bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3817: Improved T cell activation bioassays to facilitate the development of bispecific antibodies and engineered T cell immunotherapies
Authors: Stecha Pete , Garvin Denise , Hartnett Jim , Krishnan Gopal B. , Fan Frank , Cong Mei , Cheng Zhi-jie Jey
Published: August 2018

T cells play a critical role in cell-mediated immunity and can mediate long-term, antigen-specific, effector and memory responses. A variety of immunotherapy strategies have been developed recently that are aimed at inducing, strengthening or engineering T cell responses. These have emerged as promising approaches for the treatment of diseases such as cancer and autoimmunity. Current methods used to measure TCR-mediated T cell proliferation and cytokine production rely on primary PBMCs as a source of T cells, which must be stimulated via co-culture with APCs or anti-TCR/CD3 antibodies. These assays are laborious and highly variable due to their reliance on donor primary cells, complex assay protocols and unqualified assay reagents. As a result, these assays are difficult to establish in quality-controlled drug development settings. To overcome this barrier, we developed two reporter-based bioluminescent T cell activation bioassays that can be used for the development of bispecific antibodies and engineered T cell immunotherapies. The assays consist of Jurkat T cells genetically engineered to express luciferase downstream of either NFAT or IL-2 response elements. The T cell activation bioassays reflect the mechanisms of action of biologics designed to induce TCR and/or CD28-mediated T cell activation, as demonstrated using anti-CD3 and/or anti-CD28 antibodies as well as blinatumomab, a bispecific antibody that simultaneously binds CD3 expressed on T cells and CD19 expressed on malignant B cells. The bioassays are pre-qualified according to ICH guidelines and demonstrate assay specificity, precision, accuracy and linearity required for routine use in potency and stability studies. Finally, our data illustrate the use of reporter-based T cell activation bioassays for characterizing and measuring the activity of engineered chimeric antigen receptor T cells.

Citation Format: Pete Stecha, Denise Garvin, Jim Hartnett, Gopal B. Krishnan, Frank Fan, Mei Cong, Zhi-jie Jey Cheng. Improved T cell activation bioassays to facilitate the development of bispecific antibodies and engineered T cell immunotherapies [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 3817.
Abstract 3830: Monoclonal and bispecific antibodies against human insulin-like growth factors in oncology
Authors: Zhao Qi
Published: August 2018

The insulin-like growth factor (IGF) axis plays an important role in cell growth and differentiation. During the past decade, IGF type 1 receptor (IGF-1R) was deemed a promising target for cancer therapy. The major challenge is that abnormal activation of IGF-1R ligands (IGF-I and IGF-II) exists in series of cancers and contributes to the resistance in IGF-1R-targeted therapies. The IGF-I/II actions are mediated through the IGF receptor type 1 (IGF-1R) and the insulin receptor (IR), which are overexpressed in multiple types of tumors. Up to date, we have developed different antibody formats against IGF-II or IGF-I/II, including one fully human IgG m708.5 with picomolar affinity[1], two bispecific antibodies (BsAbs) m660 [2, 3] and m67 [4], and one small-size nanobody s7g1 (15 kDa) (unpublished data). BsAb m67 has exhibited in cynomolgus macaques and high stability in serum. Compared with large-size antibodies, nanobody s7g1 has distinct advantages because of greater and more rapid tissue accumulation. All antibodies can eliminate the efflux of IGFs from tumor tissues and circulating IGFs in the serum. The m708.5 shows dual-specific for IGF-I/IGF-II with the highest affinities. The m708.5 has shown satisfying preclinical activities against a broad spectrum of human cell lines. When tested in neuroblastoma cell lines, m708.5 showed strong antitumor activity alone and in combination with chemotherapeutic agents [5]. Therefore, the anti-IGF-I/II antibodies offers an alternative approach to target both the IGF-1R and IR-A signaling pathways through the inhibition of IGF-I/II. These results support the clinical development of anti-IGF antibodies for solid tumors with potential for synergy with chemotherapy. Acknowledgements: This work was supported by the Science and Technology Development Fund of Macau (FDCT/131/2016/A3). References: 1. Zhao, Q., et al.. Mol Cancer Ther, 2011. 10(9): p. 1677-85. 2. Chen, W., et al.. Mol Cancer Ther, 2012. 11(7): p. 1400-10. 3. Feng, Y., et al.. Mol Cancer Ther, 2009. 8(5): p. 1113-8. 4. Feng, Y., et al.. Exp Mol Pathol, 2014. 97(3): p. 359-67. 5. Zhao, Q., et al.. Int J Cancer, 2015. 137(9): p. 2243-52.

