bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 39: Discovery of PEN-221, an SSTR2-targeting maytansinoid conjugate with potent activity in vitro and in vivo
Authors: White Brian H. , Bazinet Patrick , Whalen Kerry , DuPont Michelle , Quinn James M. , Alargova Rossitza , Yeung Tsun Au , Brockman Adam , Gifford James , Oller Haley , Kriksciukaite Kristina , Lemelin Charles-Andre , Soo Patrick Lim , Moreau Benoit , Perino Samantha , Sharma Gitanjali , Shinde Rajesh , Sweryda-Krawiec Beata , Simcox Mary , Wooster Richard , Bilodeau Mark T.
Published: July 2017

Here we describe the discovery and the structure of PEN-221, a somatostatin receptor 2 (SSTR2) targeting peptide conjugated to DM1. PEN-221 is the first clinical compound from Tarveda’s Pentarin platform, which utilizes miniaturized drug conjugates that diffuse rapidly and deeply into solid tumors. Antibody drug conjugates (ADCs) have garnered a significant amount of attention in their ability to direct cytotoxic drugs to cancer cells; however, the efficacy of ADCs in solid tumors is limited by the slow diffusion of such large molecules through solid tumor tissue. Pentarins are designed to improve the efficacy of targeted therapies through effective tumor cell targeting and enhanced tumor penetration.

SSTR2, a GPCR overexpressed in multiple types of neuroendocrine tumors, including small cell lung cancers, internalizes rapidly upon agonist stimulation, making it an ideal vector for delivering cytotoxic payloads. Examination of a variety of SSTR2 targeting ligands, as well as several potential conjugation sites, led to the identification of the C-terminal side chain of [Tyr3]-octreotate amide as the best conjugation site for a lipophilic payload. The use of DM1 as a payload afforded superior receptor affinity and receptor internalization when compared to other similarly potent microtubule-targeting agents. In vitro studies show that PEN-221 has receptor-dependent cytotoxic effects, and preclinical studies demonstrate PEN-221 induces tumor regression in several SSTR2 expressing xenograft models.

Citation Format: Brian H. White, Patrick Bazinet, Kerry Whalen, Michelle DuPont, James M. Quinn, Rossitza Alargova, Tsun Au Yeung, Adam Brockman, James Gifford, Haley Oller, Kristina Kriksciukaite, Charles-Andre Lemelin, Patrick Lim Soo, Benoit Moreau, Samantha Perino, Gitanjali Sharma, Rajesh Shinde, Beata Sweryda-Krawiec, Mary Simcox, Richard Wooster, Mark T. Bilodeau. Discovery of PEN-221, an SSTR2-targeting maytansinoid conjugate with potent activity in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 39. doi:10.1158/1538-7445.AM2017-39
Abstract 390: Identification and characterization of survival-associated genomic features across tumor types
Authors: Smith Joan , Lin Ann , Giuliano Chris , Sheltzer Jason M.
Published: July 2017

The basic biology that differentiates a primary cell from a transformed cell has been elucidated over the past several decades. Mutations in a limited number of oncogenes and tumor suppressors free the cell from growth-inhibitory checkpoints and allow proliferation in the absence of external stimuli, leading to tumor formation. Yet, primary tumors themselves are rarely deadly: greater than 90% of cancer mortality results from metastasis. What drives metastasis, and, more broadly, what distinguishes a lethal cancer from one with a favorable prognosis, is poorly understood.

To address this question, we have performed a comprehensive meta-analysis on genomic data from primary tumors that are linked to patients’ clinical outcomes. Utiliizing data from >20,000 patients, we have identified protein-coding genes, lncRNAs, miRNAs, methylation sites, and CNVs in primary tumors that are significantly associated with patient prognosis across cancer types. Multivariate analysis reveals a hierarchy of survival determinants: the strongest mortality-associated factors are enriched for components of the mitotic cell cycle, while secondary clusters of genes are involved in extracellular matrix remodeling, cell motility, and angiogenesis. Survival-associated genomic features are indicative of immune infiltration into primary tumors and oxidative phosphorylation activity.

In vitro and in vivo analyses reveal that mortality-associated genes rarely promote the direct transformation of primary cells. Instead, CRISPR mutagenesis reveals that these genes largely represent tumor dependencies, and in their absence cancer cells fail to proliferate. Through this analysis, we have identified new genetic dependencies common across cancer cells, including C16ORF59 and C5ORF46. In total, our results represent the largest assessment of genomic features linked to cancer prognosis completed to date, and offer several lines of insight into the biological differences between fatal and benign cancers.

Note: This abstract was not presented at the meeting.

Citation Format: Joan Smith, Ann Lin, Chris Giuliano, Jason M. Sheltzer. Identification and characterization of survival-associated genomic features across tumor types [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 390. doi:10.1158/1538-7445.AM2017-390
Abstract 3904: Inhibition of the ERK5 pathway as a novel approach to target human chronic myeloid leukemia stem cells
Authors: Tusa Ignazia , Cheloni Giulia , Gray Nathanael , Gozzini Antonella , Sbarba Persio Dello , Rovida Elisabetta
Published: July 2017

Tyrosine kinase inhibitors (TKi) targeting BCR/ABL are very effective for the treatment of Chronic Myeloid Leukaemia (CML). However, resistance mechanisms or their inefficacy on CML leukaemia stem cells (LSC) may lead to relapse. Therefore, there is urgent need to identify new molecular targets. The Extracellular signal-Regulated Kinase 5 (ERK5) is a Mitogen-Activated Protein Kinase involved in cancer. Our aim was to study a possible role of the ERK5 pathway in CML LSC.

