bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 176: Development of 1- and 3-phosphohistidine monoclonal antibodies utilized as novel tools to study histidine phosphorylation
Authors: Speckmann Wayne A. , Santos Mark , Shea Reshmie , Geiman Sheri , Saadat Arazo , Hoberg James , Zheng Wei
Published: July 2016

Phosphorylation of proteins is one of the most common forms of post-translational modification (PTMs) and is a mechanism for regulating enzyme activity or protein function. There are several amino acids that can be phosphorylated but despite its importance in cancer and tumor metastasis histidine phosphorylation (pHis) has been highly overlooked. One of the key reasons is the inherent challenge associated with studying pHis legitimately, due largely in part to the lack in availability of specific antibodies. Unlike serine, threonine, and tyrosine which all form acid-stable phosphoester (P-O) bonds upon phosphorylation, histidine forms heat and acid-labile phosphoramidate (P-N) bonds. Because of the instability of the P-N bond, it is nearly impossible to assess the prevalence of pHis accurately.

Here, we outline the development of selective, sequence independent anti-1-pHis and anti-3-pHis monoclonal antibodies which have addressed this current need for specific antibodies to detect the phosphorylation of histidine. The antibodies developed specifically recognize 1-pHis and 3-pHis and are isomer specific. In addition, they do not cross-react with phospho-tyrosine (pTyr), whereas, the first and second generation “pan-pHis” polyclonal antibodies against 3-pHis have been reported to do so. Moreover, it has been suggested that pHis could be as abundant as pTyr, comprising of approximately 1% of all known phosphorylation in cells, making specific monoclonal antibodies for detection highly desirable.

Validation data of the four monoclonal phospho-Histidine antibodies developed (clones SC50-3 and SC1-1 for assessment of 1-pHis activities; clones SC56-2 and SC39-6 for assessment of 3-pHis activities) demonstrate the specificity and broad utility of these monoclonal antibodies when identifying pHis substrates, making them useful tools in a variety of immunological and biological assays.

Citation Format: Wayne A. Speckmann, Mark Santos, Reshmie Shea, Sheri Geiman, Arazo Saadat, James Hoberg, Wei Zheng. Development of 1- and 3-phosphohistidine monoclonal antibodies utilized as novel tools to study histidine phosphorylation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 176.
Abstract 1791: Effects of calcium and vitamin D supplementation on circulating markers of inflammation
Authors: Fedirko Veronika , Latash Julia , Bostick Roberd M. , Barry Elizabeth L. ,   John A. Baron on behalf of Investigators in the PPS4 Study
Published: July 2016

Chronic inflammation plays a major role in colon carcinogenesis. Recent evidence from experimental models suggests that vitamin D and calcium may reduce systemic inflammation and local inflammation in the colorectum, thus contributing to reducing risk for colorectal neoplasia. However, human experimental evidence to support these purported anti-inflammatory effects remains inconclusive, coming from very few, small, mainly short-term clinical studies with a variety of doses and administration regimens, with and without calcium supplementation in very different patient populations, and with a limited number of inflammatory biomarkers measured. To address this, we tested the effects of calcium (1.2 g/d) and/or vitamin D3 (1,000 IU/d) supplementation for 3 or 5 years on circulating pro- (IFNγ, TNFα, IL-6, IL-8, IL-12p40) and anti-inflammatory (IL-10) cytokines in 471 participants with previous colorectal adenoma in a recently completed, multi-center, randomized, double-blind, placebo-controlled, modified 2 × 2 factorial chemoprevention clinical trial (NCT00153816). The cytokines were measured in plasma using multiplex electrochemiluminescence detection-based immunoassays. Over a 3- or 5-year treatment period, we found no appreciable effects of calcium and/or vitamin D supplementation on pro-inflammatory cytokines (all P-values > 0.3). However, there was a suggestion that vitamin D may favorably modulate concentrations of IL-10, an anti-inflammatory cytokine. In the calcium, vitamin D, and calcium plus vitamin D groups, the relative increase vs. placebo in IL-10 from baseline to end of follow-up was 3% (P = 0.78), 15% (P = 0.15), 13% (P = 0.35). Our preliminary results indicate no considerable effects of calcium and vitamin D treatment on circulating pro-inflammatory cytokines, and might suggest that vitamin D may increase circulating levels of anti-inflammatory cytokine IL-10, which plays an important role in the Th2-type immune response.

Citation Format: Veronika Fedirko, Julia Latash, Roberd M. Bostick, Elizabeth L. Barry, John A. Baron on behalf of Investigators in the PPS4 Study. Effects of calcium and vitamin D supplementation on circulating markers of inflammation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1791.
Abstract 1795: The feasibility study of a randomized cancer screening trial in China
Authors: Hu Ping , Dai Min , Shi Jufang , Ren Jiansong , Li Jiang , Liao Xianzhen , Du Lingbin , Liu Yuqing , Chen Zhaoli , Wu Ning , Liu Qian , Pinsky Paul , Prorok Philip , Fagerstrom Richard , Taylor Martina , Kramer Barnett , He Jie
Published: July 2016

Lung cancer has been the leading cause of cancer death in China since the 1990s and the disease burden likely will increase. The US National Lung Screening Trial (NLST) showed a 16-20% mortality reduction in a high-risk population that had undergone Low-Dose Computed Tomography (LDCT). China would like to confirm the main result from the US NLST in the Chinese population. The aim of this study is to obtain necessary information on performance of the organizations involved in designing, conducting, monitoring and managing the study. Another goal is to test colorectal cancer screening strategies in China. This is a first randomized trial on lung and colorectal cancer screening in China. The feasibility study will aid in the development of a practical design for the Randomized Cancer Screening Trial in China.

