bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2298A: Enhanced consideration of context dependence reveals the scope of somatic missense drivers in human cancers
Authors: Tokheim Collin J. , Nanda Neha , Roberts Nicholas J. , Karchin Rachel
Published: August 2018

Large-scale sequencing studies of patient cohorts have enabled identification of many driver genes whose somatic alterations confer a selective advantage to cancer cells. However, not every mutation in a driver gene is necessarily a driver of cancer. As mutation-based biomarkers are increasingly used to decide drug regimens and placement in clinical trials, it is important to distinguish specific driver mutations. Improved computational methods are necessary for identifying the particular mutations in a given cancer that are drivers, especially for rare mutations (<1% of patients). We have previously developed the CHASM algorithm to identify cancer-specific driver missense mutations. Here, we present a completely re-worked version that leverages the context-dependence of driver mutations. We show that the new CHASM2 consistently outperforms comparable methods across a wide variety of benchmarks, including in vivo and in vitro experiments. Applied to 8,657 TCGA cancer samples in 32 cancer types, CHASM2 identifies 3,527 distinct driver somatic missense mutations. Our results suggest substantial heterogeneity of the driver mutation landscape across cancer types and a strikingly large prevalence of rare driver mutations. For particular cancer types driven by common mutations, current sample sizes are already saturating driver discovery. We performed validation experiments on missense mutations of rare and intermediate frequency in the tumor suppressor gene ATM and show CHASM2 distinguishes a radiosensitive phenotype in cultured cells. To our knowledge CHASM2 is the first sufficiently powered method to predict individual driver somatic missense mutations at multiple scales – whole exome, targeted gene panel, and within a single gene.

Citation Format: Collin J. Tokheim, Neha Nanda, Nicholas J. Roberts, Rachel Karchin. Enhanced consideration of context dependence reveals the scope of somatic missense drivers 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 2298A.
Abstract 2305: Targeting 6-phosphofructo-2-kinase to increase the efficacy of ER and CDK4/6 inhibitors against breast cancer
Authors: Imbert-Fernandez Yoannis , Lanceta Lilibeth , Telang Sucheta , Chesney Jason
Published: August 2018

We demonstrated that estradiol stimulates the expression and activity of an enzyme, 6-phosphofructo-2-kinase (PFKFB3), that regulates glucose metabolism via allosteric stimulation of 6-phosphofructo-1-kinase. PFKFB3 expression is increased in breast cancer cells that have metastasized to lymph nodes and the combination of the anti-estrogen, fulvestrant, with a PFKFB3 inhibitor, PFK158, synergistically increases apoptosis in ER+ breast cancer cells in vitro and in vivo. Given that estradiol simultaneously increases the expression of both the regulatory subunit of CDK4/6, cyclin D1, and PFKFB3, we postulated that the combination of palbociclib with PFK158 would yield synergy against breast cancer cells. In new preliminary studies, co-treatment with palbociclib and PFK158 resulted in a marked decrease in cell proliferation in two ER+ breast cancer cell lines that was accompanied by a synergistic decrease in phospho-Retinoblastoma (Rb). Importantly, exposure of ER+ breast cancer cells to palbociclib and PFK158 causes an increase in cell cycle arrest and apoptotic cell death in vitro. Notably, this combination has no effect on cell cycle arrest or apoptosis in normal mammary cells suggesting that this therapeutic strategy may be well tolerated and selective for breast cancer.

Citation Format: Yoannis Imbert-Fernandez, Lilibeth Lanceta, Sucheta Telang, Jason Chesney. Targeting 6-phosphofructo-2-kinase to increase the efficacy of ER and CDK4/6 inhibitors against breast 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 2305.
Abstract 2312: Phospho-proteomics analysis to determine the signaling pathways affected by a novel CDC7-selective inhibitor TAK-931
Authors: Iwai Kenichi , Gotou Masamitsu , Yamanaka Kazunori , Ohashi Akihiro
Published: August 2018

Background: Cell division cycle 7 (CDC7) is a serine/threonine kinase, which plays important roles in initiation of DNA replication by phosphorylating MCM2. We developed the CDC7-selective small molecule inhibitor TAK-931. Treatment with TAK-931 results in delayed S phase progression, DNA replication stress (RS), centrosome disorganization, mitotic aberrations, and potent growth suppression in cancer cells. In this study, we conducted the time-dependent phospho-proteomics analysis to determine the signaling pathways affected by TAK-931.

Materials and Methods: TAK-931 was synthesized by Takeda Pharmaceutical Company, Ltd. The SILAC-labeled COLO205 cells were treated with TAK-931 at 100 nM for 0, 4, 24 hours, applying for the phospho-proteomics analysis in the PhosphoScout® Service at Evotec AG (Munich, Germany).

Results: The SILAC-labeled COLO205 cells with TAK-931 treatment were subjected to quantitative mass spectrometry, detecting 15,128 phosphorylation sites of the 4,426 phosphoproteins. At 4 h of treatment, only 2 phosphorylation sites were detected as significantly downregulated: MAX gene-associated protein (MGA) and alpha-enolase (ENO1). At 24 h of treatment, however, 51 phosphorylation sites were identified as being significantly altered, including 44 upregulation and 7 downregulation sites. The gene ontology (GO) enrichment analysis in the categories of biological process (BP) and cellular component (CC) revealed that 10 GO terms from BP and 2 GO terms from CC were significantly enriched in the treated cells after 24 h: cell cycle, cell cycle phase, cell cycle process, mitosis, nuclear division, organelle fusion, organelle organization, mitotic cell cycle, M phase of mitotic cell cycle, M phase, chromosomal part, and spindles. Network analysis on the changed phosphorylation prolife also revealed that significant network modules existed at 24 h of treatment, which included DNA replication/damage, mitotic progression/spindle assembly, nucleus assembly, GTPase activity, cell migration, cell adhesion, TP53, and protein translation.

