bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2289: Precision-weighted RNA-seq analyses of molecular abundance (RoMA) for detecting differential gene expression
Authors: Cai Guoshuai , Franks Jennifer M. , Whitfield Michael L.
Published: August 2018

In this study, we propose a novel method, RoMA, to accurately detect differential expression and unlock the integration with numerous upstream and downstream analyses on mRNA abundance in RNA-seq studies. Various methods have been proposed, each with its own limitations. Some naive normal-based tests have low testing power with the invalid normal distribution assumptions for RNA-seq read counts, whereas count-based methods lack a biologically meaningful interpretation and have limited capability for integration with other analysis packages for mRNA abundance. RoMA incorporates information from both mRNA abundance and raw counts by modeling RPKM (reads per kilobase per million), which represents the relative abundance of mRNA transcripts, and borrowing mean-variance dependency from CPM (counts per million) as a precision weight accounting for the variability in sequencing depth. Studies on simulated data and two real datasets showed that RoMA provides an accurate quantification of mRNA abundance and a value adjustment-tolerant DE analysis with high AUC, low FDR and a desirable type I error rate. This study provides a valid strategy for mRNA abundance modeling and data analysis integration for RNA-seq studies, which will greatly facilitate the identification and interpretation of DE genes. The methods is implemented in a user-friendly R package (RoMA).

Citation Format: Guoshuai Cai, Jennifer M. Franks, Michael L. Whitfield. Precision-weighted RNA-seq analyses of molecular abundance (RoMA) for detecting differential gene expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2289.
Abstract 2293: Outstanding chromosomal expression patterning in human cancers
Authors: Barnes Spencer , Song Mingzhou
Published: August 2018

Cancer transcriptome is an effect of abnormal genetic and/or epigenetic regulation. In the past, transcriptome was studied at the level of individual transcripts for their differential expression. Spatial gene expression patterns along chromosomes were not yet characterized for cancer. We hypothesized that gene expression patterning is not randomly distributed along the chromosomes in cancer versus normal tissues. We analyzed cancer transcriptome from the National Cancer Institute Genomic Data Commons, and identified genomic regions with significant shifts in expression from normal tissues to tumors. We used a novel highly efficient method in the Ckmeans.1d.dp package for clustering genes by chromosomal location and expression level. We identified two chromosomal zones of note that exhibit similar expression patterns across several cancer types. In breast, lung, liver, and esophageal cancers, a zone near 6p22.2 contains genes that are similarly highly upregulated in tumor tissues compared to matched normal tissues. This zone contains a group of histone cluster 1 genes, among which are 14 known cancer genes. In colorectal, lung, liver, and esophageal cancers, a zone on 14q32.33 exhibits either heavy up- or downregulation, dependent on the specific cancer. This zone is rich in immunoglobulin heavy variable genes. Table 1 lists genomic zones of note specific to six cancer types. Our study provides an atlas of outstanding chromosomal expression patterning that can be used as markers for cancer in general and also for specific cancer types.

Table 1. Outstanding genomic zones specific to six human cancer types.Cancer type Chr:start-end (Mb) Cytogenetic band # Matched tumor/normal pairsBreastchr2:88-92p11.2112Colorectalchr7:141-146q3541Esophagealchr11:7-12p15.48Kidneychr12:85-91q21.3323Liverchr8:15-22p2250Lungchr8:121-126q24.1357

Citation Format: Spencer Barnes, Mingzhou Song. Outstanding chromosomal expression patterning in human cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2293.
Abstract 23: Chemotherapy and tau interplay facilitates breast-to-brain metastasis
Authors: Saatian Behnaz , Deshpande Krutika , Julian Alex , Nakamura Brooke Naomi , Shao Ling , Neman Josh
Published: August 2018

Breast cancer represents the second most common cause of brain metastasis with the rate of 10-16% after lung cancer. The overall survival in brain metastatic patients is very short ranging from 2 to 25 months. Considering the high incidence of brain metastasis and the limitation of available therapies, developing preventive strategies and early detection methods are required to improve the survival rate in the patients. Many primary and brain metastatic breast cancer patients represent cognitive deficits. Besides, chemo-treated breast cancer patients show memory problems before brain metastases diagnosis. Although the cause of cognitive impairments after chemotherapy have not been explored, a recent study revealed that chemotherapy promotes breast cancer metastasis. Correspondingly, our mRNA expression analysis showed downregulation of the tight junction markers in chemo-treated choroid plexus cells (CPs), suggesting that chemotherapy is likely to cause an increase in the blood-cerebral spinal fluid barrier (BCSFB) permeability. Chemotherapy also induces upregulation of tau expression in CP cells. Tau is known as a pathological hallmark of Alzheimer's disease (AD) and neurodegenerative disorders. The abnormally phosphorylated and aggregated form of tau play critical role in neurodegenerative diseases as of AD. Tau expression, both at the mRNA and protein level, is significantly elevated in breast to brain metastases (BBMs). Therefore, we hypothesized that chemotherapy upregulates tau expression in breast cancer cells leading to the BCSFB and blood-brain barrier (BBB) permeability, and consequently, breast to brain metastasis and neurodegeneration. To test our hypothesis, the effect of chemotherapy on the BCSFB and BBB permeability were investigated using the trans endothelial electrical resistance (TEER) and migration assays in the in vitro models. Our results show an increase in abnormal tau expression in chemo-treated primary breast cancer cells. BBM-derived tau is abnormal and forms paired helical filaments (PHFs), similar to Alzheimer's. We conclude that breast to brain metastasis is facilitated through an interplay between chemotherapy, and tau expression and release from breast cancer cells. Additionally, we suggest that tau plays a key role in neurodegeneration defects in brain metastatic patients. This study opens a new path towards prevention and early detection of breast cancer metastasis to the brain, cognitive decline and neurodegeneration in metastatic patients.

