bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2109: The bromodomain-containing protein ATAD2 promotes GR-mediated enzalutamide resistance in castration-resistant prostate cancer
Authors: De La Rosa Richard
Published: July 2021

The purpose of this study is to investigate how the activity of the glucocorticoid receptor (GR) is orchestrated to drive the development of enzalutamide resistance in metastatic, castration resistant prostate cancer (mCRPC). Enzalutamide (Enz) is the second-generation antagonist of the androgen receptor, the fuel for the development and progression of prostate cancer. Even though enzalutamide is an effective and well-tolerated treatment in a broad spectrum of patients with castration-resistant prostate cancer, almost all the patients become resistant to the drug eventually. One of the plausible mechanisms to account for the development of enzalutamide resistance is the activation of GR signaling. Upon Enz treatment, GR is upregulated, binds to the cis-regulatory elements of some AR target genes, activates the transcription of these downstream genes and thus circumvents the inhibitory effect of the AR antagonist on prostate cancer cell growth. Unfortunately, it is still unknown how GR-mediated enzalutamide resistance is controlled. Recently, a group of epigenetic regulators, bromodomain (BRD)-containing proteins, have been indicated to promote prostate cancer progression. They function as AR cofactors and dictate the transcriptional activity of the nuclear receptor in response to different ligands. Our prior work found that one of the BRD family members, called ATAD2, plays an essential role in orchestration of GR signaling in the presence of enzalutamide. ATAD2 is significantly elevated upon GR activation and positively regulates GR expression in return. Coimmunoprecipitation demonstrated that ATAD2 binds to GR in enzalutamide-resistant CRPC cells, and it is required for the GR-dependent transcriptional reprogramming in the presence of Enz. Most importantly, we demonstrated that CRPC cells that are Enz-refractory due to the activation of GR signaling become Enz sensitive when ATAD2 is depleted All these findings suggest that ATAD2 acts as a cofactor for GR to induce a gene expression pattern that results in Enz-refractory phenotype in CRPC cells. Our work will not only provide an in-depth insight into the mechanisms underlying enzalutamide resistance, but also lay the foundation for the development of novel therapeutic strategies targeting glucocorticoid receptor (GR) signaling to treat this lethal disease.

Citation Format: Richard De La Rosa. The bromodomain-containing protein ATAD2 promotes GR-mediated enzalutamide resistance in castration-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2109.
Abstract 2118: Non-coding germline GATA3 variants alter chromatin topology and contribute to pathogenesis of acute lymphoblastic leukemia
Authors: Yang Hongbo , Zhang Hui , Luan Yu , Liu Tingting , Roberts Kathryn , Qian Mao-xiang , Zhang Bo , Yang Wenjian , Perez-Andreu Virginia , Xu Jie , lyyanki Sriranga , Kuang Da , Reshmi Shalini , Gastier-Foster Julie , Smith Colton , Pui Ching-Hon , Evans William , Hunger Stephen , Hunger Stephen , Platanias Leonidas , Relling Mary , Mullighan Charles , Loh Mignon , Yue Feng , Yang Jun
Published: July 2021

Inherited non-coding genetic variants confer significant disease susceptibility in many cancers. However, the molecular processes of by which germline variants contribute to somatic lesions are poorly understood. We performed targeted sequencing in 5,008 patients and identified a key regulatory germline variant in GATA3 strongly associated with Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL). By creating an isogenic cellular model with CRISPR-Cas9 system, we showed that this variant activated a strong enhancer that significantly upregulated GATA3 transcription, which in turn reshaped the global chromatin accessibility and 3D genome organization. Remarkably, this genotype switch induced a chromatin loop between the CRLF2 oncogene and a distal enhancer, similar to the somatically acquired super-enhancer hijacking event in patients. GATA3 genotype-related alterations in transcriptional control and 3D chromatin organization were further validated in Ph-like ALL patients. Finally, we showed that GATA3 directly regulates CRLF2 and potentiates the oncogenic effects of JAK-STAT signaling in leukemogenesis. Altogether, our results provide evidence for a novel mechanism by which a germline non-coding variant contributes to oncogene activation epigenetic regulation and 3D genome reprogramming.

Citation Format: Hongbo Yang, Hui Zhang, Yu Luan, Tingting Liu, Kathryn Roberts, Mao-xiang Qian, Bo Zhang, Wenjian Yang, Virginia Perez-Andreu, Jie Xu, Sriranga lyyanki, Da Kuang, Shalini Reshmi, Julie Gastier-Foster, Colton Smith, Ching-Hon Pui, William Evans, Stephen Hunger, Stephen Hunger, Leonidas Platanias, Mary Relling, Charles Mullighan, Mignon Loh, Feng Yue, Jun Yang. Non-coding germline GATA3 variants alter chromatin topology and contribute to pathogenesis of acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2118.
Abstract 2123: Chromatin profiling of glioblastoma tissues identifies core oncogenic dependency and therapeutic opportunities
Authors: Xu Liang , Chen Ye , Huang Yulun , Sandanaraj Edwin , Yu John S. , Lin Ruby Yu-Tong , Dakle Pushkar , Ke Xin-Yu , Chong Yuk Kien , Koh Lynnette , Mayakonda Anand , Nacro Kassoum , Hill Jeffrey , Huang Mo-Li , Gery Sigal , Lim See Wee , Huang Zhengyun , Xu Ying , Chen Jianxiang , Bai Longchuan , Wang Shaomeng , Wakimoto Hiroaki , Yeo Tseng Tsai , Ang Beng Ti , Müschen Markus , Tang Carol , Tan Tuan Zea , Koeffler Phillip H.
Published: July 2021

Background: Glioblastoma (GBM) is the most aggressive and therapy-refractory brain tumor in adults. Molecular profiling of GBM on the basis of gene expression, DNA methylation and genomic variations has advanced both cancer research and clinical diagnosis. However, the enhancer architectures and regulatory circuitries governing tumor-intrinsic transcriptional diversity and subtype identity are still elusive.