Citation Format: Qi Zhao. Monoclonal and bispecific antibodies against human insulin-like growth factors in oncology [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 3830.
Abstract 3835: Targeting loss of MTAP expression in glioblastoma multiforme with a combination of 2-fluoroadenine and methylthioadenosine
Authors: Tang Baiqing
Published: August 2018

Methylthioadenosine Phosphorylase (MTAP) is a key enzyme in the purine and methionine salvage pathways that convert the polyamine byproduct 5′-deoxy-5′-methylthioadenosine (MTA) into adenine and methylthioribose-1-phosphate. Homozygous deletion of the MTAP gene is observed in 55% of all glioblastoma multiforme (GBM) tumors. Since MTAP is a house-keeping enzyme found in all normal tissues, the lack of MTAP in GBM makes MTAP-loss a potential therapeutic target. In this study, we explore a novel strategy to target MTAP-loss in GBM by combining 2-fluoroadenine (2FA), a highly toxic adenine analogue, with MTA. Because MTAP is expressed in all normal tissues, MTA will be converted to adenine, which will protect against 2FA toxicity. However, in tumor cells, the effects of 2FA will not be ameliorated because they lack MTAP. Here we show that there is undetectable expression of MTAP in A172 and U87 GBM cell lines by both western blot and MTAP enzyme activity assays. In vitro toxicity studies in which cells were cultured with either 2FA alone or 2FA plus MTA revealed that both cell lines were sensitive to 2FA with or without MTA. The LD50 of A172 cells was 110 nM (2FA) and 127 nM (2FA + 5 μM MTA), while for U87 cells it was 397 nM (2FA) and 323 nM (2FA + 5 μM MTA). This is in sharp contrast to MTAP+ NIH3T3 cells in which we observed a 44 fold shift in the ratio of the LD50 concentration in the presence and absence of MTA (160 nM versus 7000 nM). Addition of MTA protected normal tissues against 2FA toxicity in SCID mice. SCID mice subcutaneously injected with either A172 or U87 cells and then treated with 2FA(10 mg/kg) + MTA(50 mg/kg) showed significantly reduced tumor growth

compared to vehicle treated controls. Our results indicated that 2FA+MTA might be effective in treating GBM.

Citation Format: Baiqing Tang. Targeting loss of MTAP expression in glioblastoma multiforme with a combination of 2-fluoroadenine and methylthioadenosine [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 3835.
Abstract 3841: Biologic evaluation of heterocyclic pyrimidines as novel tubulin inhibitors targeting the colchicine binding site
Authors: Arnst Kinsie
Published: August 2018

Interfering with tubulin dynamics is an attractive strategy for cancer therapy, and tubulin-targeting agents are some of the most widely used chemotherapeutic drugs. Current FDA-approved tubulin inhibitors target either the taxane site (e.g., paclitaxel) or the vinca alkaloid site (e.g., vinblastine). However, their clinical efficacy is often limited by the development of multidrug resistance and a narrow therapeutic index. Tubulin inhibitors interacting with the colchicine site have shown potent anticancer activities, are generally less susceptible to multidrug resistance, and can disrupt tumor vasculature, leading to rapid tumor cell apoptosis. Here, we report on the biologic evaluation of heterocyclic pyrimidines as a new class of tubulin inhibitors targeting the colchicine binding site with a significantly improved therapeutic index. One of the most promising compounds of the series, SB157, was able to strongly inhibit tubulin polymerization and induce cytotoxicity in cancer cells in vitro. We have also obtained the high-resolution X-ray crystal structure for SB157, confirming its direct binding to the colchicine binding site in tubulin, and elucidated the detailed molecular interactions. SB157 significantly inhibited tumor growth in an A375 melanoma xenograft model, and the tumors revealed elevated apoptotic activity and extensive disruption of tumor vasculature. Finally, we demonstrated that SB157 significantly overcame clinically relevant multidrug resistance in a paclitaxel-resistant PC-3/TxR prostate cancer xenograft mouse model. Collectively, these studies provide preclinical proof of concept and structural basis to support the continued development of SB157 and optimization of this scaffold as an improved class of tubulin inhibitors for cancer therapy.

Citation Format: Kinsie Arnst. Biologic evaluation of heterocyclic pyrimidines as novel tubulin inhibitors targeting the colchicine binding site [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 3841.
Abstract 3842: Potent antitumor activity of the HSP90 inhibitor SNX-2112 in type 1 papillary renal cell carcinoma
Authors: Pahwa Roma , Giri Abhigya , Sourbier Carole , Thomas Craig , Linehan W Marston , Neckers Len , Srinivasan Ramaprasad
Published: August 2018

Papillary renal-cell carcinoma (pRCC) is the second most common form of kidney cancer, accounting for 15 to 20% of all cases. There are currently no treatment options of proven benefit for patients with advanced pRCC. The type 1 subset of pRCC tumors are associated with gain of chromosome 7 and activating mutations of MET that could lead to aberrant MET pathway activation resulting in cell proliferation and anti-apoptotic signaling. While MET kinase inhibitors are active in some patients with type 1 pRCC, primary and acquired resistance is common, prompting us to investigate possible therapeutic alternatives. Heat shock protein 90 (Hsp90), which stabilizes numerous oncogenic proteins, including MET and several downstream mediators of MET activity, is an important regulator of cell proliferation and survival, and has attracted wide interest as a novel target for cancer therapy. Hsp90 inhibitors were also identified as molecules of interest based on antitumor activity against patient derived type 1 pRCC cell lines following quantitative high throughput screening (qHTS) with small molecule libraries. We hypothesized that inhibition of Hsp90 might provide a novel therapeutic approach for patients with pRCC tumors with MET pathway activation. SNX-2112 was chosen as a candidate Hsp90 inhibitor for further evaluation. Our data demonstrate that inhibition of the Hsp90 pathway is lethal to pRCC cell lines with MET pathway activation, and that treatment with SNX-2112 leads to the degradation of MET/pMET and its downstream signaling intermediates p-AKT, AKT, p-ERK. Furthermore, SNX-2112 leads to inhibition of critical downstream transcriptional targets of AKT and ERK, including VEGF, c-Myc and BIRC5. SNX-2112 has potent anti-tumor activity in vitro, inhibiting proliferation, clonogenicity and 3D-anchorage independent colony formation of MET activated pRCC cell lines. These data demonstrate that Hsp90 inhibition is a promising therapeutic strategy in Type 1 pRCC and is worthy of further preclinical and clinical evaluation.