Conclusion: The time-dependent phospho-proteomics analysis revealed the dynamics of phospho-signal transduction affected by TAK-931 in the cancer cell, which may help to provide a deeper understanding of the molecular mechanisms of TAK-931.

Citation Format: Kenichi Iwai, Masamitsu Gotou, Kazunori Yamanaka, Akihiro Ohashi. Phospho-proteomics analysis to determine the signaling pathways affected by a novel CDC7-selective inhibitor TAK-931 [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 2312.
Abstract 2318: Palbociclib, a CDK4/6 inhibitor, suppresses proliferation of triple negative breast cancer
Authors: Min Ahrum , Kim Yu Jin , Hang Hyemin , Lim Jee Min , Kim Seongyeong , Kim So Hyeon , Suh Koung Jin , Lee Kyung-Hun , Kim Tae-Yong , Im Seock-Ah
Published: August 2018

Background: Abnormal cell cycle progression is a characteristic of cancer and targeting the cell cycle is a strategy for cancer treatment. TCGA reported that Cyclin D1 is overexpressed in 25% to 60% of invasive breast carcinomas, and CDK4/6 and CDKN2A/B aberrations are observed in 7.6% of breast cancer cases. Palbociclib is a specific inhibitor of CDK4/6, a key regulator of the G1 checkpoint. In hormone receptor positive breast cancer, the addition of palbociclib to endocrine treatment either aromatase inhibitor or fulvestrant significantly prolonged PFS in phase III trials. However, the effects of palbociclib have not been fully examined in triple negative breast cancer (TNBC) yet, although CCND amplification and CDKN2A loss are frequently observed in TNBC. Therefore, we investigated the effects of palbociclib on TNBC cells and attempted to identify the underlying mechanisms of palbociclib.

Methods: The cytotoxicity assay, cell cycle analysis and western blotting were conducted to determine anti-tumor effect and action mechanisms of palbociclib on breast cancer cell lines. These in vitro data were validated in vivo model as well.

Results: Human breast cancer cell lines showed heterogeneous response to palbociclib. Palbociclib induced G1 cell cycle arrest by blocking Rb phosphorylation, and inhibited cell proliferative signaling. In sensitive TNBC cells, palbociclib promoted senescence rather than apoptosis. In addition, palbociclib enhanced the antitumor effect of 5-FU by modulating thymidine synthase (TS) and E2F1 expression. The anti-tumor effects of palbociclib on TNBC cells were validated in TNBC-xenograft model as well.

Conclusions: CDK4/6 inhibitor palbociclib showed anti-tumor effect in vitro and in vivo xenograft model of TNBC and acts synergistically with 5-FU by downregulating TS. Our results suggest that palbociclib has therapeutic potential for the treatment of TNBC, not limited in hormone positive breast cancer type. Our results provide a rationale for the future clinical trials of palbociclib in the treatment of breast cancers.

Citation Format: Ahrum Min, Yu Jin Kim, Hyemin Hang, Jee Min Lim, Seongyeong Kim, So Hyeon Kim, Koung Jin Suh, Kyung-Hun Lee, Tae-Yong Kim, Seock-Ah Im. Palbociclib, a CDK4/6 inhibitor, suppresses proliferation of triple negative breast 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 2318.
Abstract 2320: Heme oxygenase-1 mediates ferroptosis
Authors: Chiang Shih-Kai , Chen Shuen-Ei , Chang Ling-Chu
Published: August 2018

Ferroptosis is a form of oxidative cell death and has become a chemotherapeutic target for cancer treatment. BAY 11-7085 (BAY), which is a well-known IκBα inhibitor, suppressed viability in cancer cells via induction of ferroptotic death in an NF-κB-independent manner. This study was aimed to characterize the underlying mechanism by BAY and demonstrate its potential chemotherapeutic targets for cancer treatment. Reactive oxygen species scavenging, relief of lipid peroxidation, replenishment of glutathione and thiol-containing agents, as well as iron chelation, rescued BAY-induced cell death in human breast cancer cells (MDA-MB-231 and MDA-MB-468) and glioblastoma cells (DBTRG-05MG). The results from quantitative real-time PCR and western blotting analysis showed that BAY upregulated a variety of nuclear factor-E2-related factor 2 (Nrf2) target genes related to redox regulation, particularly heme oxygenase-1 (HO-1) and solute carrier family 7 membrane 1 (SLC7A11). Studies with pharmacological inhibitors and shRNA interventions suggested that the hierarchy of induction is Nrf2-SLC7A11-HO-1. The ferroptotic process induced by hHO-1 overexpression further indicated that HO-1 is a key mediator of BAY-induced ferroptosis that operates through cellular redox regulation and iron accumulation. Our study discovered that BAY induced ferroptosis via Nrf2-SLC7A11-HO-1 pathway and provided new insights to understand the mechanism of HO-1-induced ferroptosis, suggesting HO-1 as a new chemotherapeutic target for cancer treatment.