Methods: Chromatin immunoprecipitation followed by sequencing analysis was applied to examine H3K27ac deposition and to map the active regulatory landscapes across 95 GBM biopsies and 12 normal brain tissues. RNA-sequencing analysis was performed to measure transcriptome and to classify transcriptional subtypes. Super-enhancer associated genes and master transcriptional factors were identified. Function of novel cancer associated genes was explored using both patient-derived GBM propagating cells and orthotopic xenograft models.

Results: Analyses of differentially regulated enhancers and super-enhancers uncovered previously unrecognized layers of inter-tumor heterogeneity. Integrative analysis of variant enhancer loci and transcriptome identified topographies of transcriptional enhancers and core regulatory circuitries in four molecular subtypes of primary tumors: AC1-mesenchymal, AC1-classical, AC2-proneural and AC3-proneural. Subtype-specific enhancer domains contributed to transcriptional diversity and shaped subtype identity. Moreover, this study reveals novel oncogenic dependency of GBM on various super-enhancer-driven transcriptional factors and druggable targets (e.g., BRD4).

Conclusion: Through profiling of transcriptional enhancers, we provide clinically relevant insights into the molecular classification, pathogenesis and therapeutic intervention of GBM.

Citation Format: Liang Xu, Ye Chen, Yulun Huang, Edwin Sandanaraj, John S. Yu, Ruby Yu-Tong Lin, Pushkar Dakle, Xin-Yu Ke, Yuk Kien Chong, Lynnette Koh, Anand Mayakonda, Kassoum Nacro, Jeffrey Hill, Mo-Li Huang, Sigal Gery, See Wee Lim, Zhengyun Huang, Ying Xu, Jianxiang Chen, Longchuan Bai, Shaomeng Wang, Hiroaki Wakimoto, Tseng Tsai Yeo, Beng Ti Ang, Markus Müschen, Carol Tang, Tuan Zea Tan, Phillip H. Koeffler. Chromatin profiling of glioblastoma tissues identifies core oncogenic dependency and therapeutic opportunities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2123.
Abstract 215: Developing an integrated prognostic score using germline risk variants and RNA expression from primary breast cancers for prognostication of survival across subtypes
Authors: Rajagopal Padma Sheila , Liang Yanyu , Barbeira Alvaro , Melia Owen , Zheng Jiamao , Zheng Yonglan , Yoshimatsu Toshio , Huo Dezheng , Gao Guimin , Olopade Olufunmilayo F. , Im Hae K.
Published: July 2021

Introduction: RNA expression offers prognostication in early-stage, hormone-receptor positive breast cancer. We hypothesized that germline risk variants may influence patients after breast cancer onset, and that integration of common germline risk variants with RNA expression may broaden prognostication across subtypes.

Methods: We designed two prognostic scores for women of European ancestry using genome-wide association study (GWAS) summary statistics on cancer risk from the Breast Cancer Association Consortium (133,384 cases and 113,789 controls) and individual-level genotype/clinical information from 694 women in The Cancer Genome Atlas (TCGA). PrediXcan, a transcriptome-wide association tool family, was used to generate gene-level weights of association to breast cancer risk that were then multiplied by gene expression levels. A predicted transcriptomic score (PTRS) used predicted expression levels based on germline genotypes, and an observed transcriptomic score (OTRS) used normalized RNA expression. Cox proportional hazards tested score performance on overall survival (OS) and progression-free survival (PFS).

Results: Estimation performance is shown in Table 1. A 1388-gene PTRS with range from -0.5 to 4 offered initial prognostic ability in OS across subtypes (HR: 0.69 per 1-point increase, 95% CI: 0.48-1.00, p-value 0.05). This trend persisted with clinical covariates in a multivariate regression (HR: 0.61, 95% CI: 0.36-1.00, p=0.06). No associations were seen between PTRS/PFS or OTRS/OS. OTRS/PFS trended toward statistical significance with clinical factors included (age, AJCC staging). PTRS for OS did not show subtype-specific performance.

Conclusion: Patients with breast cancer do not have broad access to prognostic data across subtypes. Integration of germline and somatic data types offers potential to improve current limitations in prognostic testing for patients.