Citation Format: Roma Pahwa, Abhigya Giri, Carole Sourbier, Craig Thomas, W Marston Linehan, Len Neckers, Ramaprasad Srinivasan. Potent antitumor activity of the HSP90 inhibitor SNX-2112 in type 1 papillary renal cell 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 3842.
Abstract 3862: Empowering research on ubiquitin and ubiquitin-like protein modification cascade using recombinant enzyme systems
Authors: Yao Eric , Lai Shenshen , Yan Jun
Published: August 2018

Protein modifications by the covalent attachment of ubiquitin (Ub) or ubiquitin-like proteins (UBLs) such as SUMO, ISG15 or NEDD8 participate in many critical cellular processes. These include cell-cycle regulation, DNA repair, endocytosis, mitophagy, oncogenesis, post-membrane-protein trafficking, antiviral pathways and most notably, proteasomal degradation of target proteins. Ubiquitination and modification by UBLs share a similar catalytic cascade which requires the sequential action of three classes of enzymes: E1 activating enzymes, E2 conjugating enzymes and E3 ligases. While Ub/UBLs are activated by E1 through an ATP-dependent thioester bond formation and transferred to E2 from E1, the exact mechanism of how they are subsequently attached to a protein substrate depends on the type of E3's containing characteristic catalytic domain - RING, HECT or RBR. It has been generally accepted that E3's mediate substrate specificity and E2's are the main determinants for selection of the lysine to construct the modifier chains, which thereby directly control the cellular fate of the substrate. Recent research has linked dysregulation of the Ub/UBLs modification system to numerous diseases including cancer, immunological disorders and neurodegeneration. The high substrate specificity provided by combinations of over 30 E2's and over 600 E3's in the human genome makes these enzymes emerging drug targets. In response to a growing market demand, SignalChem has developed an extensive array of products encompassing enzymes, Ub/UBL modifiers and substrates in the ubiquitination, SUMOylation, ISGylation and NEDDylation processes. With Promega's AMP-GloTM reagents and an optimized assay protocol, we have identified and validated a variety of functionally competent combinations of these enzyme components. With the established protocol, each enzyme in the catalytic cascade has been assessed for their specific activity towards generation of free AMP. In addition, inhibition profiles of all three enzymes have been obtained using the assay system, further demonstrating their potential to be used in high-throughput screening to identify potential lead compounds for drug discovery and development programs.

Citation Format: Eric Yao, Shenshen Lai, Jun Yan. Empowering research on ubiquitin and ubiquitin-like protein modification cascade using recombinant enzyme systems [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 3862.
Abstract 387: Glutamine metabolism reprogramming in adipocyte-tumor microenvironment promotes triple-negative breast cancer growth
Authors: Liu Yang , Zhao Tingli , Li Zhengzheng , Wang Lai , Yang Mei , Yuan Shengtao , Sun Li
Published: August 2018

OBJECTIVE

Adipocytes are the most abundant stromal partners in breast tissue. However, the crosstalk between breast cancer cells and adipocytes has been given less attention compared to cancer-associated fibroblasts. We try to explain whether cancer associate adipocytes (CAAs) in microenvironment can lead to metabolic remodeling of triple-negative breast cancer, promoted cancer proliferation, migration and the potential mechanism.

METHOD

A Transwell co-culture model in vitro and orthotopic mouse model of human breast tumor formation in vivo were used to study the crosstalk between adipocytes and breast cancer cells, and then using clinical breast cancer sample analysis to study the mechanism of glutamine reprograming.

RESULTS

We find that while cocultivated adipocytes with breast cancer cells, it exhibited enhanced oxygen consumption rate (OCR) and ATP, accompanied by increasing glucose and glutamine uptake. Interestingly, the metabolic way is significantly different between phenotype-specific breast cancer cells. Compared with non-triple-negative breast cancer cells, triple-negative breast cancer cells (MDA-MB-231, MDA-MB-468) are more dependent on glutamine. We show that cancer associate adipocytes (CAAs) promote growth of breast cancer cells through key enzymes such as GLUL/GLS/GDH and transporters such as SLC1A5/SLC38A1/SLC38A2 related to glutamine metabolism in vitro. Using cytokine antibody microarrays, IL-6/IL-8/ leptin/TIPM2 is secreted by CAAs and is required to enhance breast cancer cells malignancy in vitro. Consistently, the correlation of inflammatory cytokines and adipokines with prognosis is analyzed by the Kaplan-Meier plotter, the data suggest that inflammatory cytokines and adipokines is associated with glutamine metabolism in triple-negative breast cancer microenvironment.