Citation Format: Behnaz Saatian, Krutika Deshpande, Alex Julian, Brooke Naomi Nakamura, Ling Shao, Josh Neman. Chemotherapy and tau interplay facilitates breast-to-brain metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 23.
Abstract 2301: Dual targeting of CDK4/6 and MET in metastatic uveal melanoma
Authors: Ohara Masahiro , Saito Kengo , Kageyama Ken , Terai Mizue , Cheng Hanyin , Aplin Andrew E. , Sato Takami
Published: August 2018

Uveal melanoma (UM) is the most common cancer of the eye in adults. Up to 50% of UM patients subsequently develop metastases, especially in the liver, for which no effective treatment has been identified. It has been reported that the retinoblastoma (RB) pathway is deregulated in more than 90% of UM despite the rarity of mutations in the RB1 gene itself. Cyclin D1 over-expression and inactivation of p16INK4a in UM are frequently present and make the RB protein constitutively hyperphosphorylated and functionally inactivated; therefore, CDK4/6 inhibition (CDK4/6i) is a rational strategy for treatment of UM. In this report, we investigated the antiproliferative activity of a selective CDK4/6 inhibitor on metastatic UM. A CDK4/6 inhibitor induced G0-G1 arrest in all three metastatic UM cell lines and suppressed their growth in in vitro and in vivo experiments. Interestingly, hepatocyte growth factor (HGF) decreased the effect of CDK4/6 inhibitor on metastatic UM cell lines and protected the metastatic UM cells from CDK4/6 inhibitor-induced cellular senescence. When CDK4/6i was combined with MET inhibitor, enhanced growth suppression was observed in metastatic UM tumors grown in human-HGF knock-in xenograft mouse models. HGF is enriched in the liver and the majority of liver metastases from UM express activated forms of MET; therefore, signaling through MET could contribute to the resistance mechanisms against CDK4/6i, especially in UM patients with hepatic metastasis. Together, these results provide a rationale for the use of MET inhibitor in combination with a CDK4/6 inhibitor for the treatment of metastatic UM.

Citation Format: Masahiro Ohara, Kengo Saito, Ken Kageyama, Mizue Terai, Hanyin Cheng, Andrew E. Aplin, Takami Sato. Dual targeting of CDK4/6 and MET in metastatic uveal melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2301.
Abstract 2308: Salicylic acid metabolites and derivatives inhibit CDK activity: Novel insights into aspirin's chemopreventive effects against colorectal cancer
Authors: Sankaranarayanan Ranjini , Dachineni Rakesh , Kumar D. Ramesh , Callegari Eduardo , Kesharwani Siddharth S. , Seefeldt Teresa , Tummala Hemachand , Gunaje Jayarama B.
Published: August 2018

Aspirin's potential as a drug continues to be evaluated for the prevention of colorectal cancer (CRC). Although multiple targets for aspirin and its metabolite, salicylic acid, have been identified, no unifying mechanism has been proposed to clearly explain its chemopreventive effects. Our goal here was to investigate the ability of salicylic acid metabolites, known to be generated through cytochrome P450 (CYP450) enzymes, and also its derivatives as cyclin dependent kinase (CDK) inhibitors to gain new insights into aspirin's chemopreventive actions. Using in vitro kinase assays, for the first time, we demonstrate that salicylic acid metabolites, 2,3-dihydroxybenzoic acid (2,3-DHBA) and 2,5-dihydroxybenzoic acid (2,5-DHBA), as well as derivatives 2,4-dihydroxybenzoic acid (2,4-DHBA), 2,6-dihydroxybenzoic acid (2,6-DHBA), inhibited CDK1 enzyme activity. 2,3-DHBA and 2,6-DHBA did not inhibit CDK2 and 4; however, both inhibited CDK-6 activity. Interestingly, another derivative, 2,4,6-trihydroxybenzoic acid (2,4,6-THBA) was highly effective in inhibiting CDK1, 2, 4 and 6 activity. Molecular docking studies showed that these compounds potentially interact with CDK1. Immunoblotting experiments showed that aspirin acetylated CDK1, and preincubation with salicylic acid and its derivatives prevented aspirin-mediated CDK1 acetylation, which supported the data obtained from molecular docking studies. We suggest that intracellularly generated salicylic acid metabolites through CYP450 enzymes within the colonic epithelial cells, or the salicylic acid metabolites generated by gut microflora may significantly contribute to the preferential chemopreventive effect of aspirin against CRC through inhibition of CDKs. This novel hypothesis and mechanism of action in aspirin's chemopreventive effects opens a new area for future research. In addition, structural modification to salicylic acid derivatives may prove useful in the development of novel CDK inhibitors in cancer prevention and treatment.