Table 1.Performance of predicted transcriptomic score (PTRS) and observed transcriptomic score (OTRS)Cox regression hazard ratio95% CIp-valuePolygenic score - OS0.960.64-1.400.84PTRS-OS0.690.48-1.000.05OTRS-OS0.990.97-1.000.66Polygenic score - PFS1.100.77-1.600.55PTRS - PFS1.100.80-1.600.49OTRS - PFS0.980.96-1.000.10

Citation Format: Padma Sheila Rajagopal, Yanyu Liang, Alvaro Barbeira, Owen Melia, Jiamao Zheng, Yonglan Zheng, Toshio Yoshimatsu, Dezheng Huo, Guimin Gao, Olufunmilayo F. Olopade, Hae K. Im. Developing an integrated prognostic score using germline risk variants and RNA expression from primary breast cancers for prognostication of survival across subtypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 215.
Abstract 2154: Expression of androgen receptor splice variant, AR-V7, in high grade serous ovarian cancer
Authors: Wilson Benjamin , Xiu Joanne , Ulm Michael , Redfern Tiffany , Narayanan Ramesh , ElNaggar Adam
Published: July 2021

Objective: Androgen receptors (AR) are expressed in up to 55% of ERα-negative breast cancers overall and up to 35% of those classified as TNBC. Similarly, AR is expressed in approximately 40% of high grade serous ovarian cancer (HGSOC). The development of AR splice variants (AR-SV), particularly AR-V7, has been shown to be a poor prognostic feature, and mechanism of androgen resistance in breast and prostate cancers. We sought to determine if AR-V7 was present in HGSOC and if there was a correlation with platinum sensitivity and/or treatment outcomes.

Methods: Pair-matched chemo-naïve and platinum-treated HGSOC specimens were identified from our tissue repository. Formalin-fixed paraffin-embedded tumor (FFPE) blocks were collected and tested for expression of AR-V7. Gene fusion detection and variant detection were performed on mRNA isolated from a formalin-fixed paraffin-embedded tumor sample using the Illumina NovaSeq platform (Illumina, Inc., San Diego, CA) and Agilent SureSelect Human All Exon V7 bait panel (Agilent Technologies, Santa Clara, CA). FFPE specimens underwent pathology review to diagnose percent tumor content and tumor size; a minimum of 10% of tumor content in the area for microdissection was required.

Results: Twelve chemo-naïve specimens collected at time of diagnosis by initial biopsy (n=8) or primary debulking surgery (n=4) were pair-matched with the patients' subsequent platinum-treated specimens from either interval debulking surgery (n=6) or biopsy of recurrence (n=6). Following penultimate platinum, 50% were platinum resistant and 50% were platinum sensitive at time of second pathology. Only 19 of the 24 specimens submitted had sufficient RNA for testing, 8 chemo-naïve and 11 platinum-treated. AR-V7 variant transcript was not detected in any of the specimens submitted.

Conclusions: AR-V7 was not expressed in any specimen before or after platinum based chemotherapy suggesting it may not be a useful prognostic marker in the management of HGSOC. Further investigation into gynecologic malignancies with high AR expression and treated with androgen deprivation therapy may elucidate different results.

Citation Format: Benjamin Wilson, Joanne Xiu, Michael Ulm, Tiffany Redfern, Ramesh Narayanan, Adam ElNaggar. Expression of androgen receptor splice variant, AR-V7, in high grade serous ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2154.
Abstract 2162: Spatially resolved image-based transcriptomics using high-throughput single-molecule fluorescence in situ hybridization (HITSFISH)
Authors: Coleski Andrej , Pitchiaya Sethu , D'Silva Jeremy , Cieslik Marcin , Chinnaiyan Arul
Published: July 2021

Single-cell transcriptomics enables the study of heterogeneity by uncovering the transcriptome of individual cells, which is averaged out in bulk analyses. This facilitates the grouping of cells based on similar transcriptomic profiles, potentially identifying cell subpopulations, such as rare tumor cells that mediate drug resistance or undergo metastasis. Image-based transcriptomic techniques can go a step further in uniquely profiling cells, by providing information about the spatial co-ordinates of transcripts and cells, whilst shedding light on cell morphology for phenotypic characterization. Single-molecule Fluorescence in situ hybridization (smFISH) is one image-based method in which fluorescently labeled tiling-oligonucleotide (TO) probes are bound along the length of complementary RNA strands, allowing for the detection of transcripts at nanometer-scale resolution. However, smFISH data analysis has been difficult to implement at scale due to the tradeoff between transcript resolution and imaging area, as well as the lack of automated feature detection tools. To address this issue, we have developed a pipeline called HIgh-Throughput Single-molecule FISH (HITSFISH), which uniquely combines smFISH with structured-illumination microscopy allowing for large-area high-resolution imagery with automated feature detection and analysis. We have validated our platform by showing high transcriptomic concordance between HITSFISH and other standard methods such as bulk RNA-seq and qPCR, demonstrating its ability to accurately measure RNA levels. With our pipeline we look to answer questions pertaining to the spatiotemporal landscape of gene expression within and across normal and cancerous cells. In addition, by using immunofluorescence in tandem with our platform, we can study transcript localization at various subcellular landmarks, such as membraneless organelles, to gain insight on subcellular mechanisms of gene regulation.