CONCLUSIONS

These results suggest that adipocytes promote proliferation of triple-negative breast cancer via metabolism reprogramming potentially by inducing inflammatory cytokines and adipokines upregulation of cancer associate adipocytes (CAAs).

Citation Format: Yang Liu, Tingli Zhao, Zhengzheng Li, Lai Wang, Mei Yang, Shengtao Yuan, Li Sun. Glutamine metabolism reprogramming in adipocyte-tumor microenvironment promotes triple-negative breast cancer growth [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 387.
Abstract 388: p16/MTAP status as a model for investigating gene-environment interactions in metabolism
Authors: Sanderson Sydney M. , Mikhael Peter , Dai Ziwei , Locasale Jason W.
Published: August 2018

While the consequences of tumor suppressor gene deletions have been extensively characterized, little is known about how concurrent deletions of neighboring genes can influence the interaction of cancer cells with the environment. Using a panel of tissue-matched cancer cell lines exhibiting either wild-type or homozygous deletions of the p16 locus, metabolomics analysis showed that concurrent deletion of the proximal gene encoding the methionine salvage enzyme methylthioadenosine phosphorylase (MTAP) is associated with a subtle but reproducible global shift in metabolite levels. However, MTAP status does not appear to be predictive of metabolic responsiveness to restriction of key nutrients that feed into and out of the methionine cycle. Of particular interest is the finding that cell lines exhibit remarkable heterogeneity in responsiveness to nutrient availability, seemingly independent of MTAP status or tissue of origin. Similarly, MTAP-deleted cell lines exhibit variable global metabolic reprogramming in response to acute ectopic expression of MTAP, and maintain their heterogeneous responsiveness to nutrient restriction. These findings illustrate the exceptional metabolic diversity across cancer cell lines despite similar culture conditions, demonstrating the usefulness of examining a variety of cell lines in studies of cancer cell metabolism.

Citation Format: Sydney M. Sanderson, Peter Mikhael, Ziwei Dai, Jason W. Locasale. p16/MTAP status as a model for investigating gene-environment interactions in metabolism [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 388.
Abstract 3886: A large cancer pharmacogenomics combination screen powering crowd-sourced advancement of computational drug synergy predictions
Authors: Dry Jonathan R. , Menden Michael P. , Bulusu Krishna , Guinney Justin , Saez-Rodriguez Julio
Published: August 2018

This abstract has been withheld from publication due to its inclusion in the AACR Annual Meeting 2018 Official Press Program. It will be posted online following its presentation.

Citation Format: Jonathan R. Dry, Michael P. Menden, Krishna Bulusu, Justin Guinney, Julio Saez-Rodriguez. A large cancer pharmacogenomics combination screen powering crowd-sourced advancement of computational drug synergy predictions [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 3886.
Abstract 3894: DPYD-Varifier, a computational model to identify 5-FU toxicity-associated DPYD variants
Authors: Shrestha Shikshya , Zhang Cheng , Jerde Calvin R. , Li Hu , Offer Steven M. , Diasio Robert B.
Published: August 2018

5-Fluorouracil (5-FU) is a cornerstone therapy for the treatment of several aggressive tumors. However, nearly 1/3 of patients receiving 5-FU experience severe (grade 3+) adverse 5-FU toxicity. Genetic variations in dihydropyrimidine dehydrogenase (DPD; DPYD gene) are major contributors of 5-FU toxicity. With continued progress in next-generation sequencing technologies, the number of reported genetic variants is substantially increasing; to date, there are more than 450 reported missense variants in the DPYD gene. Only a handful of those variants, including *2A, I560S, D949V, and rs75017182, have been established as predictors of toxicity. Because most of the remaining variants are unstudied, mainly due to their rare occurrence and unequal racial distribution, it is challenging to conduct statistically powered clinical association studies. Therefore, in vitro and computational methods to identify potential toxicity-associated variants have major appeal. The primary goal of the current study was to develop a highly accurate gene-specific in silico classifier to predict deleterious DPYD variants. For this purpose, using a recombinant cellular model, we characterized 69 missense DPYD variants reported in online databases. We then utilized DPD activity data from the 69 variants in combination with data from 87 previously characterized variants (see Offer, SM et al. Cancer Research 2013 & 2014) to develop the DPYD variant classifier model (DPYD-Varifier). DPD protein-specific biochemical and structural features and amino acid conservation scores were integrated into the model's random forest classifier-based algorithm. DPYD-Varifier showed improved accuracy (85% accurate) compared to other in silico tools. Additionally, the model provided mechanistic insights into alterations of DPD function conferred by DPYD variants. In general, deleterious variants were located in closer proximity to important binding/coordinating sites of the DPD protein. Additionally, DPYD-Varifier allowed us to estimate the cumulative allele frequency of all reported deleterious variants in various geographical/racial groups, which can substantially add to our knowledge of DPD deficiency and 5-FU toxicity observed in these populations. In conclusion, the integration of in vitro and accurate computational data from our study permitted classification of relevant DPYD variants, which could be useful to more effectively individualize 5-FU therapy in the future.