Cells used were: human CML cell lines K562 and KCL22 that express constitutively active ERK5; CML patient-derived cells and CD34+ cells from heathy donors (after informed consent had been obtained). Cells were incubated in normoxic (routine) or hypoxic (0.2% O2) primary cultures (LC1) in the presence or the absence of drugs. Day-7 LC1 cells were transferred to drug-free, non-selective normoxic secondary cultures (LC2), to measure LC2 repopulation as a read-out of progenitor/stem cell potential (CRA assay). Compounds: XMD8-92 (ERK5 inhibitor) and BIX02189 (MEK5 inhibitor); imatinib (BCR/ABL inhibitor, IM).

We previously showed that stem cell potential of CML LSC is maintained in severe hypoxia. In K562 and KCL22 cells and in primary cells derived from 9 CML patients, the treatment in hypoxic LC1 with XMD8-92 or BIX02189, but not with IM, impaired progenitor/stem cell potential. The same results were obtained by combined treatment of XMD892 with IM. Importantly, XMD8-92 did not affect progenitor/stem cell potential of CD34+ cells from heathy donors. In colony formation ability assays ERK5 inhibition decreased colony formation by human primary CML cells to a higher extent than that by normal human CD34+ hematopoietic cells. Interestingly, in hypoxia, combined treatment XMD8-92/IM decreased the expression of genes relevant for stem cell maintenance such as p21, nanog and c-myc and the expression of CD26, a CML LSC marker. Moreover, combined XMD8-

92/IM maintained low the expression of p27, another gene involved in stem cell maintenance, that is increased by either drug when administered alone.

This indicate that the ERK5 pathway inhibitors impaired LSC maintenance of CML cell lines and primary CML cells.

Citation Format: Ignazia Tusa, Giulia Cheloni, Nathanael Gray, Antonella Gozzini, Persio Dello Sbarba, Elisabetta Rovida. Inhibition of the ERK5 pathway as a novel approach to target human chronic myeloid leukemia stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3904. doi:10.1158/1538-7445.AM2017-3904
Abstract 391: A comparative genomics approach to understanding the control of cell context dependent P53 binding
Authors: Sundaresan Varsha , Li Ying , DiCiaccio Benedetto , Lin Victor T. , Zhou Lei
Published: July 2017

Cancer is considered to be a genetic disease characterized by sequential accumulation of mutations. Recent studies have provided evidence that epigenetic changes and regulatory sequence mutations could also dysregulate oncogenes and tumor suppressors. As a transcription factor, P53 is activated in response to oncogenic stress and exerts distinct anti-proliferative functions based on the stressor and cell type. Though a number of ChIP-Seq studies have identified thousands of P53 binding sites in mammalian genomes, the functionality of these binding sites remains to be established. In addition, we know little about what controls stress and cell context specific binding profile of P53.

Traditionally, mutations in the coding regions of p53 have been extensively studied to gain insights on its role in cancer and to identify strategies to restore the functions of p53 in cancer cells. We hypothesize that mutations in or epigenetic silencing of functionally important P53 binding sites play an important role in tumorigenesis as well. Since functional regulatory regions tend to be more evolutionarily conserved, in this project we propose using a comparative genomics approach to identify functional P53 binding sites and determine if these regions are involved in tumorigenesis.

In order to study how P53 binding following DNA damage differs between cell types, we aim to perform comparison between P53 ChIP-Seq data generated in our lab from Drosophila embryos at different developmental stages and a cell line (Kc167) as well as published datasets from mammalian stem cells and differentiated cells. Differential expression analysis using RNA-Seq exhibited that at an early stem cell-like stage there is P53-dependent induction of pro-apoptotic genes in response to DNA damage but not in the differentiated stages. We are seeking to establish functionally significant P53 binding sites by combining ChIP-Seq data and gene expression data obtained using RNA-Seq. The importance of these binding sites will be verified by CRISPR-Cas9 -mediated genome editing. We will also perform comparison studies of DNA damage-induced P53 binding in human, mouse and Drosophila to identify analogous patterns.

Knowledge gained from this study will help us to understand the role of non-coding regulatory regions in tumorigenesis, and predict patient response to apoptosis inducing therapeutic agents. It may also lead to novel strategies to restore cellular sensitivity to chemotherapy or radiation.