Citation Format: Shih-Kai Chiang, Shuen-Ei Chen, Ling-Chu Chang. Heme oxygenase-1 mediates ferroptosis [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 2320.
Abstract 2327: Azole antifungal drugs induce cell death by suppressing mTOR through PI3K/Akt inhibition in human breast cancer
Authors: Park Ju Ho , Kim Hyesook , Kim So Hee
Published: August 2018

Breast cancer is a major cancer in women and its incidence rate increases every year. So it needs a perfect treatment. Azole drugs structurally have an imidazole [clotrimazole (CTZ) and ketoconazole (KCZ)] or a triazole [fluconazole (FCZ) and itraconazole (ICZ)]. Recently, azole antifungal drugs have shown anticancer efficacy in clinical trial. However, their antitumor mechanism are vague. Here, we evaluated the effects, as apoptosis, cell arrest, and autophagy and tumor growth using human breast cancer cells. Apoptosis assay showed imidazole-induced cell death, pro-apoptosis maker, as cleaved PARP, cleaved caspase-3, Bax, increased and anti-apoptosis maker, as Bcl-2, decreased by CTZ and KCZ in both MCF-7 and MDA-MB-231 cells. We also checked imidazole of azole drugs blocked the cell cycle in G1 phage by downregulating the protein levels of cyclin D1, CDK2, and CDK6 and increased p21 and p27 in breast cancer. Further experiments showed that azole drugs also induced autophagy through the increased accumulation of LC 3B inside the cells by immunoblot analysis and immunofluorescence staining, increased beclin-1 and decreased p62 protein. With azole decreased that though 3-MA. Azole drugs inhibited PI3K/Akt/mTOR signaling pathway. Tumor growth was also inhibited in nude mice bearing MDA-MB-231 xenograft by CTZ, KCZ and ICZ through the accumulation of LC 3B in tumor xenograft. In conclusion, imidazole antifungal drugs induced apoptosis, cell cycle and autophagy by suppressing mTOR through the regulation of PI3K/Akt pathway in human breast cancer cells, suggesting azole antifungal drugs could be new potential therapeutics for breast cancer.

Citation Format: Ju Ho Park, Hyesook Kim, So Hee Kim. Azole antifungal drugs induce cell death by suppressing mTOR through PI3K/Akt inhibition in human breast 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 2327.
Abstract 2328: ADCs that induce necrosis and thwart apoptotic resistance
Authors: Prudent James R. , Marshall David
Published: August 2018

Purpose: We have previously shown that targeting the Na,K-ATPase (NKA) with antibody drug conjugates is a powerful method for reducing tumors at levels found to be safe in cynomolgus monkeys. Currently no approved cancer drugs induce a potent necrotic type cell death even though experimental evidence shows that necrotic cell death has many notable and untapped benefits. Therefore, we wanted to better understand the driving mechanism by which ADCs targeting NKA inhibition induce necrosis.

Experimental Design: Various cancer cell types were subjected to increasing and decreasing concentrations of Na+, K+ and Ca++, as well as various ADCs using bufalin as the warhead/payload/toxin. The cells were also subjected to siRNAs specific to the NKA and bufalin alone. The cells were then analyzed for the appearance and/or presence of necrosis, apoptosis, and markers indicating various stress pathways.

Results: Conditions found to induce necrosis included: ADCs targeting specific NKA-protein complexes, cells subjected to media containing increased concentrations of K+, the specific Na,K-ATPase siRNAs, and bufalin alone.

Conditions not found to induce necrosis included: media containing increased Na+, decreased Na+ or K+, control siRNAs, or ADCs that targeted proteins not complexed to the NKA or expressed on the cell surface.

All cells undergoing necrosis lost their actin filaments, rounded up, and adherent cells detached from the surface and lost their motility. Markers indicated initial activation of the integrated stress responses (ISR). Minimal to no annexin staining indicated that apoptosis was not the dominate pathway in any experiment.

Conclusion: Proper ion flux is important for maintaining cellular homeostasis. Cell swelling is a key process that is used to correct for changes in ion flux, induce inflammation and kill unwanted cells. The NKA is a key regulatory protein in this process and when blocked, leads to uncontrolled cells swelling and necrotic death. Precise inhibition of ion flux focused on cancer cells should be further investigated as a potential method for treating patients.

Citation Format: James R. Prudent, David Marshall. ADCs that induce necrosis and thwart apoptotic 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 2328.
Abstract 2329: Noncanonical mitophagy induced by Mieap and its role in tumor suppression via ferroptosis-like cell death
Authors: Nakamura Yasuyuki , Yamamoto Makoto , Arakawa Hirofumi
Published: August 2018

Parkin/Pink1-mediated mitophagy plays a critical role in mitochondrial quality control, in which the damaged mitochondria are sequestered by autophagosomes, and degraded by fusion between autophagosomes and lysosomes. Here, we report Mieap-mediated noncanonical mitophagy as a new function of tumor suppressor p53. Mieap was originally identified as a p53-target gene. Overexpression of exogenous Mieap induces large vacuoles in cancer cells. Mieap-induced vacuoles are generated from the mitochondria, and directly eat and degrade the cancer mitochondria. UVRAG regulates the Mieap-mediated noncanonical mitophagy. Surprisingly, Mieap-mediated noncanonical mitophagy induces cell death via the iron-dependent production of reactive oxygen species (ROS). Introduction of exogenous Mieap strikingly suppresses in vivo tumor growth of Mieap-deficient colorectal cancer cells. Deficiency of endogenous Mieap dramatically promotes cancer development, malignancy, and aggressiveness in colorectal and gastric cancer model mice. The cell death induced by Mieap-mediated noncanonical mitophagy occurs via the ROS production in in vivo hypoxic tumor microenvironment. These results suggest that Mieap-mediated noncanonical mitophagy plays a critical role in tumor suppression via ferroptosis-like cell death.