Citation Format: Shikshya Shrestha, Cheng Zhang, Calvin R. Jerde, Hu Li, Steven M. Offer, Robert B. Diasio. DPYD-Varifier, a computational model to identify 5-FU toxicity-associated DPYD variants [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 3894.
Abstract 3896: A big data approach to predicting context specific synergistic drug combinations
Authors: Gilvary Coryandar M. , Madhukar Neel , Elemento Olivier
Published: August 2018

Combination therapies for various cancers have been shown to increase efficacy, lower toxicity and escape resistance. However, systematically interrogating all possible synergistic therapies is experimentally unfeasible due to the sheer volume of possible combinations. Computational approaches have proven to be an invaluable tool within pharmacogenomics and have helped with prioritizing the development of perspective therapeutics as well as matching the right drugs with the right patients. Here we apply a novel big data approach in the evaluation and ultimately the prediction of drug synergy by using the recently released NCI-ALMANAC, the largest publically available synergistic drug efficacy dataset to date. First, to better understand drug combinations, we distinguished between those that were synergistic and adverse and evaluated various drug similarity metrics for all pairs. We found that certain features showed significant differences between adverse and synergistic drug combinations, such as post-treatment transcriptional effects similarity (D = 0.17, p < 0.001) and chemical structure similarity (D = 0.25, p < 0.001). By exploiting these significant similarities and dissimilarities and incorporating cell line specific data we developed a machine learning model to predict context specific drug synergy and achieved significant predictive performance (AUC = 0.823). We find that our model can be used to both identify novel synergistic drug pairs, as well as find novel indications for known drug combinations by identifying new sensitive cell lines. Moreover, the interpretability of our model allows for the interrogation of features for a deeper understanding of why certain combinations are predicted synergistic. In addition to identifying the cancer types and subtypes a combination therapy would be most synergistic within, we set out to identify the molecular indication for highly synergistic pairs. Specifically, we systematically identified candidate predictive biomarkers which could be used to stratify patient cohorts. Overall, our model and methodology can expedite the development and expansion of combination therapeutics, which can help battle acquired resistance and increase therapeutic efficacy. The thorough understanding of specific combination efficacy dependencies allows for a true precision medicine application of these therapeutics.

Citation Format: Coryandar M. Gilvary, Neel Madhukar, Olivier Elemento. A big data approach to predicting context specific synergistic drug combinations [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 3896.
Abstract 3897: Pharmacogenomic landscape of long non-coding RNAs in human cancers
Authors: Nath Aritro , Huang R. Stephanie
Published: August 2018

Extensive efforts in recent large-scale drug screens have enabled identification of protein-coding genes (PCGs) as biomarkers for anti-cancer agent sensitivity, but information on the role of long-non coding RNAs (lncRNAs) in drug response is sparse. Lower abundance, difficulty in expression profiling and limited functional knowledge have so far hindered global analysis of lncRNAs in drug screens. However, lncRNAs represent nearly 60% of the human genome (vs. 3% for PCGs) and contain a vast majority of pharmacogenomic SNPs. Thus, it is critically important to evaluate the role of lncRNAs in drug sensitivity. Here, we perform a comprehensive reanalysis of large-scale drug screens using novel computational methods to create a landscape of pharmacogenomic interactions of lncRNAs in human cancers.

We developed an algorithm to accurately impute the lncRNA transcriptome using PCG expression. This approach allowed us to generate lncRNA transcriptome in high throughput drug screening data sets and enabled systematic evaluation of lncRNAs for hundreds of drugs. For example, we applied our method to the Genomics of Drug Sensitivity in Cancer (GDSC) study, where sensitivity to 265 anti-cancer agents were quantified in 963 cancer cell lines. Sparse regression analysis showed lncRNA transcriptome were similar in accuracy at predicting drug response compared to PCGs (P = 0.9), indicating lncRNAs may be equally important biomarkers.

Subsequently, we identified drug-specific lncRNA biomarkers using ANOVA while adjusting for tissue type. Our analysis recapitulated literature-verified lncRNAs associated with drug response, e.g. HOTAIR expression with cisplatin resistance (FDR < 0.05). In addition, we identified several compelling novel lncRNA associations for specific drugs along with previously uncharacterized multi-drug response predictors.

We next identified lncRNAs that may serve as biomarkers for drug response while controlling for known cancer functional events. As an example, we analyzed EGFR-inhibitors (erlotinib, gefitinib, pelitinib) while controlling for EGFR somatic point mutations and amplification status and identified an anti-sense lncRNA overlapping UGT1A locus that may be a potential regulator of erlotinib metabolism. These findings reveal important lncRNAs that predict response independent of known cancer functional events and are currently being evaluated in non-small cell lung cancer patients.

In conclusion, our comprehensive analysis has generated unprecedented insights into the role of lncRNAs in anti-cancer drug response and will be an invaluable resource for researchers studying drug response in various cancer types.