This study has been carried out in three cites (Changsha city, Hunan province; Lanzhou city, Gansu province; and Haining city, Zhejiang province) since August, 2014. Individuals at elevated risk of lung cancer were randomized into three arms: annual low-dose helical CT exams for three years (T0, T1, T2) and baseline colonoscopy (T0); two low-dose helical CT exams (T0, T2) plus annual faecal immunochemical test (OC-faecal immunochemical tests at T0, T1, T2); and annual InSure-faecal immunochemical tests combined with Septin 9 test (T0, T1, T2). All participants will be followed for at least three years from randomization to yield data relevant to answering the study objectives.

As of March 31, 2015, a total of 2696 eligible participants were enrolled at 3 cancer hospitals and randomly assigned to three study arms (894 in arm 1, 902 in arm 2, and 900 in arm 3). The gender distribution is 53% male and 47% female. Baseline characteristics of the study population by age, gender and total pack-year exposure were balanced. The rate of adherence to LDCT screening at the first round (T0) was 89.0%. 1598 participants received LDCT in the first round. The rate of abnormality was 77.4% with LDCT and 6.5% and 6.1% with suspicious for lung cancer for arm 1 and arm 2 respectively at T0. 2150 participants received colorectal cancer screening. The positivity was 34.1% with colonoscopy screening, 8.0% with OC-FIT test, 4.5% with InsureFIT test, and 5.8% with Septin9 test.

Citation Format: Ping Hu, Min Dai, Jufang Shi, Jiansong Ren, Jiang Li, Xianzhen Liao, Lingbin Du, Yuqing Liu, Zhaoli Chen, Ning Wu, Qian Liu, Paul Pinsky, Philip Prorok, Richard Fagerstrom, Martina Taylor, Barnett Kramer, Jie He. The feasibility study of a randomized cancer screening trial in China. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1795.
Abstract 1801: FOXA1, a novel regulator of neuroendocrine differentiation
Authors: Kim Jung , Jin Hongjian , Zhao Jonathan , Parimi Vamsi , Yang Ximing , Yu Jindan
Published: July 2016

Neuroendocrine prostate cancer (NEPC) is a subtype of prostate cancer that is highly aggressive and exhibits a neuroendocrine phenotype, being distinct from prostate adenocarcinoma. Cases of NEPC are predicted to increase rapidly following broader use of new-generation hormonal therapies including abiraterone and enzalutamide. Although androgen deprivation and cytokine induction have been previously suggested to induce neuroendocrine differentiation of prostate cancer, little is known regarding the underlying molecular mechanisms. FOXA1 is a forkhead box family transcription factor that has been shown to act as a pioneer factor for androgen receptor (AR), thereby defining the prostatic transcriptional program. We have recently shown that FOXA1 plays an important role in suppressing epithelial-to-mesenchymal transdifferentiation, but is often downregulated in castration-resistant prostate cancer (CRPC). Along this line, in the present study, we showed that FOXA1 loss, achieved by shRNA-mediated knockdown, led to cell morphology changes typical of neuroendocrine differentiation, accompanied by a strong increase of NEPC marker enolase2 (ENO2), while ectopic FOXA1 overexpression reverses these alterations. This was further linked to ERK phosphorylation, which has been previously reported to be up-regulated following neuroendocrine differentiation. Moreover, to understand the mechanism underlying FOXA1 regulation of neuroendocrine differentiation, we performed bioinformatics analysis and nominated interleukin 8 (IL8) as a direct target of FOXA1 and its up-regulation is critical in mediating neuroendocrine differentiation of FOXA1-depleted cells. Finally, we validated a negative correlation between FOXA1 and neuroendocrine differentiation in primary prostate cancer specimens through analysis of publically available RNA-seq data as well as direct IHC staining of various NEPC markers. In summary, we report a new role of FOXA1 as a critical inhibitor of neuroendocrine differentiation and the importance of balanced FOXA1 expressing in the maintenance of the epithelial phenotype of prostate cancer.

Citation Format: Jung Kim, Hongjian Jin, Jonathan Zhao, Vamsi Parimi, Ximing Yang, Jindan Yu. FOXA1, a novel regulator of neuroendocrine differentiation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1801.
Abstract 1802: ER-positive breast cancer prevention with the use of hormone replacement therapy
Authors: Dembo Anna G. , Greene Geoffrey L.
Published: July 2016