Citation Format: Yasuyuki Nakamura, Makoto Yamamoto, Hirofumi Arakawa. Noncanonical mitophagy induced by Mieap and its role in tumor suppression via ferroptosis-like cell death [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 2329.
Abstract 2334: Sialylation of the TNFR1 death receptor promotes cancer cell survival
Authors: Holdbrooks Andrew T. , Schultz Matthew J. , Liu Zhongyu , Bullard Daniel C. , Bellis Susan L.
Published: August 2018

Studies have well-established that activation of TNFR1, a ubiquitously expressed cell surface receptor, by TNF can induce either cell survival or cell death signaling cascades, however, the mechanisms regulating TNFR1 to initiate these disparate outcomes are poorly understood. TNF-induced apoptosis is initiated by clustering of activated TNFR1 at the cell surface, followed by internalization of the receptor complexes and subsequent caspase activation. Conversely, surface retention of activated TNFR1 promotes cell survival signaling mediated by NFκB and MAPKs pathways. Our group has identified a glycosylation-dependent mechanism that controls this TNFR1 signaling switch (i.e. apoptosis vs. survival). Specifically, we have found that TNFR1 activity is modified by the addition of a distinct sugar, an α2-6 linked sialic acid, by the Golgi sialyltransferase, ST6Gal-I. Importantly, ST6Gal-I is highly upregulated in numerous cancer types, and through its sialylation of a select cohort of cell surface receptors, various studies have indicated ST6Gal-I as a potent tumor cell survival factor. The effect of sialylation on TNFR1 was examined in multiple cancer cell models, including leukemia and epithelial cancer cell lines with forced overexpression or knockdown of ST6Gal-I. These models show that TNFR1 sialylation blocks TNF-induced apoptosis (indicated by the inhibition of caspase activation), and data from our group suggests that the underlying mechanism of this inhibition of apoptosis involves the sialylation-driven interference of TNFR1 oligomerization and internalization. Considering sialylated TNFR1 is retained on the cell surface following activation, we hypothesize that cells with abundant ST6Gal-I not only block TNFR1-mediated apoptosis but also divert signaling towards survival, indicated by increased activity and expression of many pro-survival factors (NFkB, AKT, etc.). Based on these collective findings, we conclude that the ST6Gal-I-mediated sialylation of TNFR1 controls the cellular response to TNF by blocking TNFR1 internalization, resulting in apoptosis inhibition and the promotion of survival signaling. Considering the tumor microenvironment is rich in immune cell-derived TNF, we posit that ST6Gal-I protects tumor cells within this inflammatory milieu by providing a mechanism by which tumor cells can evade immune cell killing.

Citation Format: Andrew T. Holdbrooks, Matthew J. Schultz, Zhongyu Liu, Daniel C. Bullard, Susan L. Bellis. Sialylation of the TNFR1 death receptor promotes cancer cell survival [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 2334.
Abstract 2339: Targeting ICMT in pancreatic cancer
Authors: Manu Kanjoormana A. , Chai Tin F. , Teh Jing T. , Zhu Wan , Casey Patrick J. , Wang Mei
Published: August 2018

Introduction: Pancreatic Cancer remains one of the most difficult to treat human cancers with the 5-year survival as low as 5%. No targeted therapy yet developed against Pancreatic cancer. Here we want to target Isoprenylcysteine carboxylmethyltransferase (ICMT), an enzyme that post translationally modifies a group of proteins including several small GTPases against Pancreatic Cancer. Method: ICMT inhibition was achieved using lentivirus expressing ICMT shRNA and a pharmacological inhibitor named Cysmethynil. We evaluated a panel of human pancreatic cancer cell lines and identified those that are sensitive to ICMT inhibition by cell viability assay. This responsiveness to ICMT inhibition was confirmed in in vivo xenograft tumor mouse models using both Cysmethynil and shRNA-targeting ICMT. Flow cytometry analysis after DAPI or PI staining is used to detect cell cycle arrest and apoptosis. We used western blot analysis to study autophagy, apoptosis and cell cycle markers. ShRNA for P21 is used for p21 knock down studies. SiRNA for p53 is used to study whether p21 induction is p53 dependent. BNIP3, ULK1 and ATG5 knock down was done using specific shRNAs to study their role in ICMT inhibition induced autophagy and apoptosis. BCL-xL is over expressed in the cells using retroviral vectors to study the role of apoptosis in ICMT inhibition induced cell death. Cell and mitochondrial respiration was determined by oxygen consumption rate analysis using Seahorse XF24 analyzer. Summary: In sensitive pancreatic cancer cells, ICMT inhibition induces mitochondrial respiratory deficiency and cellular energy depletion, leading to significant upregulation of p21 and p21 dependent induction of BNIP3 leading to autophagy and apoptosis. But unlike in prostate and liver cancer, ICMT inhibition induced apoptosis not the autophagy responsible for inhibition of cell viability in pancreatic cancer. Conclusion: These findings suggest ICMT is a potential therapeutic target against pancreatic cancers and atleast in pancreatic cancer, ICMT inhibition induced apoptosis but not autophagy responsible for its therapeutic efficacy.