Citation Format: Varsha Sundaresan, Ying Li, Benedetto DiCiaccio, Victor T. Lin, Lei Zhou. A comparative genomics approach to understanding the control of cell context dependent P53 binding [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 391. doi:10.1158/1538-7445.AM2017-391
Abstract 392: Application of somatic mutation-based expression profiles for high-throughput phenotyping in cancer
Authors: Varn Frederick S. , Wang Yue , Cheng Chao
Published: July 2017

Somatic mutations have been heavily implicated in initiating and driving cancer growth and metastasis. These alterations serve as upstream events that have broad transcriptional effects by altering protein structure and function and interfering with signaling pathways. Here, we apply an iterative approach that utilizes patient gene expression profiles to define mutation-associated transcriptomic signatures. We use these signatures to quantify the level at which a given somatic mutation affects its gene’s downstream activities. Using TP53 in breast cancer as an example, we show that while TP53 mutations are the primary predictor of altered p53 activity, events such as copy number alterations and methylation changes may represent alternative routes to high p53 disruption. Conversely, we identify a class of TP53 mutations that have minimal downstream transcriptomic effects. By expanding this approach to other genes and tumor types, we show that mutation signatures derived from different tissue types exhibit varying levels of conservation, suggesting a degree of tissue-specificity in the downstream effects of somatic mutations. To provide functional context to our results, we examine the associations between different mutation signatures and various cancer-related features, including patient survival, immune infiltration, and pathway activity. Together, our results indicate the utility of using patient gene expression profiles to study mutation function. Going forward, these results may be useful in functionally annotating variants of unknown significance and broadening the potential of precision medicine-based therapeutic approaches.

Citation Format: Frederick S. Varn, Yue Wang, Chao Cheng. Application of somatic mutation-based expression profiles for high-throughput phenotyping in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 392. doi:10.1158/1538-7445.AM2017-392
Abstract 3920: Tumor volume as a predictor of lymph node metastases in prostate cancer
Authors: Molokwu Chidi N. , Yuen Keith , Addla Sanjay , Singh Rajindra , Chahal Rohit
Published: July 2017

Introduction: Pelvic lymph node dissection (PLND) is recommended during radical prostatectomy (RP) in patients with high risk prostate cancer (PCa). Guidelines recommend extended PLND in patients who have >5% probability of lymph node involvement, calculated from pre-operative parameters. PLND increases procedure time and potential morbidity to patients. Tumour size has been reported to be predictive of biochemical recurrence after RP. We analysed the utility of several parameters including tumour size in predicting lymph node metastases in order to further refine the population who require PLND during RP.

Materials & Methods: We maintained a prospective database of Robotic Assisted Radical Prostatectomy (RARP) from July 2012 to August 2016 consisting of a cohort of 531 patients. Patients with >5% calculated probability of lymph node metastasis had PLND during RARP. We performed multiple linear regression using age, PSA, D’Amico risk score, ISUP prognostic group, clinical T-stage, radiological T-stage, tumour volume, tumour volume relative to prostate size (relative tumour volume), and number of nodes excised during PLND to determine if any of these parameters could predict the presence of lymph node metastases.

Results: Of the cohort of 531 patients, 154 had PLND during RARP. Of the PLND cohort, 15 (10%) had pN1 disease. Tumour volume (r2 0.044, p= 0.0098), relative tumour volume (r2 0.033, p=0.026) and number of lymph nodes excised (r2 0.046, p=0.0075) were independent predictors of pN1 disease. D’Amico risk score (r2 0.019, p=0.089) approached but did not reach statistical significance. Patients with pN1 disease all had primary tumours ≥6cc.

Conclusions: In our cohort of PLND patients, tumour volume was an independent risk factor for pN1 disease. With the increasing availability of multi-parametric MRI, tumour volume can be measured pre-operatively as an adjunct to PIRADS scoring. The role of tumour volume and relative tumour volume in predicting lymph node metastasis in conjunction with other pre-operative parameters requires further investigation. The use of tumour volume in determining the need for PLND in patients undergoing RP may help to further refine the population at highest risk for pN1 disease and avoid the additional morbidity of PLND in those at low risk. In addition, our findings support the recommendation of performing an extended PLND to maximise regional lymph node clearance.

Citation Format: Chidi N. Molokwu, Keith Yuen, Sanjay Addla, Rajindra Singh, Rohit Chahal. Tumor volume as a predictor of lymph node metastases in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3920. doi:10.1158/1538-7445.AM2017-3920
Abstract 3926: Is FGD3 a potentially prognostic marker for breast cancer
Authors: Sun Yuliang , Willis Scooter , Lin Xiaoqian , Achua Justin , Williams Casey , Leyland-Jones Brian
Published: July 2017

Background: Prognostic factors are capable of providing information on clinical outcomes at the time of diagnosis; they are usually indicators of growth, invasion, and metastatic potential (Gasparini G. et al. 1993; Hayes DF. et al. 1998). Tissue marker is one of the prognostic factors; to date, only a small proportion of markers are ultimately clinically useful, including Ki-67 and HER2. Faciogenital dysplasia 3 protein (FGD3), a putative regulator of cell morphology and motility, has been shown to regulate cell migration. In a study of 3,256 tumors, low expression of FGD3 mRNA indicates poor outcome (Hayakawa M. et al. 2008; Scooter W. et al. 2014). However, an immunohistochemistry (IHC) study to evaluate FGD3 protein expression has not been done. We hypothesize that the expression levels of FGD3 protein by IHC might improve the prediction of patient outcomes, and FGD3 might be a potentially prognostic marker of breast cancer.

Materials and Methods: 322 cases of breast cancer Tissue Microarrays (TMA) (BR1504a, BR1505b, HBre-Duc068Bch-01, and BR20837) were purchased from US Biomax, Inc (Rockville, MD). Rabbit polyclonal antibody against FGD3 was purchased from Novus Biologicals, LLC (Littleton, CO). Tissue cores were stained for FGD3 with Dako’s EnVision + Dual Link System. Image acquisition was performed using an Olympus camera and software. FGD3 protein expression by IHC was quantitatively determined in the range of 0-4. Unpaired t-test was used for data analyses.