Citation Format: Andrej Coleski, Sethu Pitchiaya, Jeremy D'Silva, Marcin Cieslik, Arul Chinnaiyan. Spatially resolved image-based transcriptomics using high-throughput single-molecule fluorescence in situ hybridization (HITSFISH) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2162.
Abstract 2197: Dysregulated alternative splicing landscape identifies intron retention as a hallmark and spliceosome as a therapeutic vulnerability in aggressive prostate cancer
Authors: Zhang Dingxiao , Hu qiang , Liu Xiaozhuo , Ji Yibing , Chao Hsueh-Ping , Tracz Amanda , Kirk Jason , Buonamici Silvia , Zhu Ping , Wang Jianmin , liu song , Tang Dean
Published: July 2021

Dysregulation of mRNA alternative splicing (AS) has been implicated in development and progression of hematological malignancies. How the global AS dysregulation contributes to the development and progression of solid tumors is under-studied and remains generally unclear. Here we describe the first comprehensive AS landscape in the spectrum of human prostate cancer (PCa) development, progression and therapy resistance. We find that the severity of splicing dysregulation correlates with disease progression and establish intron retention (IR) as a hallmark of PCa stemness and aggressiveness. Systematic interrogation of 274 splicing-regulatory genes (SRGs) uncovers prevalent SRG mutations associated with, mainly, copy number variations leading to mis-expression of ~68% of SRGs during PCa evolution. Consequently, we identify many SRGs as prognostic markers associated with splicing disruption and patient outcome. Interestingly, androgen receptor (AR) controls a splicing program distinct from its transcriptional regulation. The spliceosome modulator, E7107, reverses cancer aggressiveness and inhibits the growth of castration-resistant PCa (CRPC) xenograft models and of the autochthonous Hi-Myc tumors. Altogether, our studies establish aberrant AS landscape caused by dysregulated SRGs as a hallmark of PCa aggressiveness and a novel therapeutic vulnerability for CRPC.

Citation Format: Dingxiao Zhang, qiang Hu, Xiaozhuo Liu, Yibing Ji, Hsueh-Ping Chao, Amanda Tracz, Jason Kirk, Silvia Buonamici, Ping Zhu, Jianmin Wang, song liu, Dean Tang. Dysregulated alternative splicing landscape identifies intron retention as a hallmark and spliceosome as a therapeutic vulnerability in aggressive prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2197.
Abstract 2460: Oncogenic regulation of lipogenesis in docetaxel resistant prostate cancer cells
Authors: Martinez Shannalee R. , Aybar Adrián Pérez , Osterman Carlos J. Díaz
Published: July 2021

Prostate cancer is the second leading cause of cancer related deaths in men in the United States. Docetaxel is the standard chemotherapeutic agent available for patients with advanced prostate cancer, in both hormone sensitive and castration resistant settings. However, the development of resistance to docetaxel curtails the efficacy of the drug in prostate cancer patients. Aberrant lipid metabolism has been linked to poor outcomes and therapy resistance in prostate cancer, particularly through overexpression and activation of lipogenic enzymes including ATP citrate lyase, acetyl coA carboxylase, fatty acid synthase, and stearoyl coA desaturase. Together, these enzymes drive fatty acid biogenesis, which has been linked to docetaxel resistance. The purpose of this study is to delineate the molecular mechanisms underlying the lipogenic reprogramming of docetaxel resistant prostate cancer cells, as these represent targets for reversal of this drug resistance. Docetaxel resistant sublines of LNCaP, 22rv1, PC3 and DU145 cell lines were established through chronic continuous exposure to incrementally increasing concentrations of docetaxel, followed by authentication by single tandem repeat (STR) analysis and validation of resistance using MTT viability assays and immunoblot detection of multidrug resistance protein 1 (MDR1/p glycoprotein/ABCB1). Enhanced intracellular accumulation of lipids using BODIPY 493/593 staining and flow cytometric detection was used to confirm the lipogenic phenotype of docetaxel-resistant cells. Oncogenic cMYC expression was also detected by immunoblotting, and its role in regulating lipogenic enzyme expression was evaluated by siRNA interference in docetaxel resistant prostate cancer sublines. cMYC knockdown suppressed the expression of lipogenic enzymes including acetyl coA carboxylase, fatty acid synthase, and stearoyl CoA desaturase in docetaxel-resistant prostate cancer cells. Furthermore, pharmacological inhibition of cMYC production (JQ1) or activity (10058-F4) effectively resensitized resistant cells to docetaxel, as did targeting of key lipogenic enzymes. These data indicate that a cMYC associated lipogenic program is associated with docetaxel resistance in prostate cancer and may represent targets for interventions to enhance docetaxel response in patients with advanced prostate cancer.