Citation Format: Kanjoormana A. Manu, Tin F. Chai, Jing T. Teh, Wan Zhu, Patrick J. Casey, Mei Wang. Targeting ICMT in pancreatic 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 2339.
Abstract 2351: Transcriptome-wide association study reveals candidate causal genes for lung cancer
Authors: Clemenceau Alisson , Lamontagne Maxime , Torres Robert Carreras , Obeidat Ma'en , Timens Wim , Joubert Philippe , Amos Christopher I. , McKay James D. , Bossé Yohan
Published: August 2018

Genome-wide association studies have identified robust susceptibility loci associated with lung cancer. As part of the OncoArray-TRICL consortium, we have recently completed the largest GWAS on lung cancer including 29,266 cases and 56,450 controls of European descent. The goal of this study is to integrate the complete GWAS results with a large-scale expression quantitative trait loci (eQTL) mapping study in human lung tissues (n=1,038) to identify candidate causal genes for lung cancer. Transcriptome-wide association study (TWAS) was used to integrate GWAS and lung eQTL signals and identify genes whose levels of expression in lung tissue are causally related to lung cancer. TWAS was performed on six histologic and smoking subgroups, namely overall lung cancer, adenocarcinoma, squamous cell carcinoma, small cell carcinoma, never-smokers, and ever-smokers. As expected, the main TWAS signal for all histologic subtypes and ever-smokers was on chromosome 15q25. The genes most strongly associated with lung cancer at this locus were IREB2 (PTWAS=4.97E-104), and to a lower extent, CHRNA5 (PTWAS=5.26E20) and HYKK (PTWAS=2.04E-17). TWAS identified causal genes were different from those reported in GWAS, including JAML on 11q23.3 in overall lung cancer (PTWAS=1.39E-6) and adenocarcinoma (PTWAS=2.09E-8), NOTCH4 on 6p21.32 in squamous cell carcinoma (PTWAS=1.24E-12), ZNRD1 on 6p22.1 in overall lung cancer (PTWAS=3.41E-14) and ever-smokers (PTWAS=1.29E-9), HIST1H2BD on 6p22.2 for small cell carcinoma (PTWAS=1.54E-6), and NEXN on 1p31.1 in never-smokers (PTWAS=2.64E-5). In addition, a new small cell carcinoma susceptibility locus was identified on 4q32.2 and associated with the expression levels of TMA16 (PTWAS=4.2E-6). In conclusion, lung tissue TWAS on lung cancer, histologic subtypes and smoking subgroups revealed novel causal genes in GWAS-nominated loci. A new locus for small cell carcinoma (4q32.2-TMA16) was also identified and will require further validation.

Citation Format: Alisson Clemenceau, Maxime Lamontagne, Robert Carreras Torres, Ma'en Obeidat, Wim Timens, Philippe Joubert, Christopher I. Amos, James D. McKay, Yohan Bossé. Transcriptome-wide association study reveals candidate causal genes for lung 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 2351.
Abstract 2363: Novel CDH1 mutations in breast invasive lobular carcinoma
Authors: Frampton Garrett M. , Hartmaier Ryan , Sokol Ethan , Gupta Anika , Greenbowe Joel , Roels Steven , Gay Laurie , Stephens Philip J.
Published: August 2018

Breast invasive lobular carcinoma (ILC) is the second most common type of breast cancer, making up approximately 10% of all invasive breast cancers. Breast ILC is characterized by loss of e-cadherin protein expression (CDH1), which is usually caused by loss of function mutations in the CDH1 gene. A subset of breast ILC cases have no identifiable CDH1 mutation, but presumably inactivate CDH1 through currently unknown mechanisms.

Tumors from more than 150,000 unique patients including more than 15,000 breast cancers were examined by comprehensive genomic profiling to detect base substitutions, indels, copy number alterations, and genomic rearrangements in the full coding regions of up to 465 genes. Statistical significance of the disease distributions was calculated using Fisher's exact test with false discovery rate multiple hypothesis testing correction.

Of 760 breast ILC cases, 583 (76.7%) had a known loss-of-function mutation in the CDH1 gene. To identify previously unknown mutations that result in or substitute for CDH1 loss-of-function we performed statistical analysis on all mutations, in any gene, occurring more than once in the 177 ILC cases without an identifiable CDH1 mutation. The most statistically significant mutation was very near to a splice site of CDH1 (chr16:68863554 c.2296-3A>G), occurring in 7 CDH1 negative ILC cases as well as 9 other cases in cancer types that frequently harbor CDH1 mutations (7 breast, 2 gastroesophageal junction) and 1 colorectal cancer. Based on tumor-only test based zygosity modeling, this mutation is only observed as somatic and usually homozygous in the tumor. Four other positions near to splice sites of CDH1 (chr16:68844247 c.832+3A>G, chr16:68849667 c.1565+5G>A, chr16:68856133 c.1936+5G>A, chr16:68857532 c.2164+3A>C), mutated in 13 CDH1 negative ILC cases, were also statically significant, somatic, and usually homozygous. No statistically significant mutations were found in genes other than CDH1.

