bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 5371: Multimodal CRISPR activation screens reveal new mechanisms to sensitize cancer cells to targeted T cell mediated elimination
Authors: Akana Reece V. , Sun Chang , Kim YoungMin , Cai Yuxin , Kim Subin , Yoe Jeehyun , Laveroni Olivia , Jerby-Arnon Livnat 
Published: March 2024

Abstract
Cell autonomous resistance mechanisms can protect cancer cells from targeted T cell elimination, resulting in resistance to a broad range of immunotherapies. High-throughput gene knockout and gene inhibition screens have been instrumental in identifying key drivers of such mechanisms, yet often uncovering gene activity that is essential but not sufficient for response. Here we set out to leverage gene activation as an orthogonal approach to identify novel regulators of the cancer-immune interface. First, we performed CRISPR activation screens in melanoma cells co-cultured with antigen-specific primary CD8+ T cells. Our findings revealed a collection of novel genes that substantially sensitize melanoma cells to targeted T cell-driven elimination when activated, including DNA damage and unfolded protein response genes (e.g., ETS1, TSPYL2), Wnt ligands (e.g., WNT3A, WNT1), and others that are deleterious only in combination with targeting T cells. Among the resistance genes we found regulators of insulin signaling (e.g., HGF, TCF7L2) and numerous glycoproteins (e.g., PDPN, CD44). Second, we validated top novel hits in 2D and in 3D spheroid models, using optical readouts to investigate their impact on cancer cell elimination and cancer-T-cell spatial dynamics. Third, we performed a Perturb-seq screen to stratify dozens of hits into functional groups based on the transcriptional readouts in the cancer cells. Lastly, we developed an in situ Perturb-Seq method, where we combine high-plex spatial transcriptomic readouts at subcellular resolution with in situ detection of genetically modified cells, and applied it to track the impact of the (de)sensitizing perturbations not only on the cancer cells themselves, but also on T cell recruitment and suppression. Taken together, our study provides a catalog of immune evasion mechanisms and an array of novel targets, demonstrating that synthetic gene activation can diminish immune evasion and open novel avenues for development of agonist-based and RNA-based interventions.
Citation Format: Reece V. Akana, Chang Sun, YoungMin Kim, Yuxin Cai, Subin Kim, Jeehyun Yoe, Olivia Laveroni, Livnat Jerby-Arnon. Multimodal CRISPR activation screens reveal new mechanisms to sensitize cancer cells to targeted T cell mediated elimination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5371.
Abstract 5388: FOSL1 promotes glioma tumorigenesis and stemness through NF-κB and STAT3 pathways
Authors: Liu Mingli , Ramar Vanajothi , Khedri Azam , Hudson BreAnna , Guo Shanchun 
Published: March 2024