Menopause occurs in women between the ages of 45 and 55 and often results in undesirable vasomotor symptoms. Hormone replacement therapy (HRT) alleviates these symptoms, including hot flashes, difficulty sleeping, fatigue, and vaginal atrophy. HRT also prevents osteoporosis. PremPro, a currently available HRT formulation that combines conjugated equine estrogens (CE) with medroxyprogesterone acetate, increases the risk of breast cancer. Due to the perceived risk based largely on the results of the Women's Health Initiative (WHI) trial, the number of women taking HRT has decreased dramatically. Duavee, a new form of HRT that combines CE and bazedoxifene (BZA), a selective estrogen receptor modulator (SERM) and degrader (SERD), has recently been approved by the FDA for treatment of moderate to severe hot flashes and to reduce the risk of osteoporosis. Importantly, this CE/BZA mixture not only relieves symptoms associated with menopause, but it also does not stimulate the breast or uterus. Although several preclinical studies suggest that CE/BZA might be protective in the breast, the mechanism of action of this new combination therapy is not known. Our goal, therefore, is to elucidate the underlying molecular mechanisms by which CE/BZA differentially affects estrogen receptor alpha (ERα) action in the mammary gland, using transcriptome and whole genome occupancy analysis in breast cancer cell lines. We are also studying the effects of CE/BZA on early mammary cancer progression in the polyoma middle T antigen (PyMT) transgenic mouse model, which has been shown to be sensitive to estrogens. In addition, we are studying the effects in an ERα-positive patient-derived xenograft mouse model. We have determined that CE modulates gene expression in MCF7 cells similar to 17β-estradiol (E2), and that BZA is able to inhibit these effects. We have also observed that CE increases ERα occupancy versus E2 at response elements associated with some estrogen target genes, whereas CE/BZA decreases this occupancy. In the PyMT mouse model, we have determined that CE/BZA significantly delays the onset of mammary tumors in ovariectomized mice and prolongs their survival when compared to E2 and CE treatment alone. An improved understanding of the molecular mechanisms of CE/BZA action in hormone sensitive breast cancer cell and animal models should have important implications for women considering HRT.

Citation Format: Anna G. Dembo, Geoffrey L. Greene. ER-positive breast cancer prevention with the use of hormone replacement therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1802.
Abstract 1806: TRIM24 is an oncogenic transcriptional activator in prostate cancer
Authors: Groner Anna C. , Cato Laura , de Tribolet-Hardy Jonas , Bernasocchi Tiziano , Zhong Qing , Fankhauser Christian , Fritz Christine , Poyet Cédric , Wagner Ulrich , Garraway Levi A. , Wild Peter J. , Theurillat Jean-Philippe , Brown Myles
Published: July 2016

Androgen receptor (AR) signaling is a key driver of prostate cancer (PC). While androgen-deprivation therapy is transiently effective in advanced disease, tumors often progress to a lethal castration-resistant state (CRPC). We show that recurrent PC-driver mutations in SPOP stabilize the TRIM24 protein, which promotes proliferation under low androgen conditions. TRIM24 augments AR signaling, and AR and TRIM24 co-activated genes are significantly up-regulated in CRPC. Expression of TRIM24 protein increases from primary PC to CRPC, and both TRIM24 protein levels and the AR/TRIM24 gene signature predict disease-recurrence. Analyses in CRPC cells reveal that the TRIM24 bromodomain and the AR-interacting motif are essential to support proliferation. These data provide a rationale for therapeutic TRIM24 targeting in SPOP-mutant and CRPC patients.

Citation Format: Anna C. Groner, Laura Cato, Jonas de Tribolet-Hardy, Tiziano Bernasocchi, Qing Zhong, Christian Fankhauser, Christine Fritz, Cédric Poyet, Ulrich Wagner, Levi A. Garraway, Peter J. Wild, Jean-Philippe Theurillat, Myles Brown. TRIM24 is an oncogenic transcriptional activator in prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1806.
Abstract 1808: Nuclear receptor, ESRRG functions as tumor suppressor by preventing Helicobacter pylori infection in gastric cancer
Authors: Kang Myoung-Hee , Lee Yong-Soo , Jung Jin-Hak , Park Young Soo , Jeong Jin-Yong , Kweon Mi-Na , Paik Chan-Ki , Myung Seung-Jae , Park Yun-Yong
Published: July 2016

Gastric cancer (GC) is the most common cancer type in Asia. Surgical resection and chemotherapy is standard treatment of GC. Molecular therapeutic targets and mechanism contributing to develop the gastric cancer is poorly understood. To discover the molecular target in gastric cancer, we analyzed gene expression data from gastric cancer patients. Data analysis reveals that nuclear receptor ESRRG is top candidate gene conferring the tumor suppressive property in gastric cancer. ESRRG expression is down-modulated by Helicobacter pylori (H. pylori) infection, which is one of causing factor triggering gastric cancer. In addition, ectopically over-expressed ESRRG protects the cell from H. pylori infection and suppressed cancer cell growth in vitro and in vivo mouse model. Gene expression profile reveals that TFF1, which is well known as tumor suppressor in gastric cancer is down-stream target of ESRRG and ESRRG well reflects clinical outcome in gastric cancer. Our study provides new molecular insights how ESRRG regulates gastric cancer cell growth induced by H. pylori and suggests possibility that ESSRRG can be therapeutic target for the treatment of gastric cancer patients.