Results: 1) Benign tumor and breast adenocarcinomas in lower stages showed strong expression of FGD3, whereas the breast adenocarcinomas in higher stages showed mild ~ weak expression. 2) Invasive breast cancer in stage IIA showed strong FGD3 expression compared to matched metastatic carcinoma which showed mild expression of FGD3. 3) FGD3 protein levels for tumors (n=135) with N0 indicating no regional lymph node metastasis were compared with tumors with lymph node metastasis (N1-3; n=98) and corresponding matched metastatic tissue (n=103). An unpaired t-test comparing N0 vs. N1-3 showed lymph node metastasis is associated with lower FGD3 protein levels (p<0.0001).

Conclusion: FGD3 protein expression levels within breast tumors were different according to metastatic status. Our IHC results suggest the possibility of FGD3 to be a prognostic marker in patients with breast cancer.

Citation Format: Yuliang Sun, Scooter Willis, Xiaoqian Lin, Justin Achua, Casey Williams, Brian Leyland-Jones. Is FGD3 a potentially prognostic marker for breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3926. doi:10.1158/1538-7445.AM2017-3926
Abstract 3927: Programmed cell death ligand 1 expression in resected colorectal adenocarcinomas: association with micrometastasis
Authors: Hwang Ilseon , Park Keon Uk , Kim Jin Young , Ryoo Hun-Mo , Lee Yun-Han
Published: July 2017

Background: Programmed cell death 1 (PD-1) and its ligand (PD-L1) are key suppressors of the cytotoxic immune response. PD-L1 expression on tumor cells may be induced by the immune microenvironment, resulting in immune escape, and an adverse prognosis in many malignancies. In colorectal carcinoma the response to PD-1/PD-L1 inhibition is correlated with microsatellite instability. However, little is known about the clinicopathologic, molecular, and prognostic characteristics of colorectal carcinoma with PD-L1 expression.

In surgically resected colorectal adenocarcinoma, micrometastasis should be crucial for recurrence, and micrometastasis may be related to PD-L1. The aim of this study is to assess the PD-L1 expression and its association with clinicopathologic manifestations.

Methods: PD-L1 expression was evaluated in 176 resected colorectal adenocarcinomas using tissue microarrays. Immunohistochemical staining was performed to evaluate the expression of PD-L1. The relationship of clinicopathologic manifestations and PD-L1 expression in colorectal cancer were evaluated by chi-squared test, Kaplan-Meier survival and Cox regression test.

Results: High PD-L1 expression was present in 52.8% colorectal adenocarcinoma and was not related pathologic T or N stage. High PD-L1 expression was associated with decreased recurrence rate (p < 0.001), better disease-free survival (p < 0.001). Cox regression analysis revealed that pathologic N stage and High PD-L1 expression was an independent prognostic factors of disease-free survival (p < 0.001).

Conclusion: High PD-L1 expression is an independent prognostic factor, such as pathologic stage in colorectal adenocarcinoma. PD-L1 expression is independent prognostic factor, relating immune response to micrometastasis and immune suppression by PD-L1 may not be effective in micrometastasis of colorectal adenocarcinoma.

Key words: PD-L1, colon, cancer, immunology, micrometastasis

Citation Format: Ilseon Hwang, Keon Uk Park, Jin Young Kim, Hun-Mo Ryoo, Yun-Han Lee. Programmed cell death ligand 1 expression in resected colorectal adenocarcinomas: association with micrometastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3927. doi:10.1158/1538-7445.AM2017-3927
Abstract 393: Predicting DNA accessibility in the pan-cancer tumor genome using RNA-Seq, WGS, and deep learning
Authors: Wnuk Kamil , Sudol Jeremi , Rabizadeh Shahrooz , Soon-Shiong Patrick , Szeto Christopher , Vaske Charles
Published: July 2017

DNA accessibility, chromatin regulation, and genome methylation are key drivers of cancer transcription. However, there is much left to be understood about the functional implications of sequence-level data to the regulation of gene expression, especially when it comes to the noncoding genome. Recently [Kelley, D., Snoek, J., and Rinn, J., Genome Res. 2016] trained neural networks to effectively predict DNA accessibility in multiple cell types. These models make it possible to explore the impact of mutations on the predicted accessibility and thus directly link one aspect of the gene regulation puzzle all the way down to the sequence level. We present a model with improved performance on the original dataset of 164 ENCODE and Roadmap Epigenomics Consortium sample types, and then extend the method to provide predictions on any sample with RNA-Seq data without need of DNase-seq for the sample.

We first demonstrate that with several model and algorithmic changes we improve performance across 164 cell types from a mean AUC of 0.895 to a mean AUC of 0.910.

Unfortunately current accessibility models require DNase-seq for each new cell type. Models for detecting transcription factor binding sites, which rely on ChIP-seq for training data, also share this issue. In order to generalize sequence-based predictive models to apply to unseen cell types without requiring re-training we investigate using RNA-Seq as a proxy signature of cell type. The model aims to capture the interdependence of gene expression levels that characterize a cell with the regulatory logic in which sequence-level signatures are combined to determine accessibility without restriction to cell type. We explore the model’s performance when applied to held-out cell types in the ENCODE and Roadmap Epigenomics Consortium data as well as data from the TCGA Pan-Cancer initiative. We look for the impact of non-coding changes in whole-genome sequencing data in TCGA samples, and report on predicted differences in DNA accessibility across cancer subtypes.