Citation Format: Ranjini Sankaranarayanan, Rakesh Dachineni, D. Ramesh Kumar, Eduardo Callegari, Siddharth S. Kesharwani, Teresa Seefeldt, Hemachand Tummala, Jayarama B. Gunaje. Salicylic acid metabolites and derivatives inhibit CDK activity: Novel insights into aspirin's chemopreventive effects against colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2308.
Abstract 231: Identification of susceptible loci and enriched pathways for post-radiotherapy pain in breast cancer patients
Authors: Lee Eunkyung , Takita Christina , Wright Jean L. , Martin Eden , Slifer Susan , Hu Jennifer
Published: August 2018

Pain is one of the most prevalent symptoms among breast cancer survivors, which negatively impact on their quality of life (QOL). Adjuvant radiotherapy (RT) can increase the risk of development or persistence of pain; however, little is known about the mechanisms of RT-related pain or genetic susceptibility. The current study aimed to identify susceptible loci and enriched pathways for clinically relevant pain, which is defined as pain score ≥ 4, at the completion of RT (post-RT pain). We conducted a genome-wide association study (GWAS) of 1,112 breast cancer patients with 1,344,832 tagging SNPs, gene-based analysis using PLINK set-based tests for 19,621 genes, and functional enrichment analysis of a gene list (875 genes having p < 0.05) using NIH DAVID functional annotation module with KEGG pathways and GO terms (n=380). About 29% of patients reported post-RT pain, and 4 SNPs showed suggestive associations; rs16970540 in RFFL or near to LIG3 gene (p=1.7x10-6), rs4584690 and rs7335912 in ABCC4/MPR4 gene (p=5.5x10-6 and p=7.8x10-6, respectively), and rs73633565 in EGFL6 gene (p=8.1x10-6). Gene-based analysis suggested the importance (involvement) of neurotransmitter, olfactory receptor, and cytochrome P450 in post-RT pain, and functional analysis suggested glucuronidation and olfactory receptor activity as the most enriched biological processes/features. With a limited size of sample, this is the first GWA study showing (confirming) that post-RT pain is a complex polygenic trait influenced by many biological processes and functions. If validated in a larger population, the results can provide biological targets for pain management to improve cancer survivor's QOL. Additionally, these findings may be used as predictive biomarkers of personalized pain management.

Citation Format: Eunkyung Lee, Christina Takita, Jean L. Wright, Eden Martin, Susan Slifer, Jennifer Hu. Identification of susceptible loci and enriched pathways for post-radiotherapy pain in breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 231.
Abstract 2311: Determinants of CDK4 target engagement by palbociclib, ribociclib and abemaciclib
Authors: Green Jennifer L. , Nomanbhoy Tyzoon
Published: August 2018

Palbociclib, ribociclib and abemaciclib compose a class of ATP-competitive CDK4 and CDK6 inhibitors that have demonstrated preclinical and clinical efficacy against various tumor types. While these inhibitors have favorably altered the landscape of cancer therapeutics, identification of patients likely to respond to CDK4/6 inhibition remains a challenge. Previously, we used our chemoproteomics platform (KiNativ), which measures competition of an ATP- or ADP-acyl-phosphate probe, to profile lysates generated from cells that were treated with palbociclib. We showed that palbociclib-CDK4 target engagement is cell line specific and that engagement correlates with sensitivity of the cell line to palbociclib-mediated growth inhibition. However, the circumstances that determine whether target engagement is achieved were not understood. In an effort to understand the molecular mechanisms that influence CDK4 target engagement, we used the same proteomics platform to interrogate the contribution of various known CDK regulators. We identified a family of proteins that prevents palbociclib target engagement upon overexpression and whose lack of expression correlates with palbociclib sensitivity. This interaction is specific for CDK4/6 as modulation of the drug's interaction with off-target kinases was not affected by overexpression of these proteins. Furthermore, we found that these proteins similarly prevent CDK4 target engagement by ribociclib and abemaciclib. Incorporation of the proteins identified here into a biomarker strategy should help to enrich for patients likely to respond to CDK4/6 inhibition therapy and expand the utility of these inhibitors.

Citation Format: Jennifer L. Green, Tyzoon Nomanbhoy. Determinants of CDK4 target engagement by palbociclib, ribociclib and abemaciclib [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2311.
Abstract 2317: Palbociclib monotherapy exhibits potent activity in cervical cancer cell lines
Authors: Dam Peter Van , Wouters An , Dirix Luc , Rolfo Christian
Published: August 2018

Background: Over the last decades, little progress has been made in the systemic treatment of patients with advanced or recurrent cervical cancer. We recently performed an silico analysis to identify potential driver pathways of cervical carcinogenesis and candidate targets for treatment.1 Expression2Kinases (E2K) analysis revealed a protein-protein interaction (PPI) network of 162 nodes (including 20 druggable kinases) consisting of 5 signaling modules associated with MYC signaling (module 1), cell cycle deregulation and cyclin signaling (module 2), TGFβ-signaling (module 3), a PI3K - MAPK signaling (module 4) and chromatine modeling (module 5).

Methods: The cervical cancer cell lines included in the study were CaSki, SiHa and HeLa. Cells were incubated for 24 or 72 hours with the PI3K pathway inhibitor BYL719 (Novartis), the multiple pathway inhibitor INC280 (inhibits cMET-dependent PI3K and RAS signaling, Novartis), or the CDK4/6 inhibitor PD-0332991 (palbociclib, Selleck Chemicals) in monotherapy. Sensitivity to drug treatment (0 - 10 μM) was investigated using the colorimetric sulforhodamine B (SRB) assay. IC50 values were calculated using WinNonlin Software.