Citation Format: Myoung-Hee Kang, Yong-Soo Lee, Jin-Hak Jung, Young Soo Park, Jin-Yong Jeong, Mi-Na Kweon, Chan-Ki Paik, Seung-Jae Myung, Yun-Yong Park. Nuclear receptor, ESRRG functions as tumor suppressor by preventing Helicobacter pylori infection in gastric cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1808.
Abstract 1809: Endothelin causes transactivation of the EGFR and Her-2/Neu in non-small cell lung cancer cells
Authors: Moody Terry W. , Ramos Alvarez Irene , Mantey Samuel , Jensen Robert T.
Published: July 2016

Endothelin-1 (ET-1) is a 23 amino acid peptide which is present in many lung cancer cell lines (Ahmed et al., Am. J. Respir. Cell Mol. Biol. 2000; 22: 422). Expression of ET-1 or the ETA receptor is associated with poor prognosis of non-small cell lung cancer (NSCLC) patients (Boldrini et al., Eur. J. Cancer 2005; 41: 2828). ET-1 stimulates whereas the ETA antagonist ZD4054 inhibits the proliferation of cancer cells (Bagnato et al., Br. J. Pharmacol. 2011: 163; 220). Here the effects of ET-1 on the transactivation of the EGFR and Her-2/Neu were investigated in NSCLC cells. Twelve of 13 NSCLC cell lines tested had both ETA and ETB mRNA. Addition of ET-1 to adenocarcinoma cell line NCI-H838, which had both ETA and ETB mRNA, elevated cytosolic Ca2+, increased tyrosine phosphorylation of the ERK, EGFR and Her-2/Neu and increased colony number. The ability of ET-1 to increase cytosolic Ca2+, tyrosine phosphorylation of ERK, EGFR and Her-2/Neu and colony number was inhibited by ZD4054 or BQ123 (ETA antagonists). The transactivation of EGFR and Her-2/Neu by ET-1 was inhibited by lapatinib (EGFR and Her-2/Neu TKI), GM6001 (matrix metalloprotease inhibitor), PP2 (Src inhibitor) or N-acetylcysteine (anti-oxidant). ET-1 stimulated whereas lapatinib and ZD4054 inhibited the clonal growth of NSCLC cells. The results indicate that ET-1 may regulate the proliferation of NSCLC cells in an EGFR and Her-2/Neu-dependent manner.

Citation Format: Terry W. Moody, Irene Ramos Alvarez, Samuel Mantey, Robert T. Jensen. Endothelin causes transactivation of the EGFR and Her-2/Neu in non-small cell lung cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1809.
Abstract 1828: Analysis of putative serum biomarker candidates for cardiotoxicity prediction in a cohort of early stage breast cancer patients treated with docetaxel, cyclophosphamide, and bevacizumab
Authors: Murray David W. , O’Farrell Alice C. , Gullo Giuseppe , Maratha Ashwini , Dicker Patrick , Rafferty Mairin , Murphy Verena , Ky Bonnie , Gallagher William M. , O’Donovan Norma , Crown John , Byrne Annette T.
Published: July 2016

Background: Breast cancer is the leading cause of cancer in women, with 1.8 million patients diagnosed globally each year. In early stage disease, anthracycline regimens are commonly employed, but the risk of cardiotoxicity is high. Replacing anthracyclines with taxanes in combination with cyclophosphamide may improve response [1] and Bevacizumab (BVZ) may provide addtional benefit. However, as BVZ can cause hypertension and increased risk of cardiac failure, a rationale exists for studying biomarkers of cardiotoxicity in this setting.

Methods: We assessed a panel of biomarkers in n = 57 early stage breast cancer patients whom underwent docetaxel/cyclophosphamide/BVZ therapy on the TC-Avastin Trial (NCT00911716). Serum was collected prior to treatment and at 3 months. Cardiotoxicity was defined as decline in left ventricular ejection fraction ≥ 10% to <55% over 1 year treatment + 2 year follow-up. Cardiac troponin I and T [cTnI, cTnT], Endothelin-1, Galectin-3, Placental Growth Factor [PlGF], Platelet Derived Growth Factor, Vascular Endothelial Growth Factor receptors 1 and 2, total nitrous oxide NO and myeloperoxidase [MPO]) were screened by ELISA in a subgroup (n = 16). Pilot and recently published findings [2] implicated PlGF, cTn1 and MPO, which were subsequently assessed in the larger cohort.

Results: Changes in serum MPO or cTnI were not related to cardiotoxicity when analysed in the larger cohort. However, a “cut-off” PlGF fold change of 0.99 could differentiate patients who developed cardiotoxicity from those that did not. 74% of cardiotoxicity patients, compared to only 35% of non-cardiotoxicity fell below the 0.99 cut-off. Conversely, 65% of non-cardiotoxicity patients, compared to 25% of cardiotoxicity patients fell above the cut-off (p = 0.0042).

Conclusions: Our data suggest that decreased PlGF at 3 months may predict cardiotoxicity. These observations require further validation.

This work was funded by AngioTox, an EC FP7 IAPP Marie Curie Award (Grant Agreement 251528). [1]Jones et al., J. Clin. Oncol.27, 1177-1183 (2009); [2] Putt et al., Clin. Chem 61, 1164-1172 (2015).