Citation Format: Shannalee R. Martinez, Adrián Pérez Aybar, Carlos J. Díaz Osterman. Oncogenic regulation of lipogenesis in docetaxel resistant prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2460.
Abstract 2484: Detection of loss of heterozygosity in GBM revealed the role of a tumor suppressive gene in regulating glioma progression and tumor recurrence
Authors: Kiang Mei-Yee , Sun Stella , Leung Gilberto Ka-Kit
Published: July 2021

Adducin 3 (ADD3) is a major cytoskeleton protein in glioma cells. Our previous studies demonstrated a significant downregulation of ADD3 in GBM when compared with lower-grade gliomas, and that ADD3 knockdown promoted malignant phenotypes. GBMs are usually characterized by high levels of genomic instability, including frequent loss of heterozygosity (LOH) of chromosome 10q, whilst ADD3 is also located in chromosome 10q25.1-25.2 where it is known to be a tumor-suppressor region. However, it is not known whether ADD3 downregulation in malignant gliomas is associated with allelic loss in 10q. To determine 10q LOH on different polymorphic DNA loci (covering ADD3 locus) on glioma tissues, specimens of Normal, Grade II, Grade III, Grade IV GBM and recurrent GBM tissue were included. DNA fragments were evaluated by using a PCR-based microsatellite capillary electrophoresis and analyzed by a fragment analyzer. LOH status were correlated with patient survival for potential prognostic and diagnostic implications. LOH on ADD3 locus was detected in high grade and recurrent GBMs. It is revealed that downregulation of ADD3 level in GBM is associated with genetic instability of LOH in 10q and associated with recurrence disease. This study would provide significant implications, particularly for understanding pathogenesis of GBM and add valuable credits on the diagnostic as well as prognostic implications.

Citation Format: Mei-Yee Kiang, Stella Sun, Gilberto Ka-Kit Leung. Detection of loss of heterozygosity in GBM revealed the role of a tumor suppressive gene in regulating glioma progression and tumor recurrence [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2484.
Abstract 2500: The Forkhead-like protein Dach1 governs sensitivity to WEE1 kinase and PARP inhibitors
Authors: Li Zhiping , Jiao Xuanmao , Robertson A. Gordon , Ashton Anthony W. , Kossenkov Andrew , Chen Hao , Phoon Lai Yee , Lan Li , Pestell Richard G.
Published: July 2021

Molecular analysis of all cancer types in the cancer genome atlas (TCGA) demonstrated that multiple pathways may be disrupted within a given tumor including PCa. Prostate cancer (PCa), the second leading cause of death in American men, includes distinct genetic subtypes with distinct therapeutic vulnerabilities. Substratification of PCa into genetic subtypes, forms the basis of rational therapy for PCa. Known genetic drivers to PCa include deletions of target genes (PTEN and NKX3.1 deletions), rearrangements/fusions of multiple genes (including TMPRSS2 and the oncogenic ETS transcription factor, ERG) and genetic predisposing factors (including germline DNA-repair gene mutations). An increasingly common finding in PCa are defects in DNA repair. The Drosophila Dac gene was initially cloned as dominant inhibitor of Elipse. The human homologue, DACH1, encodes a winged helix/Forkhead DNA-binding protein that binds in chromatin to FOXM1 sites. Herein, reduced expression of DACH1 is shown in PCa, correlating with DACH1 promoter methylation. Homozygous deletion of DACH1 occurred in up to 18% of human PCa, associated with increased AR activity, increased somatic copy-number alteration and poor prognosis. DACH1 increased DNA repair, as shown in comet assays, enhancing nonhomologous end joining (NHEJ) while reducing homology directed repair (HDR) in DNA repair reporter assays. Laser wounding showed DACH1 was recruited to sites of DNA damage, augmenting the recruitment of Ku70/Ku80. In Dach1 deletion cells reduced expression of Dach1 was associated with resistance to inhibitors of Poly-(ADP)-ribose polymerase (PARP) and enhanced sensitivity to inhibitors of WEE1 kinase and DNA-PK. The abundance of DACH1 in prostate cancer may define a subclass of prostate cancer warranting selective therapies.

Citation Format: Zhiping Li, Xuanmao Jiao, A. Gordon Robertson, Anthony W. Ashton, Andrew Kossenkov, Hao Chen, Lai Yee Phoon, Li Lan, Richard G. Pestell. The Forkhead-like protein Dach1 governs sensitivity to WEE1 kinase and PARP inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2500.
Abstract 2565: Detection of ETV1 expression in human prostate tissue specimens using a novel and highly specific rabbit monoclonal antibody
Authors: Tan Shyh-Han , Young Denise , Elsamanoudi Sally , Kagan Jacob , Srivastava Sudhir , Dobi Albert , Petrovics Gyorgy , Sesterhenn Isabell A. , Chesnut Gregory T.
Published: July 2021

Introduction: ETV1 is frequently involved in genomic fusions and translocation events that lead to its overexpression in multiple cancers. These events occur in approximately 5% of prostate cancers, which are mutually exclusive from tumors harboring TMPRSS2-ERG fusion or PTEN deletion. Studies suggest a correlation between strong ETV1 protein expression and poor outcome in prostate cancer. ETV1 has been reported to synergistically cooperate with KIT as a lineage survival factor in gastrointestinal stromal tumors. The expression of ETV1 in a subset of sarcomas that harbor CICrearrangements or CIC-DUX4 gene fusions presents a reliable molecular signature for the diagnosis of this cancer. Our understanding of the role that ETV1 plays in the activation of prostate cancer has been limited by the lack of ETV1 specific antibodies.

Methods: A novel ETV1 monoclonal antibody (MAb) was raised by immunization of ETV1 peptides in rabbit followed by screening of hybridomas by ELISA and immunoblot assays. Further screening using exogenously expressed ETV1, ETV4, ETV5, ERG, SPDEF, and FLI1 proteins identified the clone with the most reactive MAb. Purified MAb was used to evaluate ETV1 expression on a tissue micro-array (TMA) constructed from radical prostatectomies of 50 African American (AA) and 50 Caucasian American (CA) patients by immunohistochemistry (IHC). Key residues required for Mab binding were mapped by ELISA using overlapping peptides and alanine scanning.