Results: Incubation with BYL719 or INC280 for 24 or 72 hours in concentrations up to 10 μM was not able to establish a distinct cytotoxic effect in the three cervical cancer cell lines included in the study. Incubation with 10 μM BYL719 for 72h resulted in only 20% cell kill, while 10 μM INC280 for 72h caused around 30% cell kill in all three cell lines. However, incubation with palbociclib for 72 hours clearly induced a concentration-dependent cytotoxic effect, with IC50 values of 5.32 ± 0.03 μM, 7.69 ± 0.02 μM and 5.68 ± 0.12 μM for HeLa, SiHa and CaSki cells, respectively.

Conclusion: CDK4/6 inhibition seems effective in cervical cancer cell lines and should be further evaluated alone and in combination with chemotherapy for the treatment of advanced and recurrent cervical cancer.

References:

Van Dam P, van Dam PJ, Rolfo C, Giallombardo M, Van Berckelaer C, Trin XB, Altintas S, Huizing M, Papadimitriou K, Tjalma W, Van Laere S. In silico pathway analysis in cervical carcinoma reveals potential new targets for treatment. Oncotarget 2016;7(3):2780-2795

Citation Format: Peter Van Dam, An Wouters, Luc Dirix, Christian Rolfo. Palbociclib monotherapy exhibits potent activity in cervical cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2317.
Abstract 2322: Ferroptosis induced by erastin in RAS mutant ovarian cancer cells
Authors: Takenaka Motoki , Suzuki Noriko , Mori Minako , Aoki Hitomi , Hirayama Tasuku , Nagasawa Hideko , Morishige Ken-ichirou
Published: August 2018

Although apoptosis has been thought as the principal mechanism of programed cell death in chemotherapy, other forms of non-apoptotic death also has been noted. The oncogenic RAS-selective lethal small molecule erastin triggers a unique iron-dependent form of cell death termed ferroptosis. This mechanism is expected to induce eradication of chemotherapeutic (e.g. platinum) resistant cancer cells. In this study we demonstrate this new form of cell death in ovarian cancer cells. The clear cell epithelial ovarian cancer cell lines, TOV-21G (KRAS_G13C) and KOC-7C (RAS wild type) were used in this study. Erastin dose-dependently inhibited growth in TOV-21G cells, but not KOC-7C cells. Treatment of TOV-21G cells with erastin resulted in increase of ROS. Iron chelator deferoxamine (DFO) rescued growth inhibition induced by erastin in TOV-21G cells. Although cisplatin inhibited growth in TOV-21G cells, this inhibition was not rescued by DFO. The GSH level was reduced by erastin, but this reduction was not rescued by DFO. In addition, the basal GSH level was higher and the basal ROS level was lower in KOC-7C cells than those in TOV-21G cells. Erastin induced ferroptosis by reducing the GSH level in RAS mutant ovarian cancer cells. Although DFO rescued growth inhibition induced by erastin, the pathway was not on System Xc-. The sensitivity to erastin might depends on the basal level of GSH in cells. Ferroptosis had great potential to become a new approach in anti-tumor therapies in ovarian cancer.

Citation Format: Motoki Takenaka, Noriko Suzuki, Minako Mori, Hitomi Aoki, Tasuku Hirayama, Hideko Nagasawa, Ken-ichirou Morishige. Ferroptosis induced by erastin in RAS mutant ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2322.
Abstract 2323: Involvement of glutamine in tolerance of human pancreatic cancer cell to inhibition of fatty acid synthesis
Authors: Nishi Koji , Suzuki Mina , Yamamoto Noriko , Iwase Yumiko , Yumita Nagahiko
Published: August 2018

Pancreatic cancer (PC) is the fifth leading cause of cancer-related mortality in Japan. Although the standard chemotherapy regimens, FOLFIRINOX (fluorouracil, leucovorin, irinotecan and oxaliplatin), gemcitabine and nab-paclitaxel, are widely used to treat patients with advanced pancreatic cancer, their antitumor effects (progression and survival rates) are less potent against this cancer type than other solid tumors. Previously, we found that inhibition of acetyl-CoA carboxylase (ACC), a major rate-controlling enzyme of fatty acid synthetic pathway, suppressed the proliferation of PC cells. However, in this study, we found the tolerance to the inhibition in a part of PC cells. Therefore, we investigated the mechanism of the tolerance to ACC inhibition in PC cells. The human PC cell lines, AsPC-1, BxPC-3 and PANC-1 were obtained from the American Type Culture Collection. Live and apoptotic cell numbers were determined using the MUSE Annexin V and Dead Cell Kit according to the manufacturer's instructions. PANC-1 cells showed the tolerance to inhibition of fatty acid synthesis by ACC inhibitor, 5-(tetradecyloxy)-2-furoic acid (TOFA), whereas TOFA induced apoptosis in AsPC-1 and BxPC-3 cells. In addition, PANC-1 did not show autophagy even under the condition of fatty acid starvation. Next, we investigated whether PANC-1 has the tolerance to nutrient starvation in nutrient-depletion medium, Earle's Balanced Salt Solution (EBSS) and serum-free medium respectively. PANC-1 showed the survival even in such mediums, whereas AsPC-1 and BxPC-3 almost dead. This result suggests that the tolerance of fatty acid starvation of PANC-1 are related to that of nutrient starvation. Interestingly, proliferation of PANC-1 cultured in glutamine-depletion medium dramatically was suppressed, even though low glucose medium did not affect. BPTES, a selective inhibitor of glutaminase, also suppressed the proliferation and induced apoptosis in PANC-1, whereas there are little effects of 2-dexy-glucose, inhibitor of glycolysis, on them. These results suggest that the TCA cycle plays an important role on the tolerance to inhibition of fatty acid synthesis and nutrient starvation of PANC-1, and can be novel therapeutic target of PC cells which have the tolerance to nutrient starvation like PANC-1.