Citation Format: David W. Murray, Alice C. O’Farrell, Giuseppe Gullo, Ashwini Maratha, Patrick Dicker, Mairin Rafferty, Verena Murphy, Bonnie Ky, William M. Gallagher, Norma O’Donovan, John Crown, Annette T. Byrne. Analysis of putative serum biomarker candidates for cardiotoxicity prediction in a cohort of early stage breast cancer patients treated with docetaxel, cyclophosphamide, and bevacizumab. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1828.
Abstract 1831: Cancer genomic resources and needs in the Latin American region
Authors: Oliver Javier , Perdomo Sandra , Vaca Felipe
Published: July 2016

In 2012, approximately 1,005,255 new cancer cases and 550,164 deaths from cancer occurred in both sexes in central and South American region, by 2025 and increment of nearly 30% of new cases and 35% of deaths from cancer are predicted. One of the priorities of cancer control within the region is to reduce avoidable deaths from cancer by improving early detection and personalized treatment. The development of cancer genomics and their integration to cancer care has shown a great improvement in cancer control worldwide; however, all these key advances have been mainly concentrated in highly developed nations and little is known about the capacities and needs of cancer genomics in the Latin-American (LA) context.

In order to evaluate the capacity and development of cancer genomic applications in the LA we collected available information for all countries in central, South America and Cuba. Data reviewed included: number of cancer research institutions, number of NGS platforms, research groups working in cancer genetics, publications on cancer genetics for the last 10 years, educational programs on genomics and related health policies.

In average 15 Research groups per country were registered as conducting cancer-genetics related projects. In the las 10 years, 206 publications on cancer genetics including reviews were led by (1st, 2nd or corresponding) authors affiliated to LA institutions and only publications in the last 2 years included cancer genomic analysis. Most publications related to breast, gastric and hematological cancers were underrepresented in general. In the past 5 years physical resources have grown markedly by the acquisition of nearly 150 NGS platforms in 9 countries, the majority installed in medical related services and universities. Participation in large genomic consortia has only included two countries Mexico (as partner) and Brazil (as leading). Educational programs in genomics are scarce, almost exclusive of graduate programs and few are applied to cancer.

Despite the recent advances in introducing cancer genomics knowledge and application in LA, the region lacks development of integrated genomic research projects, improved use of platforms dedicated to cancer, educational programs and health policies that focus on the most frequent cancers and could impact cancer care.

Citation Format: Javier Oliver, Sandra Perdomo, Felipe Vaca. Cancer genomic resources and needs in the Latin American region. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1831.
Abstract 1839: Analysis of cell cycle-related gene expressions in MDM2-transfected LNCaP-MST cells after inhibition of HSP90
Authors: Venkatesan Thiagarajan , Alaseem Ali , Alhazzani Khalid , Dondapati Priya , Alobid Saad , Rathinavelu Appu
Published: July 2016

MDM2 gene is a cellular proto-oncogene amplified in 25-40% of all human cancers and regulates cell proliferation, senescence, and apoptosis through targeting p53. It is widely speculated that MDM2 protein is not only responsible for tumorigenesis via p53 inactivation, but also for increasing the metastatic ability of originally non-metastatic tumor cells. HSP90 is a molecular chaperone that regulates the maturation, activation and stability of critical signaling proteins that drive the development and progression of prostate cancer. 17-Allyamino-17-demethoxygeldanamycin (17-AAG) is an inhibitor of HSP90, which causes proteasomal degradation of client proteins such as AR, HER2, Akt etc., leading to cell cycle arrest and substantial growth inhibition. HSP90 is also believed to be involved in the stabilization of MDM2 and therefore, inhibition of HSP90 is expected to accelerate degradation of MDM2 and disrupt its function. Hence, our study was designed to determine the expression profile of the genes in the cell cycle pathway following treatment of MDM2 overexpressing LNCaP-MST prostate cancer cells with 17-AAG (10 μM at 24 hrs). In the present study, Human cell cycle PCR array was used to examine the alterations in gene expression pattern following exposure to the 17-AAG in MDM2 overexpressed prostate cancer cells. In our experiments we observed that the genes impacted by MDM2 overexpression were reversed by HSP90 inhibition. The expression of genes such as CDC25A, CDC25C, AURKB and Survivin levels were significantly down-regulated after 17-AAG treatment, while p21 and GADD45A were up-regulated compared to the control LNCaP-MST. A heat map and scatterplot analysis clearly confirmed the alterations in the expression levels of these genes following 17-AAG treatment. It is expected that more thorough understanding of the consequences of HSP90 blockade, particularly those that are independent of its role as a regulator of p53, will reveal its therapeutic significance. Our results offer convincing evidence to suggest that the inhibition of HSP90 can induce cell cycle arrest in LNCaP-MST cells. (The financial support from the Royal Dames of Cancer Research Inc., Ft. Lauderdale, Florida is gratefully acknowledged).

Citation Format: Thiagarajan Venkatesan, Ali Alaseem, Khalid Alhazzani, Priya Dondapati, Saad Alobid, Appu Rathinavelu. Analysis of cell cycle-related gene expressions in MDM2-transfected LNCaP-MST cells after inhibition of HSP90. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1839.
Abstract 1845: Development of a quantitative targeted RNA-Seq methodology for use in differential gene expression analysis
Authors: Lader Eric , Hussong Melanie , Fosbrink Matthew
Published: July 2016