Citation Format: Kamil Wnuk, Jeremi Sudol, Shahrooz Rabizadeh, Patrick Soon-Shiong, Christopher Szeto, Charles Vaske. Predicting DNA accessibility in the pan-cancer tumor genome using RNA-Seq, WGS, and deep learning [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 393. doi:10.1158/1538-7445.AM2017-393
Abstract 3934: Bladder cancer tumor heterogeneity: development of a system-level mutation assay
Authors: Patsch Katherin , Matasci Naim , Soundararajan Anjana , Nicoll John , Katz Jonathan , Sanchez Antonio , Feierstein Erika , Loy Christina Van , Xu Zhao , Agus David B. , Gross Mitchell E. , Ruderman Daniel
Published: July 2017

Survival rates for patients with muscle-invasive bladder cancer have not improved in the past 20 years, and new therapies are imperative. Intratumor heterogeneity can complicate molecular profiling attempts to optimize therapy for cancers harboring several actionable tumor subclones. To develop personalized treatment strategies, there is a need for assays to measure intratumor heterogeneity in bladder cancers.

We conducted a pilot study of two muscle invasive high-grade transitional cell carcinoma cases. We used a comprehensive cancer panel (Thermo Fisher) covering >400 cancer genes to analyze distinct tumor loci and matched normal tissues. Based on the identified somatic mutations, we designed a bladder-specific panel to (1) validate our results with increased coverage, and (2) analyze liquid biopsy samples.

Using the comprehensive cancer panel, we sequenced 6 tumor loci to an average sequencing depth of approximately 100x. We detected intratumor heterogeneity in both patients: By applying a combination of frequency-based (minor allele frequency >10%) and probabilistic (probability of difference between observed frequencies due to sampling) filters, we identified 44 credible somatic SNVs, including mutations that were not shared among all three loci. We used these SNVs to design a custom amplicon panel covering 42 SNVs across 38 genes that is suitable for highly fragmented DNA. The custom panel was used to validate the SNVs in the same tumor regions and in liquid biopsy samples from plasma and urine (approximate coverage 6,000x). In both cases, we identified private mutations reported in The Cancer Genome Atlas Urothelial Bladder Carcinoma (TCGA-BLCA) data collection, reflecting tumor evolution. Liquid biopsy samples from urine revealed all trunk mutations but only 1 out of 5 private mutations.

We conclude that tumor evolution can affect distinct loci within bladder tumors, which may not be fully represented in liquid biopsy samples. These results suggest the need for analyzing multiple tumor regions to identify all actionable driver mutations. In the future, we plan to apply our assay to additional foci and patients in order to identify optimal bladder tumor sampling strategies.

Citation Format: Katherin Patsch, Naim Matasci, Anjana Soundararajan, John Nicoll, Jonathan Katz, Antonio Sanchez, Erika Feierstein, Christina Van Loy, Zhao Xu, David B. Agus, Mitchell E. Gross, Daniel Ruderman. Bladder cancer tumor heterogeneity: development of a system-level mutation assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3934. doi:10.1158/1538-7445.AM2017-3934
Abstract 3938: Dynamic interconversion of cancer cells: single-cell analysis of population-based equilibrium
Authors: Chan Yuen San , Yu Edwin-Wai Kin , Wang William Weimao , Fong Chi-Chun , Yip Timothy Tak-Chun , Au Joseph Siu-Kie , Yang Mengsu
Published: July 2017

Background Multiple subpopulations of cells with different phenotypes and genotypes coexist within malignant tumours. Different subpopulations seem to maintain themselves in an equilibrium position via stochastic interconversion. Such plasticity plays an important role in cancer progression, such as transition from epithelial states to mesenchymal state during metastasis, dedifferentiation of non-CSCs to CSCs and rapid transition to MDR cells during drug exposure. We attempt to explore this interconversion plasticity at both population and single cell levels to better understand the dynamics of phenotypic transition.

Methods: We isolated CD90+ and CD90- cells from A549 lung adenocarcinoma cell line by flow cytometry and then characterized their phenotypic behaviors and interconversion dynamics at both population and single cells level. Single cells were cultured in 96 well plate for 13 days. Viable cells were then transferred into 6 well plate for further expansion until enough cells were collected for measurement of CD90 expression in flow cytometry.

Results: A549 maintains a stable expression of CD90 (~50%) which is bimodal distributed. The sorted subpopulations eventually achieved a new equilibrium (CD90+ à 60% , CD90 - à 30%) and did not converge during the bulk cell interconversion. Individual single cell clones showed large variation of CD90 expression. The distribution of CD90 expression was significantly different between CD90+ and CD90- single cell clones, while both groups of clones achieved a similar average equilibrium as compared with the bulk cells. We noted two types of clones defined as transition clones and committed clones with differential interconversion capacities. The later one showed no sign of interconversion and was committed to the original CD90+/- expression level. Further experiments showed that the committed CD90+ cells were more mesenchymal with higher migration ability, while the committed CD90- cells were more epithelial and tumorigenic.