Citation Format: Koji Nishi, Mina Suzuki, Noriko Yamamoto, Yumiko Iwase, Nagahiko Yumita. Involvement of glutamine in tolerance of human pancreatic cancer cell to inhibition of fatty acid synthesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2323.
Abstract 2330: CaV1.3 calcium channel expression in prostate cancer cells during androgen-deprivation conditions
Authors: McKerr Niamh , Breen Conor , Srivastava Kirtiman , Mills Ian G. , McCloskey Karen D.
Published: August 2018

Background Prostate cancer (PCa) remains one of the most frequently diagnosed cancers in males in the western world. Despite efforts to reduce tumour burden through treatments including androgen-deprivation therapy (ADT), patients often relapse 2 years post-therapy due to emergence of castration-resistant disease. The importance of ion channels including calcium channels in cancer biology is increasingly recognized. CACNA1D encodes the calcium channel, CaV1.3, which is overexpressed in PCa biopsies. In addition to canonical ion transport, the c-terminal region of CaV1.3 may modulate gene expression. The purpose of this study was to elucidate the expression of CaV1.3 during PCa progression and explore the effects of calcium channel blockers. Methods A cell line panel comprising normal prostate epithelium (RWPE-1), androgen-sensitive PCa (LNCaP, VCaP) and castration-resistant PCa (C4-2B, DU145 and PC3) were used. LNCaP were cultured in charcoal-stripped serum-containing media to mimic ADT or with the anti-androgen enzalutamide (10μM) for 4-14 days. qPCR, Western blotting and immunofluorescence imaging were performed. Cell viability assays were performed in the absence/presence of ADT, enzalutamide and the CaV1.3 blocker nifedipine (1μM). Apoptosis was measured with flow cytometry in cells stained with Annexin V/Propidium Iodide (AV/PI). Results CACNA1D mRNA had higher expression in LNCaP, VCaP and C4-2B vs. RWPE-1 (p≤0.05; N=3). Western blotting/densitometry showed higher protein expression of CaV1.3 in LNCaP, VCaP and C4-2B relative to RWPE-1 (P≤0.05; N=3). Immunofluorescence indicated cytoplasmic and perinuclear localisation of CaV1.3 in the majority of cell lines and diffuse cytoplasmic staining in VCaP (N=2). ADT and enzalutamide-treated LNCaP displayed time-dependent increased CACNA1D expression over 14 days however, CaV1.3 protein expression peaked at 4 days (N=3). Cell viability assays of LNCaP treated with ADT or enzalutamide (4 days) showed significant reduction in viability when nifedipine was combined with the treatment (N=3; P≤0.05). Similarly, increased % of apoptotic LNCaP cells were observed when nifedipine was added to treated LNCaP vs. ADT or enzalutamide alone (N=2). Conclusion CaV1.3 expression is differentially expressed in cells lines modelling PCa progression and enhanced during ADT. Blockade of CaV1.3 in combination with ADT treatments further reduced cell viability and increased the apoptotic population.

Citation Format: Niamh McKerr, Conor Breen, Kirtiman Srivastava, Ian G. Mills, Karen D. McCloskey. CaV1.3 calcium channel expression in prostate cancer cells during androgen-deprivation conditions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2330.
Abstract 2331: Antioxidant enzymes mediate the survival of ECM-detached ovarian cancer cells
Authors: Versagli Calli A. , Libbing Cassandra
Published: August 2018

Epithelial ovarian cancer (EOC) is the deadliest gynecological cancer and is one of the leading causes of cancer death in women. It has been speculated that the high death rate associated with EOC is due to the fact that approximately 70% of all cases are diagnosed after the disease has already metastasized, thus making the illness more difficult to treat. In order to successfully survive the process of metastasis, EOC cells must be able to survive without anchorage to the extracellular matrix (ECM); however, the molecular mechanisms behind how EOC cells accomplish this feat has yet to be unveiled. Here, we present data that suggest antioxidant enzymes, namely Catalase, Peroxiredoxin 1, and Peroxiredoxin 2 all play a unique role in the survival and proliferation of ECM-detached SKOV3 cells. SKOV3 cells were genetically engineered to be deficient in Catalase, Peroxiredoxin 1, or Peroxiredoxin 2, and all three engineered cell types show abrogated anchorage-independent survival in soft agar. Furthermore, Catalase elimination in ECM-detached SKOV3 cells results in increased cell death at 48 hours, whereas elimination of Peroxiredoxin 1 or Peroxiredoxin 2 in ECM-detached SKOV3 cells results in decreased proliferation at 72 hours. Our current research is working to understand the molecular mechanisms behind the changes in survival and proliferation seen in antioxidant enzyme deficient SKOV3 cells. In aggregate, our data reveal that antioxidant enzymes may serve as attractive targets for the development of chemotherapeutics to eliminate metastatic ovarian cancer.