Previously unappreciated somatic, non-canonical splice site mutations in CDH1 occur in ~10% (20/177) of breast ILC cases with no known CDH1 mutation. These mutations occur almost exclusively in tumor types known to harbor frequent CDH1 mutations and are usually homozygous in the tumor, strongly indicating that they are CDH1 loss-of-function mutations.

Citation Format: Garrett M. Frampton, Ryan Hartmaier, Ethan Sokol, Anika Gupta, Joel Greenbowe, Steven Roels, Laurie Gay, Philip J. Stephens. Novel CDH1 mutations in breast invasive lobular carcinoma [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 2363.
Abstract 2364: The phosphatase PRL-3 is a pro-survival oncogene in acute lymphoblastic leukemia
Authors: Wei Min , O'Leary Kristin , Liu Jinpeng , Wang Chi , Blackburn Jessica
Published: August 2018

PRL-3, phosphatase of regenerating liver 3, is one member of a subfamily of protein tyrosine phosphatases that belongs to the dual-specificity phosphatases (DSPs). Accumulating evidence shows that PRL-3 is a biomarker of tumor progression and metastasis and high PRL-3 expression is associated with reduced overall and disease-free survival in a variety of solid tumors, suggesting it's importance in tumor recurrence. However, the role for PRL-3 in leukemia relapse has not yet been described.

We have found that PRL-3 contributes to Acute Lymphoblastic Leukemia (ALL) malignancy by acting as a pro-survival gene, both in vitro and in vivo. Expression of PRL-3 in a MYC-induced zebrafish T-cell ALL model showed that PRL-3 expressing ALL had shorter latency and higher penetrance, compared with the fish injected with rag2:Myc; rag2:GFP alone. Growth after transplantation at low cell number was also significantly faster in the PRL3 expressing T-ALL, suggesting that PRL-3 plays an important role in ALL development and progression.

We found PRL-3 to be highly expressed in a majority of human ALL compared to normal peripheral blood mononuclear cells (PBMNC), which suggests clinical relevance. Analysis of gene expression microarray data of 119 patient samples reveals that PRL-3 was highly expressed in 58% of the patients. In addition, PRL-3 had the highest expression in the most aggressive and treatment resistant subgroups of T-ALL, the TAL/LMO and Immature subtypes. PRL-3 is also expressed by human T-ALL cell lines using quantitative RT- PCR and immunoblotting. PRL-3 knockdown leads to significant apoptosis in T-ALL cells. The treatment of T-ALL cells using small molecule drugs including 2-cyano-2-ene-ester compound (Analog 3) and thieno [3,2-b] pyridine 1-oxide (thienopyridone) , also resulted in significant growth inhibition of T-ALL cells in part through >4-fold increase in apoptosis, measured by annexin V staining.

In summary, our study provides evidence for a role of PRL-3 in T- ALL malignancy, and suggests that PRL3 may be a useful drug target in the treatment of this disease. Our future studies are focused on identifying the mechanisms by which PRL3 promotes T-ALL progression and survival.

Citation Format: Min Wei, Kristin O'Leary, Jinpeng Liu, Chi Wang, Jessica Blackburn. The phosphatase PRL-3 is a pro-survival oncogene in acute lymphoblastic leukemia [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 2364.
Abstract 2366: The tyrosine phosphatase, SHP-2, participates in regulating PD-L1 expression in non-small cell lung cancer
Authors: Toral Keller , Harris Brent , Creameans Jarrod , Black Penni
Published: August 2018

Immune checkpoint inhibitors, specifically those which target T cell surface receptor Programmed Cell Death-1 (PD-1) and its ligand, Programmed cell death ligand-1 (PD-L1), have demonstrated clinical benefit in patients with non-small cell lung cancer (NSCLC). Currently, tumor expression of PD-L1 serves as the FDA-approved companion diagnostic for response to anti-PD-1/PD-L1 therapy. However, a fraction of patients who express PD-L1 exhibit a response to treatment, and in some cases, patient tumors that do not express PD-L1 demonstrate response to therapy, making this a poor predictive biomarker. Further, the mechanism by which PD-L1 expression is controlled remains under-studied.

In this study, we seek to determine the impact of the protein tyrosine phosphatase, SHP-2, on PD-L1 expression through Jak/STAT- mediated signaling in NSCLC cell lines. SHP-2 can harbor activating mutations which prevent auto-inhibition of the catalytic phosphatase domain, and that may be pro-tumorigenic in some tumors. To understand the impact of SHP-2 activity on PD-L1 expression, we first assessed activation of STAT3 in NSCLC tumor cells after treatment with IFNγ and other mitogens. We then transfected NSCLC cells with wildtype and hyperactive SHP-2 mutants or ablated SHP-2 with siRNA specific for SHP-2 and measured both STAT3 and PD-L1 expression. Cell lines A549 and H661 exhibit low endogenous expression of both SHP-2 and PD-L1, and thus were chosen for transfection with hyperactive SHP-2 mutants (E76K and G503V), which also altered PD-L1 expression by western blot analysis. Quantitative real-time PCR (qPCR) and western blot analysis revealed that siRNA ablation of SHP-2 led to increased PD-L1 expression in the H460 cell line. Cells transfected with the SHP-2 activating mutants will also be assessed alteration of cell proliferation. Finally, using data obtained from The Cancer Genome Atlas project, STAT3 and PD-L1 expression levels are being evaluated in NSCLC tumor samples in which SHP-2 is mutated (E76K or G503V). With these data, we expect to gain a more complete understanding of the role of SHP-2 within the tumor microenvironment and the extent to which it controls PD-L1 expression, leading to new drug targets or combinations of drugs and better predictive biomarkers of response to PD-1 and PD-L1 inhibitors.