Conclusions: Interconversion plasticity varied from cell to cell and the committed cells prohibited the return of sorted cell subpopulations returned back to the original equilibrium. Phenotypic studies of CD90+ and CD90- revealed that the interconversion between them was EMT related.

Citation Format: Yuen San Chan, Edwin-Wai Kin Yu, William Weimao Wang, Chi-Chun Fong, Timothy Tak-Chun Yip, Joseph Siu-Kie Au, Mengsu Yang. Dynamic interconversion of cancer cells: single-cell analysis of population-based equilibrium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3938. doi:10.1158/1538-7445.AM2017-3938
Abstract 395: Activation mechanisms of cancer associated MEK1 mutants
Authors: Gao Yijun , Chang Matthew T. , McKay Daniel , Yaeger Rona D. , Torres Merna , Muniz Keven , Matthias Drosten , Abdel-Wahab Omar I. , Barbacid Mariano , Caponigro Giordano , Stuart Darrin , Solit David , Taylor Barry S. , Yao Zhan , Rosen Neal
Published: July 2017

Activating BRAF mutants drive human tumors by dysregulating ERK signaling despite ERK-dependent feedback suppression of RAS. These RAF mutants become RAS independent, by either of two mechanisms, and they are thus unaffected by feedback inhibition of upstream signaling. Recently, it has become clear that mutation activation of MEK1 or MEK2 occur at appreciable frequency in human tumors, but the mechanism of activation of these mutants and whether they remain dependent on upstream activation of RAS or RAF signaling remains unknown. Here we characterized the mechanism of activation of 18 recurrent MEK1 mutants identified in human cancer. Based on these data, they fall into three classes. The kinase activity of first class remains dependent on RAF mediated phosphorylation of S218 and S222; the second class of MEK mutants has basal RAF-independent activity, but can be further stimulated by RAF and the third class no longer requires phosphorylation of S218 and S222 and signals in a RAF independent manner. These features determined their sensitivity to ERK-dependent feedback regulation and the ability to drive ERK signaling output in cells. The more RAF-independent activity the mutants acquired, the better they could activate downstream ERK pathway, leading to increased transforming activities in MEF cells in the absence of RAF. This is consistent with their genetic association with RAS, RAF and NF1 mutations in human tumors. The Class 3 RAF-independent MEK1 mutants tend to be mutually exclusive with those mutants, while coexistence with RAS/RAF/NF1 mutations is frequently observed in tumors with Class 1 or 2 MEK1 mutants. Moreover, functional class correlated with sensitivities of ERK signaling driven by these mutants to MEK inhibitors that function by different mechanisms. Unlike RAF dependent or regulated MEK1 mutants, the ERK signaling driven by RAF independent MEK1 mutants is insensitive to an allosteric MEK inhibitor that functions by preventing RAF mediated MEK phosphorylation. However, signaling driven by all classes of MEK1 mutants is sensitive to an ATP competitive MEK1 inhibitor which targets MEK1 kinase activity.

Citation Format: Yijun Gao, Matthew T. Chang, Daniel McKay, Rona D. Yaeger, Merna Torres, Keven Muniz, Drosten Matthias, Omar I. Abdel-Wahab, Mariano Barbacid, Giordano Caponigro, Darrin Stuart, David Solit, Barry S. Taylor, Zhan Yao, Neal Rosen. Activation mechanisms of cancer associated MEK1 mutants [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 395. doi:10.1158/1538-7445.AM2017-395
Abstract 3952: Rapid autopsy programs: A systematic review
Authors: Li Brian , Dhani Neesha , Yu WeiYang , Khan Shawn , Grose Elysia
Published: July 2017

A Rapid Autopsy Program (RAP) is an advanced method of biospecimen procurement whereby autopsies are conducted within 2-6 hours postmortem in order to obtain substantial amounts of high-quality, fresh tissue in support of current and future research. Rapid autopsies were first introduced in the late 1980’s and are still a relatively novel approach, but growing in popularity as a viable alternative to traditional tissue sampling methods for cancer research. In oncology, primary tumour, metastatic, and normal tissue from uninvolved organ sites are sampled and can be subsequently snap frozen in liquid nitrogen or preserved in formalin-fixed paraffin-embedded blocks. Fresh tissue can also be distributed immediately to cancer researchers allowing for analyses incompatible with archival frozen or fixed tissue. Tissue degradation has been found to be minimized in different tissue types by decreasing the postmortem interval – the time between death and tissue preservation. Institutions capable of executing the collection and maintenance of such a repository of biospecimens are a monumental asset to clinicians and researchers who are interested in investigating tumour biology, metastatic disease, and treatment response. In addition to the clear scientific benefits of RAPs, these programs offer a unique opportunity for palliative cancer patients to donate their bodies to aid cancer research. This systematic review was conducted to examine the scope of impact of RAPs thus far on biomedical, translational, and clinical research in the field of oncology, in hopes to inform best practice for collection and use of biospecimen in oncology. Medline, Embase, Cochrane Database of Systematic Reviews, Cochrane CENTRAL, and Ovid MEDLINE will be searched with devised strategies and screened from inception until present. In addition, grey literature will be reviewed. Two independent reviewers will be responsible for study selection and data extraction. The results of the systematic review are pending.