Citation Format: Aritro Nath, R. Stephanie Huang. Pharmacogenomic landscape of long non-coding RNAs 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 3897.
Abstract 39: LCN2, CD133 expressions in breast cancer spinal bone metastasis and CD151, TWIST1, EPHA2 in lung metastasis: Prognostic value and organ-specific relapse
Authors: Maiti Aparna , Hait Nitai C.
Published: August 2018

Purpose: Breast cancer most often recurs and metastasizes to the distal organs that had their primary tumors surgically excised. Due to heterogeneous nature of the breast cancer, metastasis organotropism has poorly understood. In this study, we assessed the specific cancer-related gene expression changes occurring with metastatic breast cancer recurrence to distant organs comparing with non-metastatic breast cancer. Experimental design: We develop a clinically relevant metastatic syngeneic mouse model using two cell lines 4T1-luc2 that can only metastasize to lung and 4T1.2-luc2 metastasized to bone and lung. After the primary mammary tumor surgically excised from mice, we monitor mice using IVIS imaging for metastatic tumor lesion development in the distant organs followed by MRI scan to detect spinal bone metastases. Animals were sacrificed to collect spines, bones, and lung lesions for molecular analysis. We also validate the gene expression data using the RNA-seq cohort data from the Roswell Park Cancer Institute patient samples with advanced breast cancer bone metastases and matched non-metastatic breast cancer.

Results: Our cell lines, RNA-seq data analysis suggested that there are at least 50 genes which are statistically highly expressed in 4T1.2-luc2 compared to 4T1-luc2 cells. Out of which only a few genes are well established for cancer metastasis biology, include ANGPTL77, SERPINE2, TSPAN11, ESM1, LCN2 which are expressed more than 8 fold in 4T1.2-luc2 compared to 4T1-luc2 cells. To our surprise, we found that most of these genes are differentially expressed when metastasized to distant organ. Our data revealed that LCN2 (7 fold) and CD133 (above 20 fold) are overexpressed in the spine and bone compared to the primary or lung met lesions formed by 4T1.2-luc2. Conversely, LCN2 and CD133 genes are downregulated in breast cancer lung metastasis tissues, whereas CD151, EPHA2, and TWIST1 genes are highly overexpressed in lung metastatic lesion compared to primary, bone or spine. Further, the RNA-seq data of patient samples explained that LCN2, CD133 expressions are significantly higher (8 out of 10 patients) in advanced breast cancer bone metastases compared with matching non-metastatic breast cancer. Conclusion: Our data suggested that LCN2, CD133 would be a prognostic marker for breast cancer spinal bone metastasis, whereas CD151, TWIST1, EPHA2 would be a prognostic marker for lung metastasis and organ-specific relapse.

Citation Format: Aparna Maiti, Nitai C. Hait. LCN2, CD133 expressions in breast cancer spinal bone metastasis and CD151, TWIST1, EPHA2 in lung metastasis: Prognostic value and organ-specific relapse [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 39.
Abstract 3908: miR-3132 induces TRAIL and cell death in mutant p53-expressing cancer cells
Authors: Lulla Amriti R. , Einarson Margret B. , Zhou Yan , Lev Avital , Dicker David T. , El-Deiry Wafik S.
Published: August 2018

Tumor suppressor p53, guardian of the genome, is frequently mutated or functionally dysregulated in more than 50% of human tumors. p53 mutation is a later event in tumorigenesis and a number of p53 mutants have “Gain of Function” (GOF) properties that have been shown to promote invasive and more aggressive phenotypes in cancer cells. Mutant p53 has been an attractive and promising therapeutic target for advanced stages of tumors. Further understanding the GOF properties of mutant p53 may help with design of novel strategies to target mutant p53 and/or key GOF pathways. We developed a novel functional high throughput screening (HTS) assay to identify miRNAs that selectively target GOF in mutant p53-expressing cell lines. Of the 2754 miRNA mimics screened, we identified 56 miRNAs that selectively reduced cell viability in TP53 R175H (mutant) cells. The current study characterizes the role of our lead mimic, miR-3132 in mutant p53 cells. We found that expression of miR-3132 is lost in cancer cells and that miR-3132 expression is correlated to p53 status, with mutant p53 cells expressing lower levels of miR-3132 than those with wild-type p53. Restoring expression of miR-3132 via mimics induces anti-proliferative and pro-apoptotic effects in a broad panel of colorectal (CRC), breast and lung cancer cell lines. In addition, miR-3132 induces TRAIL-dependent cell death in mutant p53-expressing cells. All cell lines show potent upregulation of cleaved caspases -8, -9 and -3, at 72 hrs post transfection with miR-3132. DNA damaging agents and serum starvation endogenously increase mature miR-3132 levels in wild-type p53 cells. However, this increase is blunted in mutant p53 cells, indicating that p53 could have a role in modulating either the transcriptional or post-transcriptional levels of miR-3132. In summary, we have identified miR-3132 as a TRAIL and cell death-inducing miRNA whose regulation is dependent on p53. Our ongoing work aims at finding the key targets of miR-3132 that explain the cellular response, TRAIL induction and subsequent cell death. Our long-term goal is to develop miR-3132 as a novel single or combinatorial anti-cancer agent for mutant p53 cancer cells.