Citation Format: Calli A. Versagli, Cassandra Libbing. Antioxidant enzymes mediate the survival of ECM-detached ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2331.
Abstract 2333: Genetic characterization of programmed cell death in aneuploid yeast cells
Authors: Onyundo Megan , Sanborn Matthew , Timmel Victoria , Austriaco Nicanor
Published: August 2018

Aneuploidy is the genetic state of a cell that has a chromosomal number that is not an exact multiple of the haploid complement. It is a leading cause of spontaneous abortions and of mental retardation in humans, and is a characteristic defect in cancer. Yeast cells that are aneuploid manifest a diversity of phenotypes including cell cycle defects, genomic instability, protein imbalance, chaperone stress and proteotoxicity. We are investigating the links between aneuploidy on the budding yeast, Saccharomyces cerevisiae and programmed cell death. We have shown that aneuploid yeast cells are more sensitive to ethanol-induced cell death. In addition, we have also shown that caloric restriction protects aneuploid yeast from program cell death. [In addition to funding from the RI-INBRE Program, our laboratory is supported by grant NIGMS R15 GM110578, awarded to N. Austriaco.]

Citation Format: Megan Onyundo, Matthew Sanborn, Victoria Timmel, Nicanor Austriaco. Genetic characterization of programmed cell death in aneuploid yeast cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2333.
Abstract 2338: XAV-939, a Wnt/beta-catenin pathway inhibitor, sensitizes liver cancer cells to sorafenib: Implications in sorafenib resistance
Authors: Vishnoi Kanchan , Ke Rong , Vishwakarma Navin , Rana Ajay , Rana Basabi
Published: August 2018

Sorafenib is the only FDA-approved drug currently available for the treatment of advanced hepatocellular carcinoma (HCC). However, development of sorafenib-resistance in patients poses a major challenge to make this treatment successful in long-term. There is an urgent need to explore the mechanisms underlying sorafenib-resistance and to develop strategies towards overcoming sorafenib-resistance. Aberrant activation of Wnt/β-catenin signaling in HCC is suggested to maintain tumor initiating cells, drug-resistance, tumor progression, and metastasis. Combining β-catenin inhibitors with sorafenib might thus be an effective therapeutic strategy to target drug-resistant HCC. To explore this, we used wnt/β-catenin pathway inhibitor XAV-939 in combination with sorafenib to study their effect on the apoptotic potential of HCC cell lines Huh7, Hep3B and HepG2. XAV-939 was demonstrated to effectively reduce the expression of β-catenin in wild-type expressing Huh7 and Hep3B cells. Flow cytometric analysis of these cells pre-treated with XAV-939 followed by sorafenib treatment showed a significant increase in apoptosis in a time dependent manner. Furthermore, these cells revealed increased mitochondrial depolarization which was accompanied by a parallel increase in cytosolic cytochrome c release and downregulation of pro-survival protein Mcl-1. These effects of XAV-939 were further validated following knockdown of endogenous β-catenin, which showed a potentiation of sorafenib effects with β-catenin knockdown. Further, to understand the mechanism associated with sorafenib-resistance, we developed sorafenib-resistant HCC cells by repeated exposure to increasing concentration of sorafenib. To our surprise the combination of sorafenib and XAV-939 was ineffective in increasing the apoptotic potential of these resistant cells, suggesting a differential role of wnt/β-catenin axis in sorafenib-resistance. Taken together, these results suggested that combination of wnt/β-catenin pathway inhibitor and sorafenib could be effective in sensitizing HCC cells to apoptosis initially, but might be ineffective once these cells acquire sorafenib-resistance.

Citation Format: Kanchan Vishnoi, Rong Ke, Navin Vishwakarma, Ajay Rana, Basabi Rana. XAV-939, a Wnt/beta-catenin pathway inhibitor, sensitizes liver cancer cells to sorafenib: Implications in sorafenib resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2338.
Abstract 2343: Comprehensive evaluation of antibody-drug conjugate targets for drug development across tumor types: Analysis from TCGA
Authors: Groisberg Roman , Rosznik Jason , Meric-Bernstam Funda , Subbiah Vivek
Published: August 2018

Introduction: Cancer cells over-express proteins on their cell surface that can be exploited as drug targets and has led to development of two successful Anti-body Drug Conjugates (ADCs), Brentuximab and Trastuzumab Emtansine. Because a strong correlation exists between mRNA levels and protein expression we analyzed RNAseq data from TCGA to look at known ADC targets as a proof-of-concept to discovering new therapeutic targets.

Methods: We used mRNA expressions (transcripts per million, TPM) from publicly available RNA sequencing data from the Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) project to analyze the expression of ADC targets in tumor and normal tissues. We chose thirty-seven targets with known ADCs in clinical development. Graphs were generated to show distribution of each gene's expression across all tumor and normal tissue types in TCGA and GTEx. Potential targets were determined based on tumor expression of at least two times the highest expression in any normal tissue. Proteins known to be secreted in the blood stream at high levels were excluded as they are unlikely to make a good ADC target (eg. AFP, FOLH1).