Citation Format: Brian Li, Neesha Dhani, WeiYang Yu, Shawn Khan, Elysia Grose. Rapid autopsy programs: A systematic review [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3952. doi:10.1158/1538-7445.AM2017-3952
Abstract 3956: Intra-individual genomic heteroneity of ovarian cancer and clinical utility of ascitic cancer cells for mutation profiling
Authors: Choi Youn Jin , Kim Min Sung , Lee Sug Hyung
Published: July 2017

Intra-individual tumoural heterogeneity (ITH) is a hallmark of solid tumours and impedes accurate genomic diagnosis and selection of proper therapy. The purpose of this study was to identify ITH of ovarian high-grade serous carcinomas (OSCs) and to determine the utility of ascitic cancer cells as a resource for mutation profiling in spite of ITH. We performed whole-exome sequencing, copy number profiling, and DNA methylation profiling of four OSC genomes using multiregional biopsy from 13 intra-ovarian lesions, 12 extra-ovarian tumour lesions (omentum/peritoneum), and ascitic cells. We observed substantial levels of heterogeneity in mutations and copy number alterations (CNAs) of the OSCs. We categorized the mutations into 'common', 'shared' and 'private' according to the regional distribution. Six common, 8 shared, and 24 private mutations were observed in known cancer-related genes. Common mutations had a higher mutant allele frequency and included TP53 mutations in all four OSCs. Region-specific chromosomal amplifications and deletions involving BRCA1, PIK3CA, and RB1 were also identified. Of note, the mutations detected in ascitic cancer cells represented 92.3-100% of overall somatic mutations in the given case. Phylogenetic analyses of ascitic genomes predicted a poly-seeding origin of somatic mutations in ascitic cells. Our results demonstrate that despite ITH, somatic mutations, CNAs, and DNA methylations in both “common” category and cancer-related genes were highly conserved in ascitic cells of OSCs, highlighting the clinical relevance of genome analysis of ascitic cells. Ascitic tumour cells may serve as a potential resource to discover somatic mutations of primary OSC with diagnostic and therapeutic relevance.

Note: This abstract was not presented at the meeting.

Citation Format: Youn Jin Choi, Min Sung Kim, Sug Hyung Lee. Intra-individual genomic heteroneity of ovarian cancer and clinical utility of ascitic cancer cells for mutation profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3956. doi:10.1158/1538-7445.AM2017-3956
Abstract 3967: High microsomal PGE2synthase-1 levels associate with low CD8 T cells and poorer melanoma patient survival
Authors: Kim Sun-Hee , Roszik Jason , Peng Weiyi , Ekmekcioglu Suhendan , Grimm Elizabeth A.
Published: July 2017

COX-2 and its product PGE2 are reported to enhance carcinogenesis and tumor progression and to support immunosuppression, as reported by others in melanoma and other cancers. As most COX-2 inhibitors result in cardiotoxicity, the downstream microsomal PGE2 synthase-1 (mPGES1) is now also an alternative consideration for therapeutic targeting. Our previously published results (Kim, et al., 2016) showed that mPGES1 protein increased with melanoma patient clinical stage and that its intense expression was associated with reduced patient survival. We also reported that mPGES1 inhibition attenuated cell survival and increased apoptosis in human melanoma cells and significantly suppressed tumor growth in melanoma xenografts. This current study was designed to evaluate whether expression of mPGES1 contributes to immune evasion. Tumors in a Stage III melanoma TMA analysis demonstrated that high mPGES1 expression (intensity 3) was significantly associated with low CD8 levels (lower than median CD8 percent value) (r=-0.2722, p=0.0128). Importantly, stage III melanoma patients with this high mPGES1 and low CD8 signature had significantly increased risk of death as determined by Kaplan-Meier method and Univariate Cox proportional hazards regression models, and compared with patients with low mPGES1/low CD8 and low mPGES1/high CD8. Furthermore, the differential levels of several chemokines including CXCL1 and CXCL2 to regulate T cell migration were correlated the mPGES1 expression in the TCGA melanoma data set. Collectively, our study continues to resolve the potential role of mPGES1 in regulation of immune evasion and is designed as a preclinical approach to develop a rational therapeutic strategy targeting PGE2-driven inflammatory mediators as useful adjuvants for immune-based therapy.

Citation Format: Sun-Hee Kim, Jason Roszik, Weiyi Peng, Suhendan Ekmekcioglu, Elizabeth A. Grimm. High microsomal PGE2synthase-1 levels associate with low CD8 T cells and poorer melanoma patient survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3967. doi:10.1158/1538-7445.AM2017-3967
Abstract 3976: Resolving cellular complexity in renal cell carcinoma with droplet-based single cell transcriptomics
Authors: Vijayakumar Siva , Bolisetty Mohan , Sivajothi Santosh , Robson Paul
Published: July 2017

Purpose: The development, growth, invasion and metastasis of cancer is supported and sustained by complex tissue microenvironments. Interactions between the stromal cells, cancer cells and immune cells through autocrine and paracrine signaling, influences many stages of cancer progression. Furthermore, understanding the identity of the tumor infiltrated immune cell populations is known to predict survival outcomes. Kidney cancer is the 12th most prevalent cancer in the world and is predicted to affect 62,700 new cases in 2016 in the United States alone (American Cancer Society). Existing studies on renal cell carcinoma utilizing bulk profiling methods have been unable to deconvolute the role of the various cell types in promoting the progression this cancer type.