Citation Format: Keller Toral, Brent Harris, Jarrod Creameans, Penni Black. The tyrosine phosphatase, SHP-2, participates in regulating PD-L1 expression in non-small cell lung 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 2366.
Abstract 2378: Histone H4 tyr88-phosphorylation mediated by ACK1 promotes castration-resistant prostate cancer by epigenetic reprogramming of androgen receptor
Authors: Mahajan Nupam P. , Mahajan Kiran , Lawrence Nicholas , Lawrence Harshani
Published: August 2018

Androgen receptor (AR) plays a paramount role in the onset and progression of prostate cancer (PC). This very facet underlies androgen deprivation therapy (ADT), a preferred treatment to negate AR transcriptional activity. While ADT provides immediate palliative benefits, it is ineffective long term, as the recalcitrant disease recurs within 2-3 years and progresses to a lethal stage, referred to as the metastatic castration-resistant prostate cancer (mCRPC). The AR gene (AR) is amplified or mutated in >50% of mCRPCs, suggesting that PC cells may have reinvigorated AR transcription as a response to the loss of existing AR activity by ADT. Consequently, resistance to ADT has become one of the most vexing problems in PC therapy. AR antagonists are often ineffective due to their inability to repress the expression of AR or its splice variant, AR-V7. Here, we report that the tyrosine kinase ACK1 (TNK2) phosphorylates histone H4 at tyrosine 88 upstream of the AR transcription start site. The WDR5/MLL2 complex reads the H4-Y88-phosphorylation marks and deposits the transcriptionally activating H3K4-trimethyl marks promoting AR transcription. We developed a new class of ACK1 kinase-specific small molecule inhibitor (R)-9bMS that suppresses ACK1 kinase activity at IC50= 48 nM. Reversal of the pY88-H4 epigenetic marks by the ACK1 inhibitor (R)-9bMS-sensitized naive and enzalutamide-resistant prostate cancer cells and reduced AR and AR-V7 levels to mitigate CRPC tumor growth. Thus, a feedforward ACK1/pY88-H4/WDR5/MLL2/AR epigenetic circuit drives CRPC and is necessary for maintenance of the malignant state.

Citation Format: Nupam P. Mahajan, Kiran Mahajan, Nicholas Lawrence, Harshani Lawrence. Histone H4 tyr88-phosphorylation mediated by ACK1 promotes castration-resistant prostate cancer by epigenetic reprogramming of androgen receptor [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 2378.
Abstract 2383: EGFR expression increases following decline in activity of N-acetylgalactosamine 4-sulfatase (ARSB) and SHP2 (PTPN11) and increases in chondroitin 4-sulfate and JNK in prostate
Authors: Bhattacharyya Sumit , Feferman Leo , Tobacman Joanne K.
Published: August 2018

The enzyme arylsulfatase B (ARSB; N-acetylgalactosamine 4-sulfatase) removes 4-sulfate groups from the non-reducing end of dermatan sulfate and chondroitin 4-sulfate (C4S) and is required for degradation of these sulfated glycosaminoglycans. Decline in ARSB and the resultant increase in C4S was previously reported to inhibit the activity of the enzyme SHP2 (PTPN11), a ubiquitous, non-receptor tyrosine phosphatase, and to lead to transcriptional events mediated by the sustained phosphorylation of phospho-ERK1,2 and phospho-p38. Previous experiments had shown that decline in ARSB was associated with increased expression of the epidermal growth factor receptor (EGFR; Her1/ErbB1) in human prostate malignant tissue and in human prostate cells, but the mechanism for this increase was unknown. The current experiments were performed to determine how decline in ARSB leads to increased expression of EGFR. Experiments were performed in human prostate stem and epithelial cells and normal and malignant human prostate tissues. Measurements included: EGFR promoter activity, chromatin immunoprecipitation assay, and binding studies of chondroitin sulfates with SHP2. Materials used included: SR11302, an inhibitor of c-fos binding with nuclear DNA; c-Jun peptide that acts as a c-Jun mimetic and inhibits c-Jun phosphorylation by JNK; JNK-selective inhibitor peptide that inhibits JNK phosphorylation; SHP2 dominant negative and constitutively active DNA constructs; PHPS1, an inhibitor of SHP2; ARSB siRNA, and galectin-3 siRNA. Increased EGFR promoter activation and EGFR expression followed decline in ARSB, decline in activity of SHP2 (PTPN11), enhanced activity of c-Jun N-terminal kinase (JNK), increased nuclear DNA binding of c-Jun and c-Fos, and EGFR promoter activation. These effects demonstrate an impact of ARSB and chondroitin 4-sulfation on the activation of a critical dual phosphorylation threonine-tyrosine kinase and on the mRNA expression of an important tyrosine kinase receptor in prostate cells. Restoration of ARSB activity may provide a new therapeutic approach to malignancies in which there is activation of EGFR-mediated tyrosine kinase signaling pathways.