Results: IHC evaluation using the ETV1 specific rabbit Mab on a prostate cancer TMA derived from 100 patients identified five ETV1 positive cases, of whom four were CA. The index tumors for these five ETV1 cases were ERG negative. One patient harbored both ERG positive and ERG negative tumor foci, and as expected, the ETV1 positive tumor focus was ERG negative, and vice versa.

Conclusions: We developed a novel rabbit monoclonal ETV1 antibody that is suitable for IHC assay in human prostate tissue. An evaluation of prostate cancer specimens confirmed the reported frequency of ETV1 alteration. Further evaluation using tissue specimens from larger cohorts to establish the sensitivity and specificity of this antibody and validate the utility of ETV1 detection in improving the diagnosis and stratification of prostate and other cancers are in progress.

Citation Format: Shyh-Han Tan, Denise Young, Sally Elsamanoudi, Jacob Kagan, Sudhir Srivastava, Albert Dobi, Gyorgy Petrovics, Isabell A. Sesterhenn, Gregory T. Chesnut. Detection of ETV1 expression in human prostate tissue specimens using a novel and highly specific rabbit monoclonal antibody [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2565.
Abstract 2614: A deep learning model-based lung cancer risk assessment for incidental pulmonary nodules
Authors: Yuan Pengyu , He Tiancheng , Nguyen Hien , Wong Stephen T.
Published: July 2021

Lung cancer is the leading cause of cancer death among both men and women. The solitary pulmonary nodule (SPN) is frequently seen on radiographs and computed tomography (CT) and often provokes additional clinical and imaging activities as an SPN alerts possible early stage lung cancer. However, reliable diagnosis of malignant lung nodules in the workup of SPN is challenging, making it difficult for early stage cancer management to avoid morbidity due to malignant cancer, patient distress, and increased costs caused by more invasive and unwarranted procedures for benign cases. We developed a deep learning model based on convolutional neural networks (CNNs) to predict lung cancer risk for incidental SPN. The model can help better management of incidental lung cancer by providing a reliable characterization of SPNs. Deep learning models currently are applied to nodule detection in low-dose CT scans of lung cancer screening for at-risk population. Our work aims to investigate whether deep learning can differentiate benign from malignant incidental SPN during normal dose CT exams, where the number of patients with SPN is an order more than that of lung screening population. We collected an incidental SPN dataset containing 139 CT scans to generate the nodule malignancy prediction model using 3D CNN. This dataset was collected from either contrast or non-contrast CT scans with regular doses. All 139 patients went through surgeries to remove the tumors. CT images were collected within one year before the surgery. 90 cases are malignant and the rest are benign. Ground truth labels were obtained from biopsies. Data were preprocessed by resampling, normalization, and cropping to unique volume around the SPN. The radiologists annotated the SPN location. Since the two nodule classes are imbalanced, we did online data augmentation to increase the variety of lung nodule images and balance the two classes. We used 80% of the data for training and 20% for testing. The deep learning structure adapted is a 3D Resnet with 68 layers and 33M parameters. The area under the receiving operating characteristics curve (AUC) of the prediction result was 0.81 while the accuracy was 79.6%. Applying GradCam to visualize important regions in CT scans contributing to the final predictions, we found that our trained ResNet model focuses on meaningful areas in lung nodules and detects malignant SPNs in most cases. Our preliminary results shows the feasibility of using deep learning to predict incidental lung nodules and improve diagnostic speed and accuracy.

Citation Format: Pengyu Yuan, Tiancheng He, Hien Nguyen, Stephen T. Wong. A deep learning model-based lung cancer risk assessment for incidental pulmonary nodules [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2614.
Abstract 2632: Fostering engagement in biobanking and research through the NCI Cancer Moonshot Biobank patient and provider engagement website
Authors: Casas-Silva Esmeralda , Agrawal Lokesh , Ellis Helena J. , Gopalakrishnan Veena , Guan Ping , Jensen Mark , Madero Natalie , McDermott Sean , McLean Jeffrey , Rao Abhi , Suh James , Wanyiri Jane , Weil Carol J. , Williams Mickey , Moore Helen M.
Published: July 2021

The National Cancer Institute's (NCI) Cancer Moonshot Biobank (the Biobank) aims to accelerate cancer research on treatment resistance and sensitivity through collection of longitudinal biospecimens and health data donated by participants diagnosed with late-stage cancer and receiving standard of care therapy at participating NCI Community Oncology Research Program (NCORP) sites.

Participant and provider engagement are a central tenet of the Biobank. We therefore created an innovative website to interface directly with participants and providers and to meet our engagement goals of returning value to stakeholders, operating with transparency, maintaining bi-directional communication, and enrolling a diverse population that represents the U.S. including racial/ethnic minorities, rural populations and others that are historically underrepresented in clinical research. The Biobank engagement website has several features designed to provide return of value to stakeholders. This includes clear information about the program, transparency about how biospecimens and data will be used in research, and the ability to access and download documents such as clinical biomarker tests and signed consent forms. The website also includes project updates, participant and provider resources, and educational material.