Citation Format: Amriti R. Lulla, Margret B. Einarson, Yan Zhou, Avital Lev, David T. Dicker, Wafik S. El-Deiry. miR-3132 induces TRAIL and cell death in mutant p53-expressing 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 3908.
Abstract 3920: nfP2X7 mediates survival of cancer cells in the tumor microenvironment
Authors: Gilbert Simon Mark , Oliphant Christopher John , Peille Anne-Lise , Bronsert Peter , Virgilio Francesco Di , McNulty Shaun , Lara Romain
Published: August 2018

The purpose of this study was to demonstrate the effect of the tumor microenvironment (TME) on expression of nfP2X7, a cancer specific form of P2X7 involved in cancer cell survival. P2X7 is an ATP-gated receptor, involved in inflammation, cell death, cell proliferation and cell migration. Alterations in these cellular processes are associated with the development of cancer. P2X7 is characterized by its capacity to open a large molecular weight pore in response to prolonged ATP activation. A mechanism that triggers membrane depolarisation and cell death. The TME contains high concentrations of ATP sufficient to cause opening of the P2X7 pore, while ATP is almost undetectable in healthy tissue. Secretion by cancer cells and release of intracellular ATP from dying cells are thought to be the main source of extracellular ATP in the TME. Non-functional P2X7 (nfP2X7) is a cancer-specific receptor in which formation of the large molecular weight pore, has been abrogated. Acquisition of nfP2X7 enables tumor cells to survive in the presence of high ATP concentrations. Using antibodies raised against nfP2X7, we assessed the expression of the receptor in a panel of cancer types and analysed the effect of the high ATP concentrations on nfP2X7 expression. We further analyzed the effect of siRNA mediated nfP2X7 depletion on cancer cell lines' viability. Our data shows that nfP2X7 is broadly expressed at the surface of many cancer types and supports cancer cell survival. Furthermore, we demonstrate that exposure to a high ATP concentration, equivalent to those in the TME, drives nfP2X7 expression. These data demonstrate that nfP2X7 is a cancer therapeutic target, whose expression is driven by the TME.

Citation Format: Simon Mark Gilbert, Christopher John Oliphant, Anne-Lise Peille, Peter Bronsert, Francesco Di Virgilio, Shaun McNulty, Romain Lara. nfP2X7 mediates survival of cancer cells in the tumor microenvironment [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 3920.
Abstract 3922: Ganetespib demonstrates strong anti-tumor effect in acquired EGFR-TKI resistance NSCLC cells
Authors: Kurihara Eisuke , Shien Kazuhiko , Torigoe Hidejiro , Takahashi Yuta , Ogoshi Yusuke , Yoshioka Takahiro , Namba Kei , Sato Hiroki , Yamamoto Hiromasa , Soh Junichi , Toyooka Shinichi
Published: August 2018

Purpose: Epidermal growth factor receptor (EGFR) -tyrosine kinase inhibitors (TKIs) are the standard first-line treatment of advanced non-small cell lung cancer (NCSLC). However, almost all patients eventually acquire resistance to EGFR-TKIs, and novel therapeutic strategies to overcome the acquired resistance have been required. Heat-shock protein 90 (Hsp90) is a chaperon protein involved in folding and stabilization of client proteins essential for cancer cell growth and survival. Ganetespib (STA-9090) is one of the second-generation Hsp90 inhibitors with strong anti-tumor effect on NSCLC cells. In this study, we evaluated the anti-tumor effect of ganetespib in EGFR-TKI sensitive and acquired resistance NSCLC cell lines.

Materials and Methods: We treated 4 EGFR-mutant NSCLC cell lines (HCC827, HCC4006, PC-9 and HCC4011), and 14 experimentally established EGFR-TKIs (gefitinib or afatinib) resistance cell lines with ganetespib. The EGFR-TKI resistance mechanism consisted of EGFR T790M second mutation, MET amplification, epithelial-to-mesenchymal transition (EMT) and cancer stem cell-like features. We determined cell proliferation by MTS assay and calculated the IC50 values. We also performed Western blotting to investigate downstream signaling pathways.

Results: The IC50 values in parental NSCLC cell lines ranged from 1.3nM to 15nM, and those in acquired EGFR-TKI resistant cell lines ranged from 0.87nM to 25nM, which suggests strong anti-tumor effect of ganetespib. In addition, this effect was observed regardless of the resistant mechanisms, including EMT. Ganetespib suppressed EGFR activation in EGFR-TKI resistance cells harboring EGFR T790M second mutation. Also, ganetespib suppressed MET activation in EGFR-TKI resistance cells harboring MET amplification.

Conclusion: Ganetespib showed strong anti-tumor effect in acquired EGFR-TKI resistance NSCLC cells regardless of the resistant mechanisms, suggesting that ganetespib could be a promising therapeutic option in the treatment of NSCLC with acquired EGFR-TKI resistance.