Results: Using ERBB2 as the prototype gene, we were able to detect over-expression in all tumors that have HER-2 ove-rexpression in the literature including breast, bladder, NSCLC, Gastroesophageal, endometrial, HNSCC, ovarian, and colon cancer. Other genes with overexpression in select tumors including BSG (RCC, Melanoma, ACC), CD70 (RCC, DLBCL, mesothelioma, cervical), ENPP3 (RCC, endometrial, breast, lung adeno), FGFR2 (breast, gastric), FOLR1 (ovarian, lung adeno), GPNMB (lung, melanoma, HNSCC, renal), MSLN (lung adeno, mesothelioma, ovarian), SLC39A6 (breast), TDGF1 (thymoma, testicular, adrenocortical) were discovered as well. Tumors exhibited a significantly higher expression compared to normal tissue for ERBB2: breast cancer vs normal breast: p < 0.001; bladder cancer vs normal bladder: p < 0.001; lung adenocarcinoma vs normal lung: p < 0.001; ovarian cancer vs normal ovary: p < 0.001.

Conclusion: Overall, this large-scale analysis from TCGA reveals that RNA-seq can be used as an initial screen to identify novel targets for ADCs. Further analysis will identify previously unappreciated targets that can be exploited for drug development across diverse cancers. Further confirmation with protein expression is warranted.

Citation Format: Roman Groisberg, Jason Rosznik, Funda Meric-Bernstam, Vivek Subbiah. Comprehensive evaluation of antibody-drug conjugate targets for drug development across tumor types: Analysis from TCGA [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2343.
Abstract 2344: Identification and validation of synthetic lethal cancer pairs from genome-wide CRISPR screens
Authors: Schwartz Benjamin , Jing Junping , Stronach Euan , Cooper David , Wilson Boris , Middleton Sarah , Khaladkar Mugdha , Dong Coco , Shabon Usman , Rao Yanhua , Riesenberg Stefanie
Published: August 2018

Although most canonical somatic cancer mutations have now been cataloged, only a fraction are amenable to drug discovery efforts. These are limited to targets with gain of function mutations and druggable protein domains. Examples include kinases such as FLT3 and metabolic enzymes such as IDH1/2. However, excluded from this list are undruggable protein targets with gain of function mutations, such as KRAS, or tumor suppressors which acquire loss of function mutations, such as APC.

The concept of synthetic lethality is being used to address this gap in cancer discovery. Cells which carry these directly undruggable somatic mutations become susceptible to inhibition of secondary targets. This susceptibility is specific in context to cells carrying the cancer associated mutation, offering both a novel approach for establishing efficacy as well as a potential therapeutic window for normal cells. The successful development of PARP inhibitors for BRCA mutant ovarian and breast cancers has provided the first clinical validation of this idea.

Pooled, genome-wide knock out experiments are being pursued to systematically discover more of these types of synthetic lethalities. Typically, a sgRNA library is used to transfect a CAS9 expressing cancer cell line, and sequencing of the guides over time identifies lethalities specific to the cell line. Once this experiment is repeated over hundreds of cell lines, one can use the genomic information of the cell lines, such as mutations or copy number aberrations, to identify potential new combinations of molecular lethality.

We have analyzed pooled CRISPR knockout data in a large panel of cancer cell lines generated from two independent sources (Broad Institute and Open Targets) to identify these types of lethality. Besides identifying targets for prevalent cancer mutations, we have also identified lethal combinations based on genes aberrantly deleted/silenced in cancers, termed “collateral lethality.” These include pairs of paralogs, many of which code enzymes with redundant functions.

In this presentation we will outline our analysis strategies and methods, and describe the development of a downstream experimental validation pipeline, using a specific target/predictor pair to exemplify our approach.

Citation Format: Benjamin Schwartz, Junping Jing, Euan Stronach, David Cooper, Boris Wilson, Sarah Middleton, Mugdha Khaladkar, Coco Dong, Usman Shabon, Yanhua Rao, Stefanie Riesenberg. Identification and validation of synthetic lethal cancer pairs from genome-wide CRISPR screens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2344.
Abstract 2345: The genomic landscape of small cell carcinoma of the oesophagus
Authors: Wang Feng , Liu Dongbing , Zhao Qi , Chen Gang , Liu Xinming , Wang Yingnan , Su Hong , Qin Yanru , He Yifu , Zou Qingfeng , Liu Yanhui , Lin Youen , Sheng Hui , Hu Jiajia , Liu Zexian , Zuo Zhixiang , Bei Jinxin , Xu Ruihua
Published: August 2018

Background & Aims—Small cell carcinoma of the oesophagus (SCCE) is a deadly malignancy but its genetic characteristics remain unknown and treatment is commonly adopted from therapies for its histologically identical counterpart, small cell lung cancer (SCLC). We conducted a large-scale genomic analysis of SCCE to determine the mutational landscape of this deadly disease.

Methods—We performed whole-exome sequence of tumour and para normal esophageal tissues from 55 patients with SCCE who underwent surgery at seven institutions in China. Comprehensive analyses were carried out to identify key genetic events in SCCE, including significant mutations, copy number variation, pathway disruption. We also performed ultra-deep targeted sequencing on 20 specimens and microarray assays on 24 specimens to validate somatic mutations and copy number variations respectively. Using an unsupervised hierarchical clustering approach, the genomic characteristics of SCCE were compared with other types of malignancies including oesophageal squamous cell carcinoma, oesophageal adenocarcinoma and et al. In vitro proliferation and migration assays were conducted to assess the function of PDE3A.