The Cancer Moonshot Biobank Participant and Provider Engagement website serves as an important interface between the Biobank and its stakeholders. Its ultimate aim is to facilitate project engagement and return value to participants and providers.

Citation Format: Esmeralda Casas-Silva, Lokesh Agrawal, Helena J. Ellis, Veena Gopalakrishnan, Ping Guan, Mark Jensen, Natalie Madero, Sean McDermott, Jeffrey McLean, Abhi Rao, James Suh, Jane Wanyiri, Carol J. Weil, Mickey Williams, Helen M. Moore. Fostering engagement in biobanking and research through the NCI Cancer Moonshot Biobank patient and provider engagement website [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2632.
Abstract 2671: SMAD4 as a potential gatekeeper for genomic instability and mTOR-mediated tumorigenesis in esophageal adenocarcinoma
Authors: Milne Julia V. , Gotovac Jovana R. , Fujihara Kenji M. , Duong Cuong P. , Phillips Wayne A. , Clemons Nicholas J.
Published: July 2021

Esophageal cancer is the 8th most common cancer worldwide and has the 6th highest mortality rate of all cancers. The 5-year survival rate following esophageal adenocarcinoma (EAC) diagnosis is dismal at less than 15 percent, indicating a dire need for improved therapeutic strategies and early detection. EAC develops stepwise following exposure to chronic gastric reflux: From pre-malignant Barrett's metaplasia, through stages of low- and high-grade dysplasia until developing into invasive cancer. Mutation or loss of common tumor suppressor genes TP53 and SMAD4 act as markers for cancer progression, occurring in high-grade dysplastic tissue and invasive EAC, respectively. Our novel in vivo tumorigenesis model demonstrates progression of Barrett's metaplasia to EAC, in which SMAD4-deficient Barrett's metaplasia cells form tumors in immunodeficient mice after a period of latency and in a dose-dependent manner. This delayed tumor growth onset suggests further drivers are required for oncogenesis, and these SMAD4-deficient cells and tumors display a greater degree of genomic instability than wildtype-SMAD4 controls. A genome-wide CRISPR-Cas9 knockout screen unveiled a synthetic lethal relationship between SMAD4-deficiency and cell cycle checkpoint inhibition, suggesting a role for SMAD4 in maintaining genomic stability and a potential novel therapeutic avenue for SMAD4-deficient EAC. Additionally, a concurrent in vivo CRISPR-Cas9 tumorigenesis screen produced tumors 4-fold faster than the previous model and identified regulators of mTOR signaling as co-operative drivers of tumorigenesis in EAC. Wildtype-SMAD4 cells failed to generate tumors despite undergoing the same genetic perturbations, indicating a potential gatekeeping effect of SMAD4 in mTOR-mediated EAC tumorigenesis. In sum, loss of SMAD4 acts as a double-edged sword, increasing genomic instability and thereby rendering EAC cells sensitive to cell cycle checkpoint inhibition, whilst simultaneously co-operating with modulated mTOR signaling to promote tumorigenesis in EAC xenograft models.

Citation Format: Julia V. Milne, Jovana R. Gotovac, Kenji M. Fujihara, Cuong P. Duong, Wayne A. Phillips, Nicholas J. Clemons. SMAD4 as a potential gatekeeper for genomic instability and mTOR-mediated tumorigenesis in esophageal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2671.
Abstract 268: DNA-launched HPV E7 nanoparticle vaccine induces potent anti-tumor cytolytic T-cell responses
Authors: Liaw Kevin , Xu Ziyang , Tursi Nicholas , Chokkalingam Neethu , Kulp Daniel , Weiner David
Published: July 2021

Nanotechnology-based approaches have provided promising tools for development of novel cancer immunotherapeutic immunogens through the decoration of multiple antigens on nanoparticle scaffolds. However, challenges with inducing robust cytolytic T cell responses critical to anti-tumor immunity, as well as purification and scalability, have hampered their clinical translation. To address these challenges, here we present the design and characterization of a DNA-launched nanoparticle vaccine displaying the HPV E7 epitope for inducing anti-tumor immune responses. We designed a nanovaccine construct by scaffolding the HPV E7 epitope with bacterial enzyme lumazine synthase. We demonstrate that through electroporation-facilitated DNA vaccination encoding the antigen-scaffold construct, this nanovaccine self-assembles in vivo into nanoparticles displaying 60 copies of the antigen on its surface. Compared to vaccination with CpG-adjuvanted HPV E7 peptide, the DNA-launched nanovaccine induced significantly greater antigen-specific CD8+ T cell responses upon stimulation with the HPV E7 peptide as well as with the epitope natively expressed on the target tumor cells. In a flank model of TC-1 lung cancer, the DNA-launched nanovaccine achieved complete tumor protection in 70% of mice when administered prophylactically. With therapeutic treatment, the nanovaccine achieved complete tumor regression in 60% of mice. In addition, anti-tumor immunity was retained upon rechallenge over 100 days after initial tumor challenge, supporting the induction of immune memory. These promising results, along with its facile characterization and scalability, support that DNA-launched nanovaccines that self-assemble in vivo can induce significant anti-tumor cellular immunity and are ideally positioned for study in clinical translation.

