and thymic apoptosis in polymicrobial sepsis and implicate GRK5 as a potential molecular target in sepsis.
Effect of Vitamin D Receptor Activator Therapy on Vitamin D Receptor and Osteocalcin Expression in Circulating Endothelial Progenitor Cells of Hemodialysis Patients
Authors: Cianciolo Giuseppe , La Manna Gaetano , Della Bella Elena , Cappuccilli Maria Laura , Angelini Maria Laura , Dormi Ada , Capelli Irene , Laterza Claudio , Costa Roberta , Alviano Francesco , Donati Gabriele , Ronco Claudio , Stefoni Sergio
Published: March 2013

Background: The effects of vitamin D receptor (VDR) and osteocalcin (OC) expression as well as VDR agonist (VDRA) therapy on circulating endothelial progenitor cells (EPCs) has not been elucidated yet. Methods: We therefore analyzed EPCs in 30 healthy controls and 82 patients undergoing dialysis (no VDRA therapy: 28; oral calcitriol: 30, and intravenous paricalcitol, PCTA: 24). The percentage of EPCs (CD34+/CD133-/KDR+/CD45-) expressing VDR or OC, and VDR and OC expression defined by mean fluorescence intensity (MFI) were analyzed using flow cytometry. The in vitro effect of VDRAs was evaluated in EPCs isolated from each patient group. Results: The percentage of VDR+ EPCs correlated positively with VDRA therapy and 25(OH)D, and negatively with diabetes, C-reactive protein, hemoglobin and osteopontin. VDR-MFI correlated positively with VDRA therapy, parathyroid hormone (PTH) and 25(OH)D, and negatively with diabetes and osteopontin. The percentage of OC+ EPCs correlated positively with the calcium score, PTH and phosphate, and negatively with 25(OH)D. OC-MFI correlated positively with calcium score, PTH, phosphate and hemoglobin, and negatively with albumin, 25(OH)D and osteopontin. Cell cultures from patients without VDRA therapy had the highest levels of calcium deposition and OC expression, which both significantly decreased following in vitro VDRA administration: in particular extracellular calcium deposition was only reduced by adding PCTA. Conclusions: Our data suggest that 25(OH)D serum levels and VDRA therapy influence VDR and OC expression on circulating EPCs. Since OC expression may contribute to vascular calcification, we hypothesize a putative protective role of VDRA therapy.
Curse on Two Generations: A History of Congenital Syphilis
Authors: Obladen Michael
Published: March 2013

Before the microbiologic era, venereal diseases were poorly distinguished. Congenital syphilis was believed to be transmitted during conception by the father's sperm, during delivery in the birth canal, or from infected milk or breasts. The most frequent maternofetal transmission was not considered because the mother's primary infection remained undiagnosed. The concept of treating infants with mercury transmitted by nurses' milk prompted the founding of a specialized infant hospital in Vaugirard in 1780: lactating syphilitic women received mercury orally and by rubbing it into the skin. Their own infant and a second infected infant from the foundling hospital were believed to be cured by their milk. Underwood described snuffles in 1789 and Bertin periosteal bone disease in 1810. Tardive congenital lues with keratitis, deafness, and notched upper incisors were described by Hutchinson in 1863. Feeding remained difficult, as wet nursing transmitted syphilis to the nurse and other infants. Specialized institutions tried goat or donkey milk. A debate between contagionists assuming exclusively maternal infection and hereditists assuming germinal transmission by the father's sperm continued throughout the 19th century. Schaudinn and Hoffmann identified Spirochaeta pallida in 1905. When Ehrlich discovered the efficacy of salvarsan in 1910, Noeggerath treated infants with the new drug, pioneering the injection into scalp veins. In 1943, Lentz and Ingraham established penicillin treatment for congenital syphilis. Whereas this drug effectively prevented maternofetal transmission, treating infants remained difficult due to the Jarisch-Herxheimer reaction.
The British Society for Gynaecological Endoscopy Endometriosis Centres Project
Authors: Saridogan Ertan , Byrne Dominic
Published: March 2013

Management of advanced endometriosis frequently requires a multidisciplinary team approach and international guidelines suggest treatment in centres of expertise. Due to variability of published outcome data, prospective data collection and standardisation of reporting systems have been suggested to improve our understanding of surgical outcomes. The British Society for Gynaecological Endoscopy (BSGE) Endometriosis Centres were established to manage rectovaginal endometriosis, to collect treatment and outcome data, and to provide these data to patients, clinicians and healthcare commissioners. The BSGE Endometriosis Centres Project works on the principle of voluntary participation. Centres that would like to be recognised or accredited as a BSGE Endometriosis Centre need to fulfil a number of basic requirements including working in appropriate multidisciplinary clinical teams, auditing their outcome and having sufficient workload to maintain their surgical skills. The project has already had an impact on where the patients with advanced endometriosis are treated in the United Kingdom. Patients and healthcare professionals are becoming aware of their presence and more patients with the condition are being referred to these centres. It is also expected that the accredited centre status would be required for funding by healthcare commissioners for this type endometriosis.