RNA Sequencing (RNA-Seq) uses the capabilities of Next Generation high-throughput sequencing (NGS) methods to provide insight into the transcriptome of a cell as it generates millions of reads. Whole transcriptome sequencing can be used to quantify gene expression on a transcriptome-wide scale, identify splice variants, quantify allele specific expression, and characterize fusion transcripts. Development of a highly reproducible and sensitive targeted quantitative sequencing method would aid in facilitating a deeper understanding and characterization of the roles of a specific set of genes, while enabling much higher sample throughput and significant cost savings relative to whole-transcriptome sequencing. In this study, we report a targeted RNA-Seq technology, QIAseq RNA, which makes use of several methodologies to deliver an extremely flexible, highly precise tool for characterizing gene expression. QIASeq RNA incorporates 12-base random molecular barcodes into each unique target strand which benefits quantifying gene expression in a given multiplexed sample. Counting the number of molecular tags allows one to determine the sequence coverage per target and adjust experimental conditions to use the read budget of any sequencing platform most efficiently. Using either the Illumina or Ion Torrent platforms, users can choose to multiplex up to 96 RNA samples from 12 to 1000-plex expression panels. No mRNA selection or rRNA removal or blocking is required. The entire protocol, from cDNA synthesis to finished library, which is ready for sequencing, can be accomplished in under one day. Custom assays for a specific target site can add the ability to distinguish between isoforms or identify allele specific expression. We explore the capabilities of this system by profiling large numbers of genes in a cell model system's response to small molecule treatment. Changes in gene expression by these treatments were measured by targeted RNA NGS, and fold-changes in gene expression due to chemical perturbation were characterized. Complex gene relationships in perturbed pathways were mapped using QIAGEN's Ingenuity Pathway Analysis (IPA) tool. The IPA tool also facilitated the elucidation of the impact of gene expression changes in the context of biological processes, molecular interactions, cellular phenotypes and disease. This article will provide application data for GRRC including a discussion of technical challenges faced when profiling large numbers of genes in a large cohort.

Citation Format: Eric Lader, Melanie Hussong, Matthew Fosbrink. Development of a quantitative targeted RNA-Seq methodology for use in differential gene expression analysis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1845.
Abstract 1857: Dysregulation of transsulfuration enzymes contribute to malignant transformation in a murine model of colitis-associated carcinogenesis
Authors: Johnson Paul , Phillips Ches’Que M. , Grim Carl , Zatarain John R. , Gajjar Aakash H. , Qiu Suimin , Wang Rui , Chao Celia , Pinchuk Iryna V. , Hellmich Mark R.
Published: July 2016

Introduction: Ulcerative colitis (UC) is a highly morbid, chronic inflammatory disease characterized by mucosal ulceration of the colonic mucosa and is associated with the higher risk of colitis-associated cancer (CAC). The exact mechanism(s) causing UC progression to CAC is currently unknown. The balanced activity of transsulfuration pathway enzymes, cystathionine-gamma-lyase (CSE) and cystathionine-beta-synthase (CBS), and their production of endogenous hydrogen sulfide gas, are critical to maintenance of the colonic homeostasis. CSE activity is suggested to be important for wound healing and mucosal protection. Consistent with this, we have previously showed a decrease CSE expression in human colonic mucosa obtained from patients with chronic UC, compared to normal colonic mucosa by immunocytochemistry. By contrast, increased CBS expression is implicated in the progression of sporadic colorectal carcinoma. However, the role of these enzymes in CAC is unknown. We hypothesize that the dysregulation in CSE/CBS expression and activity is important to the progression from UC to CAC.

Methods: CSE null mice and wild type Sv129/B6 (control) mice were used in azoxymethane-dextran sodium sulfate (AOM-DSS) colon cancer model which mimics human CAC. The disease development was followed up to day 80. Confocal microscopy and Western blot was used to assess the gene expression during cancer development. Size, number, and time interval to tumor formation, as well inflammation were assessed.

Results:

Abrogation of CSE expression using CSE null animals in AOM-DSS model of CAC accelerated the time to tumor development and resulted in an increase in both tumor size (p<0.001) and number (p<0.001) compared to wild-type controls. CBS protein expression was increased within the colonic tumor when compared to the normal margin in AOM-DSS treated animals by Western blot analysis and tissue immunostaining. Interestingly, CAC liver metastases, an exceedingly rare finding in this mouse model, were identified.

Conclusion: Taken together, our human and murine data suggest that dysregulation in the transsulfuration pathway enzymes CSE and CBS expression/activity may be critical contributor to the CAC development in UC and may serve as a potential biomarker for disease progression in the future.

Citation Format: Paul Johnson, Ches’Que M. Phillips, Carl Grim, John R. Zatarain, Aakash H. Gajjar, Suimin Qiu, Rui Wang, Celia Chao, Iryna V. Pinchuk, Mark R. Hellmich. Dysregulation of transsulfuration enzymes contribute to malignant transformation in a murine model of colitis-associated carcinogenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1857.
Abstract 1858: Analyzing the impact of MDM2 overexpression on cell cycle pathway-related genes using polymerase chain reaction array
Authors: Dondapati Priya , Venkatesan Thiagarajan , Rathinavelu Appu
Published: July 2016