Citation Format: Kevin Liaw, Ziyang Xu, Nicholas Tursi, Neethu Chokkalingam, Daniel Kulp, David Weiner. DNA-launched HPV E7 nanoparticle vaccine induces potent anti-tumor cytolytic T-cell responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 268.
Abstract 2713: Preclinical models for translational immuno-oncology research: patient-derived xenografts on humanized mice
Authors: Stecklum Maria , Wulf-Goldenberg Annika , Brzezicha Bernadette , Klinghammer Konrad , Jöhrens Korinna , Walther Wolfgang , Hoffmann Jens
Published: July 2021

Background: The preclinical evaluation of novel immune checkpoint modulators is dependent on models with functional human immune cells. In previous experiments, we have demonstrated, that we can use either peripheral blood mononuclear cells (PBMC) or hematopoietic stem cells (HSC) to establish a humanized immune system on highly immunodeficient mice with functional T, B or NK cells. Furthermore, we determined PD-L1 expression as a predictive marker and target for immunotherapy on different patient-derived xenografts (PDX). By co-transplantation of PDX, we successfully generated a fully human tumor-immune-cell model in mice. Finally, we evaluated the functionality of the model by the treatment with checkpoint inhibitors and for combination therapies i.e. with chemotherapy or radiation.

Methods: HSC-humanized mice (HIS) were generated by i.v. transplantation of HSC. Engraftment of immune cells was monitored by FACS analysis. For PBMC-humanized mice, immune cells were implanted i.v. PD-L1 expression on PDX was determined by FACS and immunohistochemistry (IHC). PDX from 13 different entities (i.e. melanoma, lymphoma) were transplanted on HIS mice and treated with Ipilimumab (Ipi) and Nivolumab (Nivo) alone or in combination with radiation.

Results: 14 weeks after HSC transplantation up to 20% of the human immune cells in the blood were T-cells. We have transplanted more than 40 different PDX from 13 different tumor entities on HIS mice. Most of the investigated PDX (>70%) successfully engrafted and showed no significant difference in tumor growth compared non-humanized mice. However, for some PDX we observed a delayed tumor growth or a complete rejection. Engraftment delay seems to correlate with the PD-L1 expression of PDX (the higher PD-L1, then the higher growth delay). Treatment with Ipi or Nivo alone or in combination led to a minor tumor growth delay and an increased percentage of T-cells in the blood and the tumor. Response showed a correlation to innate immune response and PD-L1 expression of PDX and could further be increased by combination with radiotherapy.

Conclusions: Our humanized immune-PDX models enable appropriate preclinical translational research on tumor immune biology and the evaluation of new therapies and combinations, as well as the identification and validation of biomarkers for immune therapy.

Citation Format: Maria Stecklum, Annika Wulf-Goldenberg, Bernadette Brzezicha, Konrad Klinghammer, Korinna Jöhrens, Wolfgang Walther, Jens Hoffmann. Preclinical models for translational immuno-oncology research: patient-derived xenografts on humanized mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2713.
Abstract 2728: Modulation of NFkappaB signaling influences tumor associated macrophage phenotypes in murine models of breast cancer
Authors: Bullock Kennady K. , Parker Dominique , Glass Evan B. , Hoover Alyssa , Merkel Alyssa , Maynard Richard , Wilson Andrew , Giorgio Todd , Rhoades Julie A. , Yull Fiona E.
Published: July 2021

Tumor associated macrophages (TAMs) often exist in high densities in breast tumors and are associated with adverse clinical outcomes. TAM repolarization as a strategy to shift the breast cancer microenvironment from immunosuppressive to immunostimulatory is an active area of research. The canonical and non-canonical NfkappaB (NFkB) signaling pathways represent potential points of intervention to influence TAM phenotypes. We have found that upregulation of the canonical NFkB signaling pathway specifically in macrophages can be used to repolarize TAMs toward an M1, anti-tumor phenotype. Using our doxycycline inducible mouse model termed IKFM, we are exploring the effects of macrophage specific upregulation of NFkB signaling on different stages of tumor growth and metastasis. In studies using the IKFM mouse model, we found that doxycycline-induced NFkB activation in macrophages in established orthotopic mammary tumors has therapeutic effects on tumor outcomes. Compared to control mice, we found that IKFM mice have reduced tumor burden in the lung, indicating prevention of metastasis. Ongoing studies are focused on understanding how macrophage specific activation of NFkB influences the metastatic niches of lung and bone; two of the most common sites of breast cancer metastasis. To move our findings towards clinical translation, we have developed an M2-macrophage targeted nanoparticle that activates NFkB signaling by delivering an siRNA against IkappaBalpha, an inhibitor of the pathway. The M2 targeted nanoparticle is currently being tested in murine models of breast cancer metastasis to both lung and bone to translate our findings from the IKFM mouse model to a pharmacological manipulation.

Citation Format: Kennady K. Bullock, Dominique Parker, Evan B. Glass, Alyssa Hoover, Alyssa Merkel, Richard Maynard, Andrew Wilson, Todd Giorgio, Julie A. Rhoades, Fiona E. Yull. Modulation of NFkappaB signaling influences tumor associated macrophage phenotypes in murine models of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2728.