Results—The mutational spectrum of SCCE was dominated by C>T/G>A transitions and the mutational signatures classified the patients into 3 groups. We identified three novel significantly mutated genes (PDE3A, PTPRM and CBLN2) and mutations in five other well-known tumour-associated genes (TP53, RB1, NOTCH1, FAT1 and FBXW7). Non-silent alterations of chromatin remodeling genes were detected in 96.4%. Moreover, pathway enrichment analysis revealed that genes involved in the cell cycle, p53, Notch and Wnt signaling were frequently altered in SCCE. Mutations in NOTCH family correlated with shorter overall survival. Furthermore, comparison analysis of mutation signatures revealed that SCCE tumours were more closely related to oesophageal squamous cell carcinoma, rather than oesophageal adenocarcinoma and SCLC, suggesting that current therapies for SCCE may be inadequate. Functional characterizations suggested that PDE3A acts as a novel oncogene.

Conclusions—Our findings reveal key somatic alterations in SCCE, and provide insights to better understand the pathogenesis of SCCE and to develop better treatment strategies for patients with the condition.

Citation Format: Feng Wang, Dongbing Liu, Qi Zhao, Gang Chen, Xinming Liu, Yingnan Wang, Hong Su, Yanru Qin, Yifu He, Qingfeng Zou, Yanhui Liu, Youen Lin, Hui Sheng, Jiajia Hu, Zexian Liu, Zhixiang Zuo, Jinxin Bei, Ruihua Xu. The genomic landscape of small cell carcinoma of the oesophagus [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2345.
Abstract 2350: Efficient survival multifactor dimensionality reduction method for genome-wide association study
Authors: Gui Jiang , Ji Xuemei , Amos Christopher I.
Published: August 2018

The problem of identifying associations between SNPs and case-control outcome has been studied extensively and a number of new techniques have been developed. Little progress has been made, however in the analysis of SNP effect in relation to censored survival data. We present an extension of the two class multifactor dimensionality reduction (MDR) algorithm that enables detection and characterization of epistatic SNP-SNP interactions in the context of survival outcome. The proposed an Efficient Survival MDR (ES-MDR) method handles censored data by modifying MDR's constructive induction algorithm to use logrank Test. We applied ES-MDR to genetic data of over 470,000 SNPs from the OncoArray Consortium. We use onset age of lung cancer and case-control (n=27,312) status as the survival outcome and divided data into training and testing sets. We also adjust for subject's age, gender and smoking status. From training set, we identified strong association from SNPs in BRCA1 and IL17RC genes with lung cancer onset age. This result is validated in the testing set.

Citation Format: Jiang Gui, Xuemei Ji, Christopher I. Amos. Efficient survival multifactor dimensionality reduction method for genome-wide association study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2350.
Abstract 2354: Candidate non-coding driver mutations in super-enhancers and long-range chromatin interaction networks across 1,800 whole cancer genomes
Authors: Reimand Juri
Published: August 2018

A comprehensive understanding of the mutations that drive tumorigenesis and disease progression is essential to understanding tumor biology and designing precision therapies. The landscape of driver mutations in the protein-coding genome has been well-characterized by large exome-sequencing studies. Surprisingly, many tumors do not have mutations in any known protein-coding driver. Non-coding driver mutations are hypothesized to explain many of these cases, but aside from a few regions, like the TERT promoter, our understanding of drivers in the complex regulatory genome remains limited. To fill this gap, we analyzed 150,000 cis-regulatory modules with clustered transcription factor binding sites in 1,844 whole cancer genomes from the ICGC-TCGA PCAWG project. Using a new statistical method, ActiveDriverWGS, we identified dozens of frequently mutated regulatory elements (FMREs) enriched in non-coding SNVs and indels (FDR<0.05) with many structural rearrangements and focal copy number alterations in additional samples. The FMREs were enriched in super-enhancers, long-range chromatin interactions and H3K27ac marks derived from primary tumors, suggesting a gene regulatory role of these mutations through three-dimensional genome organization. The interaction network of chromatin loops and FMREs revealed putative target genes located dozens to hundreds of kilobases away from the mutated regulatory elements. We found known and putative oncogenes and tumor suppressors whose expression significantly correlated with mutations in FMREs, suggesting novel oncogenic mechanisms. Most of the FMREs were also confirmed by additional driver discovery methods, lending confidence to our statistical approach. We also validated ActiveDriverWGS on protein-coding sequence and accurately recovered known driver genes. The non-coding regulatory genome is characterized by diverse mutational processes, regional hypermutations and technically challenging areas with suboptimal sequencing coverage. Thus our findings, most of which are reported for the first time, should be carefully vetted and experimentally validated in future studies. Our integrative analysis of somatic mutations, cis-regulatory regions and long-range chromatin interaction networks is a novel framework for cancer discovery and reveals the currently largest set of potential non-coding drivers in a pan-cancer cohort.

Citation Format: Juri Reimand. Candidate non-coding driver mutations in super-enhancers and long-range chromatin interaction networks across 1,800 whole cancer genomes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2354.