The human homologue of the mouse double minute 2 (MDM2) oncogene has been well characterized as a negative regulator of the tumor suppressor p53. It has been established that p53 regulates genes that are involved in cell cycle arrest, apoptosis, DNA repair and senescence. MDM2 interacts primarily with p53 in an autoregulatory negative feedback loop and targets it for ubiquitylation and proteasomal degradation there by attenuating p53 cell cycle arrest and apoptosis functions. Studies have shown that MDM2 also exerts its effects via p53 independent pathways. These p53-independent functions of MDM2 may have a role in cancer etiology and progression, yet the exact function and significance of these interactions are not fully understood. MDM2 is overexpressed in more than 40 different types of malignancies, including wide varieties of carcinomas and sarcomas, which is believed to be associated with worse clinical prognosis and increased likelihood of distant metastases. Hence, MDM2 is proving to be a key player in human cancer and thus an important target for therapeutic intervention. In this study, we aimed to investigate the effects of MDM2 overexpression on cell cycle pathways by using LNCaP prostate cancer cells and MDM2 transfected LNCaP-MST. The LNCaP-MST cells expressed at least 10-folds higher levels of MDM2 mRNA due to transfection. We used the Human Cell Cycle PCR array to compare the gene expression profiles of LNCaP and LNCaP-MST cells. Our experimental results clearly indicated that MDM2 modulates the expression of cell cycle related genes. The following genes were found to be significantly down-regulated in LNCaP-MST cells compared to LNCaP: CCND2, GADD45A, RB1, RBBP8, CDKN1A and TP53. Genes such as BIRC5, CDK1 were found to be significantly up-regulated. Heat map and scatter plot analysis further confirmed the alterations in the expression levels of these genes following the MDM2 overexpression. In addition, cluster analysis further denoted that the gene expression patterns positively correlated with cell proliferation and tumor growth. Our experimental results offer conclusive evidence that MDM2 can impact the cancer growth and metastatic potential by altering the gene expressions in the cell cycle pathway. (The financial support from the Royal Dames of Cancer Research Inc., Ft. Lauderdale is gratefully acknowledged).

Citation Format: Priya Dondapati, Thiagarajan Venkatesan, Appu Rathinavelu. Analyzing the impact of MDM2 overexpression on cell cycle pathway-related genes using polymerase chain reaction array. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1858.
Abstract 186: Functional characterization of ERK3 mutants existing in human cancers
Authors: Alsaran Hadel , Elkhadragy Lobna , Long Weiwen
Published: July 2016

Extracellular signal-Regulated Kinase 3 (ERK3) is an atypical member of the mitogen-activated protein kinase (MAPK) family. Recent studies have shown that ERK3 is highly upregulated in multiple cancers, such as lung cancer and colon cancer, and plays an important role in promoting cancer migration and invasion. While the link between ERK3 and cancers has been recognized, little is known about ERK3 mutations in cancer progression. In this study, we have investigated ERK3 mutations on R64 and L290 that have been found in cancers of lung, large intestine and skin (COSMIC database). Importantly, mutations of R64C, R64H, L290P and L290V all are located in the kinase domain of ERK3. In order to characterize these mutations, we generated ERK3 mutants with each of these point mutations, and then overexpressed them in HeLa cells with or without stable knockdown of endogenous ERK3. Notably, we found that in comparison with wild type ERK3, all of these cancer-related mutations lead to reduction of phosphorylation at S189 within the activation loop (an indicator of ERK3 kinase activity). While activation loop phosphorylation is well known to result in activation of the classical MAPKs, its effects on the activity and functions of ERK3 remain elusive. To study the functional impact of these ERK3 mutations on cancer cell invasiveness, we performed trans-well migration assays using HeLa cells overexpressing each of these mutants or the wild type ERK3. In line with previous finding, overexpression of ERK3 increased cell migration. Interestingly, both L290P and L290V mutations significantly increased ERK3's ability in promoting cell migration, whereas R64C and R64H mutations resulted in a decrease in cell migration. These results suggest that S189 phosphorylation within the activation loop does not correlate with ERK3's function in promoting cancer cell motility. To conclude, we report for the first time the functional characterization of cancer-related ERK3 mutations in modulating cancer cell migratory properties.

Citation Format: Hadel Alsaran, Lobna Elkhadragy, Weiwen Long. Functional characterization of ERK3 mutants existing in human cancers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 186.
Abstract 1867: Activation of RHO-DIAPH signaling impairs RAS-driven exosome biogenesis
Authors: Davis-Turner Julie , Howard Andrew M. , Alberts Arthur S.
Published: July 2016

Like oncogenic RAS, the RAS homologues RHOA and RHOB are conventionally thought to support the malignant platform by governing cell structure remodeling by effectors that include the Diaphanous (DIAPH) family formins. RHO-activated DIAPHs generate linear actin filaments (F-actin) that physically support membrane protrusions at the leading edge of migrating cells and the trafficking of growth factor receptors, which are processes that may be required for malignant progression. However, our cellular, genetic, and pharmacological studies argue that both RHO and DIAPHs function as tumor suppressors whose loss of function triggers or augments malignant progression. DIAPH1 and DIAPH3 inhibition, for example, reduces cortical F-actin sufficiently to cause non-apoptotic blebbing. Combined with oncogenic RAS, DIAPH inhibition and blebbing accelerate exosome release. These exosomes have the capacity to foment oncogenesis by seasoning cells of the tumor microenvironment in ways that promote inflammation, survival, immune escape, and metastasis. We have identified a mechanism whereby RAS uses its own guanine-nucleotide releasing protein, SOS, to inhibit DIAPHs and facilitate its own escape within exosomes. Consistent with this idea, pharmacological activation of RHO and DIAPH signaling inhibits tumorigenesis and invasion. RHO and DIAPH agonist/activation strategies may prove to be an effective alternative for impairing RAS-driven oncogenesis where conventional inhibitors of RAS-MAPK signaling have failed or have been met by poorly understood resistance mechanisms.