Citation Format: Eisuke Kurihara, Kazuhiko Shien, Hidejiro Torigoe, Yuta Takahashi, Yusuke Ogoshi, Takahiro Yoshioka, Kei Namba, Hiroki Sato, Hiromasa Yamamoto, Junichi Soh, Shinichi Toyooka. Ganetespib demonstrates strong anti-tumor effect in acquired EGFR-TKI resistance 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 3922.
Abstract 3927: Clinicopathologic features and immune microenvironment of non-small cell lung cancer with primary resistance to epidermal growth factor receptor-tyrosine kinase inhibitors
Authors: Takashima Yuta , Sakakibara-Konishi Jun , Hatanaka Yutaka , Hatanaka Kanako C. , Ohhara Yoshihito , Oizumi Satoshi , Hida Yasuhiro , Kaga Kichizo , Kinoshita Ichiro , Dosaka-Akita Hirotoshi , Matsuno Yoshihiro , Nishimura Masaharu
Published: August 2018

Background: Approximately 20-30% of non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) activating mutations are not responsive to EGFR-tyrosine kinase inhibitors (TKIs). Although, primary resistance to EGFR-TKI is attributed to various genetic alterations, little is known about the clinical and immunopathological features of patients with primary resistance. The tumor immune microenvironment including tumor infiltrating lymphocytes (TILs) and programmed death ligand 1 (PD-L1) has been reported to play an important role in tumor progression in NSCLC. However, few studies have directly focused on the relationship between the tumor immune microenvironment and primary resistance to EGFR-TKI.

Materials and Methods: Characteristics of 124 NSCLC patients with EGFR mutations who received EGFR-TKI were analyzed. Primary resistance was defined as disease progression within 3 months after EGFR-TKI treatment. Tumor specimens obtained before EGFR-TKI treatment were assessed for the density of TILs expressing CD4 or CD8, and for the expression rate of PD-L1 on tumor cells and tumor-infiltrating immune cells, immunohistochemically.

Results: Primary resistance was observed in 13.7% (17/124) of patients. Smoking Tobacco correlated significantly with primary resistance compared to non-primary resistance. Lower density of total TILs and negative PD-L1 expression as per immunohistochemical analysis correlated significantly with primary resistance. Moreover, immune ignorant phenotype of tumor microenvironment, negative PD-L1 expression with low TIL density, was significantly observed in primary resistance.

Conclusions: Smoking and immune ignorance in the tumor microenvironment might result in primary resistance to EGFR-TKIs.

Citation Format: Yuta Takashima, Jun Sakakibara-Konishi, Yutaka Hatanaka, Kanako C. Hatanaka, Yoshihito Ohhara, Satoshi Oizumi, Yasuhiro Hida, Kichizo Kaga, Ichiro Kinoshita, Hirotoshi Dosaka-Akita, Yoshihiro Matsuno, Masaharu Nishimura. Clinicopathologic features and immune microenvironment of non-small cell lung cancer with primary resistance to epidermal growth factor receptor-tyrosine kinase inhibitors [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 3927.
Abstract 3937: Differential alteration of IL-8 in liver cancer stem cell enrichment in response to PI3K/Akt/mTOR inhibitors and sorafenib
Authors: Kahraman Deniz C. , Kahraman Tamer , Atalay Rengul Cetin
Published: August 2018

Liver cancer stem cells (LCSCs) are derived from damaged and transformed hepatic progenitor cells (HPCs) during precancerous cirrhosis stage. Ras/Raf/MAPK and PI3K/AKT/mTOR signaling pathways are significantly deregulated in liver cancer. The activation of PI3K/AKT/mTOR pathway in LCSC population is one of the reasons for acquired resistance to sorafenib in advanced hepatocellular carcinoma (HCC) patients. Therefore, identifying novel inhibitors targeting this pathway acting on LCSCs is highly essential. We, therefore, elucidated the bioactivities of small-molecule kinase inhibitors on LCSCs acting through PI3K/Akt/mTOR pathway in comparison with DAPT (CSC inhibitor), DNA intercalators and sorafenib. For this purpose, CD133+/EpCAM+ cells originated from HCC cells were analyzed by flow cytometry and effective inhibitors on LCSCs were further tested for their potential combinatorial effects. Treatment of cells with sorafenib and DNA intercalators resulted in enrichment of CD133+/EpCAM+ cells. Yet, rapamycin, LY294002 and DAPT significantly reduced CD133/EpCAM positivity. Combination studies revealed that sequential treatment strategy decreased the ratio of LCSCs as opposed to sorafenib treatment alone. The effects of the inhibitors were also demonstrated with LCSC sphere formation. Additionally, a large panel of genes involved in cancer pathways was analyzed using Nanostring® nCounter® Technology to identify the differentially expressed genes in rapamycin, sorafenib or DAPT treated cells. Systematic pathway analysis using Cytoscape ScoreFlow algorithm application allowed us to identify differential response genes involved in stemness. While the stemness genes were downregulated in cells treated with rapamycin or DAPT, JAG1 gene was found to be upregulated in sorafenib-treated cells. Interleukin 8 (IL-8) was downregulated upon treatment with DAPT, yet upregulated upon sorafenib treatment. Following IL-8 inhibition with repaxirin, CD133/EpCAM positivity of cells decreased significantly, indicating that IL-8 signaling is crucial for the conservation of stemness features of cancer cells. The combination of sorafenib and reparixin resulted in lower levels of SOX2 and Nanog proteins. Conclusion: PI3K/Akt/mTOR pathway inhibitors alter hepatic CSC composition and gene expression in favor or to the detriment of cancer stem cell survival. Blockade of IL-8 signaling provides a promising therapeutic approach for prevention of LCSC enrichment.

Citation Format: Deniz C. Kahraman, Tamer Kahraman, Rengul Cetin Atalay. Differential alteration of IL-8 in liver cancer stem cell enrichment in response to PI3K/Akt/mTOR inhibitors and sorafenib [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 3937.