Citation Format: Sumit Bhattacharyya, Leo Feferman, Joanne K. Tobacman. EGFR expression increases following decline in activity of N-acetylgalactosamine 4-sulfatase (ARSB) and SHP2 (PTPN11) and increases in chondroitin 4-sulfate and JNK in prostate [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 2383.
Abstract 24: Extracellular HMGA1 promotes tumor invasion and metastasis in breast cancer
Authors: Méndez Olga , Peg Vicente , Salvans Candida , Pujals Mireia , Fernández Yolanda , Abasolo Ibane , Perez Jose , Matres Ana , Tabernero Josep , Cortés Javier , Arribas Joaquín , Villanueva Josep
Published: August 2018

The recent in-depth study of the cancer secretome suggests that a fraction of the intracellular proteome might be playing alternative roles in the extracellular space during tumorigenesis through its unconventional secretion. A proteomic study aimed at evaluating the role of unconventional protein secretion in the context of tumor invasion led us to investigate the high mobility group A1 (HMGA1) chromatin-binding protein whose overexpression has been correlated with high-grade carcinomas and reduced patient-survival. Here we show that HMGA1 is not only oversecreted in invasive breast tumor cells, but its blockage with specific antibodies in the extracellular space impairs tumor cell migration and invasion. The invasive phenotype caused by extracellular HMGA1 (eHMGA1) is dependent on ERK1/2 signaling and a blocking-Ab against eHMGA1 decreases pERK and re-localizes HMGA1 to the nucleus concomitantly to the loss of invasion capacity. Remarkably, pre-treatment of invasive breast tumor cells with an HMGA1-blocking Ab reduces their metastatic ability in an orthotopic breast cancer xenograft model. HMGA1 also shows a cytoplasmic localization in primary human triple-negative breast cancer (TNBC) tumors that developed distant metastasis while displaying a nuclear localization in TNBC tumors that did not progress to metastatic disease. Thus, eHMGA1 could be a novel drug target in aggressive breast cancer and a biomarker predicting the onset of distant metastasis.

Citation Format: Olga Méndez, Vicente Peg, Candida Salvans, Mireia Pujals, Yolanda Fernández, Ibane Abasolo, Jose Perez, Ana Matres, Josep Tabernero, Javier Cortés, Joaquín Arribas, Josep Villanueva. Extracellular HMGA1 promotes tumor invasion and metastasis in breast 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 24.
Abstract 2409: The effect of heme on mitochondrial biogenesis and function of NSCLC cells
Authors: Sohoni Sagar Shashikant , Vidal Chantal , Zhang Li
Published: August 2018

Heme is a central molecule for mitochondrial function and for many processes involved in oxygen utilization. Heme serves as a prosthetic group or a cofactor for a number of oxidative phosphorylation enzymes and other oxygen-utilizing hemoproteins. Heme also directly regulates the synthesis, translocation, and assembly of these enzyme complexes. Most, if not all, human cells can synthesize and uptake heme from the circulation. A number of epidemiologic studies have shown that high heme intake is associated with increased risk of cancer, including lung cancer. To assess the status of heme metabolism in lung cancer cells, we performed a series of experiments in non-small cell lung cancer (NSCLC) cell lines and compared the results with an immortalized normal lung cell line, HBEC30KT. We observed significant increase in rates of heme synthesis and heme uptake in NSCLC cells compared to HBEC30KT. Previous studies carried out in our lab showed intensified mitochondrial respiration and increased levels of heme as well as hemoproteins in NSCLCs. NSCLCs were also found to exhibit significantly high expressions of proteins involved in mitochondrial biogenesis such as SIRT-1, PGC1α, NRF-1, NRF-2, and TFAM. Expression levels of these proteins correlate with oxygen consumption and total heme content in NSCLCs, suggesting a possibility of role of heme in mitochondrial biogenesis. Immunofluoresence studies showed that heme depletion and add-back significantly affected localization and expression levels of these proteins. Studies are currently under way to determine how heme depletion affects the interactions of various transcription factors involved in mitochondrial biogenesis and function.

Citation Format: Sagar Shashikant Sohoni, Chantal Vidal, Li Zhang. The effect of heme on mitochondrial biogenesis and function of NSCLC 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 2409.
Abstract 2419: Lipid droplets promote glioblastoma cell growth through maintaining cholesterol homeostasis mediated by autophagy
Authors: Geng Feng , Cheng Xiang , Cheng Chunming , Chakravarti Arnab , Guo Deliang
Published: August 2018

Lipid droplets (LDs) have been studied as the major storage site of neutral lipids for decades, especially in the progression of metabolic diseases. Recently, the functions of LDs in cancers have begun to gain significant attention. Here, we provide evidence that LDs promote glioblastoma (GBM) cell growth through autophagy mediated maintenance of cholesterol homeostasis. Our data showed that LDs in tumor tissues from GBM patients are engulfed by autophagic vacuoles and lysosomes. Cellular cholesterol deprivation or removal of plasma membrane cholesterol by methyl-β-cyclodextrin (MβCD) activates autophagy to hydrolyze LDs to support GBM cell survival, whereas inhibition of autophagy blocks transportation of cholesterol to plasma membrane, leading to cell death. Fluorescence imaging reveals direct trafficking of LD-stored cholesterol to the plasma membrane through autophagic liberation. Our data demonstrate that LDs serve as cholesterol reservoir for GBM cell growth, suggesting that targeting LD-autophagy-cholesterol homeostasis metabolic pathway represents a potential means to treat GBM and other malignancies.

Citation Format: Feng Geng, Xiang Cheng, Chunming Cheng, Arnab Chakravarti, Deliang Guo. Lipid droplets promote glioblastoma cell growth through maintaining cholesterol homeostasis mediated by autophagy [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 2419.