Citation Format: Julie Davis-Turner, Andrew M. Howard, Arthur S. Alberts. Activation of RHO-DIAPH signaling impairs RAS-driven exosome biogenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1867.
Abstract 187: ASN0007, a potent ERK1/2 inhibitor with strong antitumor activity in multiple RAS mutant models
Authors: Reddy Sanjeeva P. , Sivanandhan Dhanalakshmi , Kethiri Raghava Reddy , Nair Sreekala , Dewang Purushottam , Zainuddin Mohd , Vaithilingam Krishnakumar , Smith Roger A. , Gupta Sandeep , Thompson Scott K.
Published: July 2016

The extracellular-signal-regulated kinases, ERK1 and ERK2 (ERK1/2), play a critical role in the RAS/RAF/MEK signaling pathway which is quite frequently activated by mutations in upstream targets such as BRAF, RAS and receptor tyrosine kinases. Most of the resistance mechanisms to BRAF and MEK inhibitors ultimately lead to increase in phosphorylation of ERK1/2 suggesting the importance of this node in the RAS/RAF/MEK pathway even in the resistance setting. Therefore, inhibition of ERK1/2 offers a promising strategy to address both innate and acquired resistance to BRAF and MEK inhibitors in various solid tumors. In order to inhibit this critical node of the RAS/RAF/MEK pathway, we discovered and characterized ASN007, a novel compound that shows potent inhibition of ERK1/2 kinases in a biochemical assay with low single-digit IC50 values. In mechanistic cell-based assays, ASN007 inhibited the phosphorylation of ERK1/2 substrates such as pRSK1, FRA1 and Elk1 in various cell lines. As expected, the compound did not have any effect on the phosphorylation of ERK1/2. ASN007 showed strong anti-proliferative activity in both BRAF and RAS mutant cell lines. Importantly, when compared to other ERK1/2 inhibitors such as BVD-523 and GDC-0994, ASN007 showed potent antiproliferative activity in a much broader panel of KRAS, NRAS and HRAS mutant cell lines representing various histological tumor types. The compound also caused cell cycle arrest at the G0-G1 phase and induced apoptosis in both BRAF and RAS mutant cell lines. In a daily oral dosing regimen, ASN007 demonstrated strong inhibition of tumor growth in multiple BRAF and KRAS mutant xenograft models in mice and was well tolerated at efficacious doses. ASN007 has a good pharmacokinetic profile in pre-clinical species and does not exhibit significant inhibition of CYP enzymes or hERG channel. Based on its profile in preclinical studies, ASN007 is expected to show strong efficacy in BRAF and RAS mutant cancers. ASN007 is currently in preclinical development.

Citation Format: Sanjeeva P. Reddy, Dhanalakshmi Sivanandhan, Raghava Reddy Kethiri, Sreekala Nair, Purushottam Dewang, Mohd Zainuddin, Krishnakumar Vaithilingam, Roger A. Smith, Sandeep Gupta, Scott K. Thompson. ASN0007, a potent ERK1/2 inhibitor with strong antitumor activity in multiple RAS mutant models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 187.
Abstract 1875: Identification of LASEP3 as a serological and tissue biomarker and a therapeutic target for lung cancer
Authors: Takano Atsushi , Nakamura Yusuke , Daigo Yataro
Published: July 2016

Lung cancer is the leading cause of cancer deaths worldwide. The prognosis of patients with lung cancer remains poor, because of its occult metastasis even at earlier stage of the disease and its resistance to conventional multimodal treatments. Therefore, this study aims to identify novel serum and tissue biomarkers for personalized therapy and/or therapeutic targets for lung cancers through screening and functional analyses of cancer-specific oncoproteins. We used cDNA microarrays for screening genes encoding transmembrane/secretory proteins that are up-regulated in lung cancers, and identified a secreted protein, LASEP3 (lung cancer-associated serum protein 3) as a candidate. Immunohistochemical staining of LASEP3 showed that LASEP3 expression was observed in 198 (54.8%) of 361 NSCLCs (non-small cell lung cancer) that had undergone curative surgery. High level of LASEP3 expression was associated with poor prognosis for NSCLC patients. (P = 0.0183 by log-rank test). Serum LASEP3 levels were higher in NSCLC patients than in healthy volunteers. The proportion of serum LASEP3-positive cases was 160 (61.8%) of 259 NSCLCs (49.4% for stage I-II, 67.4% for stage III-IV), while 6 (5.5%) of 109 healthy volunteers were falsely diagnosed. Moreover, reduction of LASEP3 expression by siRNAs suppressed lung cancer cell proliferation, probably through apoptosis as suggested by flow cytometric analysis and caspase-3 and 7 assays. Furthermore, subsequent microarray analysis of these cancer cells identified several candidate downstream genes of LASEP3 that relate to cell growth and invasion signals. LASEP3 is a possible diagnostic and prognostic biomarker and a therapeutic target for lung cancer.

Citation Format: Atsushi Takano, Yusuke Nakamura, Yataro Daigo. Identification of LASEP3 as a serological and tissue biomarker and a therapeutic target for lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1875.