Method: Single cell transcriptomics of primary tumors is a powerful approach to study not only the cancer cells but also the cells of the microenvironment. We profiled the various cell types that comprise primary renal cell carcinoma using the Chromium Single Cell 3’ system (10X Genomics).

Results: With this single cell approach, we have generated thousands of single cell transcriptomes leading to the identification of numerous cell types and cell states in renal cell carcinoma and matched normal kidney. The microenvironment of renal cell carcinoma tumors contains a heterogeneous mix of stromal, immune and kidney cell types. Compared with the matched normal, the tumors are heavily infiltrated with lymphocytes, mast cells and cells of the myeloid lineage. We identified pathways enriched in specific cell types that illuminate potential mechanisms underlying the cell-cell interactions between cancer cells, stromal cells, and the infiltrating immune cells thus highlighting potentially new targeting strategies for the treatment of this disease. In addition, the single cell data described here will be used to deconvolute existing and new bulk analyses of renal cell carcinoma patients and to establish targeted antibody panels for use in flow and imaging mass cytometry. We anticipate both will lead to better patient stratification to inform patient-specific treatment regimes.

Conclusion: Droplet-based single cell transcriptomics can successfully identify the numerous cell types that comprise a tumor, providing a foundation of knowledge to inform biomarker selection and provide insight into novel treatment strategies. This approach reflects a more comprehensive view of renal tumors that is often masked by bulk profiling analyses.

Citation Format: Siva Vijayakumar, Mohan Bolisetty, Santosh Sivajothi, Paul Robson. Resolving cellular complexity in renal cell carcinoma with droplet-based single cell transcriptomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3976. doi:10.1158/1538-7445.AM2017-3976
Abstract 3986: Regulation mechanism and clinical significance of KPNA2 promotes MDSCs infiltrating in epithelial ovarian cancer
Authors: Huang Shuting , Zheng Min , Wu Yuanzhong
Published: July 2017

Myeloid-Derived Suppressor Cells (MDSCs) are a population of immune cells that negatively regulate immune responses, which could promote tumor invasion and metastasis. Recently research has revealed that tumor could actively reshape the immune-microenvironment for its progression. We recently demonstrated that KPNA2 is a candidate oncogene in ovarian cancer that was closely related to MDSCs density, and found there was a positive position feedback between KPNA2 expressed and MDSCs density, which might contribute to the poor prognosis of patients. Based on these results, we combined clinical sample analysis, animal model and experimental studies to do further molecular and genomic research. The conduction of this project will help explain the molecular mechanisms of how KPNA2 regulates ovarian cancer through suppressing the immune system, and provide new strategies by molecular targeted therapy combining biological therapy for ovarian cancer.

Note: This abstract was not presented at the meeting.

Citation Format: Shuting Huang, Min Zheng, Yuanzhong Wu. Regulation mechanism and clinical significance of KPNA2 promotes MDSCs infiltrating in epithelial ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3986. doi:10.1158/1538-7445.AM2017-3986
Abstract 3989: Different expression and prognostic effect of PD-L1/PD-1 in SCC and non-SCC of non-small cell lung cancer
Authors: Liu Yunpeng , Wang Shuo , Qu Xiujuan , Liu Zhi , Che Xiaofang , Ling Xu , Hou Kezuo , Fan Yibo , Song Na , Wen Ti , Li Ce
Published: July 2017

Although checkpoint inhibitors targeting PD-1/PD-L1 pathway have demonstrated similar clinical activity in treatment of NSCLC both for squamous(SCC) and non-squamous carcinoma (non-SCC) patients, the predictive role of PD-L1 expression is distinct between two histologies in clinical trials. In order to investigated the expression and prognostic roles of PD-1/PD-L1, data from TCGA and GEO database were analyzed and further validated with our original data by IHC in 133 resected NSCLC tumor tissue(49 SCC and 84 non-SCC). Through the analysis of online data, we concluded that PD-L1 expression in SCC was significantly higher than that of non-SCC. In addition, in non-SCC, PD-L1 was associated with immune response and indicated worse prognosis while no such results were observed in SCC. Further, our original data demonstrated PD-L1 expression in tumor cells(TC) and immune cells(IC) were significantly more often and more intense in SCC(P<0.001). In survival analysis, PD-L1 expression in TC and IC were associated with shorter overall survival in non-SCC (P=0.019 and 0.003, respectively). However, in SCC, PD-L1 did not demonstrate any significant effect neither in TC or IC (P=0.526 and 0.674, respectively) which consistent with online data. Contrary to PD-L1, no significant differences of PD-1 expression in tumor infiltrating lymphocytes(TILs) were showed between two histologies. PD-1 in TILs of SCC correlated with worse prognosis (P=0.025) while in non-SCC there was no correlation(P=0.986). Taken together, these findings suggest distinct regulation mechanisms and clinical significance of PD-L1/PD-1 pathway in NSCLC subgroups.

Citation Format: Yunpeng Liu, Shuo Wang, Xiujuan Qu, Zhi Liu, Xiaofang Che, Xu Ling, Kezuo Hou, Yibo Fan, Na Song, Ti Wen, Ce Li. Different expression and prognostic effect of PD-L1/PD-1 in SCC and non-SCC of non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3989. doi:10.1158/1538-7445.AM2017-3989