Abstract
Background: Glioblastoma multiforme (GBM) is the most common and malignant primary brain tumor, GBM’s inevitable recurrence suggests that glioblastoma stem cells (GSC) render these tumors persistent. Our previous work showed that FOSL1, which is transactivated by the STAT3 gene, functions as a tumorigenic gene in glioma pathogenesis and is a diagnostic marker and potential drug target for glioma patients. Accumulating evidence shows that STAT3 and NF-κB cooperate to promote the development and progression of various cancers. The link between STAT3 and NF-κB suggests that NF-κB can also transcriptionally regulate FOSL1 and contribute to gliomagenesis.
Methods: 1). To investigate downstream molecules of FOSL1, we analyzed the transcriptome after overexpression of FOSL1 in a PDX-L14 line, characterized by very low FOSL1 expression 2). We conducted immunohistochemical staining for FOSL1 and p65 using rabbit polyclonal anti-FOSL1 and p65 in glioma tissue microarrays (TMA) derived from 76 glioma patients and 10 normal individuals. 3). Mutants of the human FOSL1 promoter, featuring mutations in essential binding sites for NF-κB and STAT3, were generated using a Q5 site-directed mutagenesis kit. Subsequently, we examined luciferase activity in glioma cells and compared it to the wild-type FOSL1 promoter. 4). We explore the mutual regulation between NF-κB signaling and FOSL1 by modulating the expression of NF-κB or FOSL1. Subsequently, we assessed the activity of FOSL1 and NF-κB. 5) To understand the role of FOSL1 in cell growth and stemness, we conducted a CCK-8 assay, cell cycle analysis, assessing apoptosis and GSC markers, ALDH1 and CD133 under varying FOSL1 expression conditions.
Results: 1) Transcriptome analyses of downstream molecules of FOSL1 show that NF-κB signaling pathways are regulated by FOSL1. 2) p65 protein expression correlates to the expression of FOSL1 in glioma patients. 3) NF-κB is a crucial transcription factor activating the FOSL1 promoter in glioma cells. 4) Mutual regulation between NF-κB and FOSL1 contributes to glioma tumorigenesis and stemness through promoting G1/S transition and inhibition of apoptosis.
Conclusion: The FOSL1 molecular pathway is functionally connected to NF-κB and STAT3 activation, enhances stemness, and FOSL1 is a novel GBM drug target.
Citation Format: Mingli Liu, Vanajothi Ramar, Azam Khedri, BreAnna Hudson, Shanchun Guo. FOSL1 promotes glioma tumorigenesis and stemness through NF-κB and STAT3 pathways [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5388.
Abstract 5407: Temozolomide promotes matrix metalloproteinase 9 expression through p38 MAPK and JNK pathways in glioblastoma cells
Authors: Jung Chaeyong , Thanh Hien Duong , Lee Sueun , Moon Kyung-Sup 
Published: March 2024

Abstract
Glioblastoma multiforme (GBM) is a highly aggressive and deadly brain cancer. Temozolomide (TMZ) is the standard chemotherapeutic agent for GBM, but many patients experience recurrence and invasion of tumor cells. We investigated whether TMZ treatment of GBM cells regulates matrix metalloproteinases (MMPs), which have main function to promote tumor cell invasion. TMZ effectively killed GL261, U343, and U87MG cells at a concentration of 500 μM, and surviving cells upregulated MMP9 expression and its activity but not those of MMP2. TMZ also elevated levels of MMP9 mRNA and MMP9 promoter activity. Subcutaneous graft tumors survived from TMZ treatment also exhibited increased expression of MMP9 and enhanced gelatinolytic activity. TMZ- mediated-MMP9 upregulation was specifically mediated through the phosphorylation of p38 and JNK. This then stimulates AP-1 activity through the upregulation of c-Fos and c-Jun. Inhibition of the p38, JNK, or both pathways counteracted the TMZ-induced upregulation of MMP9 and AP-1. This study proposes a potential adverse effect of TMZ treatment for GBM: upregulation of MMP9 expression associated with increased invasion and poor prognosis. This study also provides valuable insights into the molecular mechanisms by which TMZ treatment leads to increased MMP9 expression in GBM cells.
Citation Format: Chaeyong Jung, Hien Duong Thanh, Sueun Lee, Kyung-Sup Moon. Temozolomide promotes matrix metalloproteinase 9 expression through p38 MAPK and JNK pathways in glioblastoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5407.
Abstract 5482: Spatial and multi-omic interrogation of the primary and recurrent glioblastoma microenvironment
Authors: Clemenceau Jean R. , Suarez-Meade Paola , Quiñones-Hinojosa Alfredo , Hwang Tae Hyun 
Published: March 2024

Abstract
Glioblastoma remains as one of the deadliest malignancies with most cases likely to recur. Recent work has focused on characterizing not only glioma cells, but the tumor microenvironment via single-cell and spatial transcriptomic technologies. This has improved our understanding of the high heterogeneity of this disease and described potential tumor domains. However, there is still a need to further understand the changes that occur between the primary and the recurrent lesion. We propose that a spatial, multi-omic characterization of matched primary and recurrent glioblastoma can yield insights into recurrence mechanisms occurring throughout the tumor microenvironment. We collected 22 IDH-WT glioblastoma samples from 12 patients (11 primary, 11 recurrent, 10 matched pairs) from the Mayo Clinic Neuroscience Biobank. We performed single-nucleus RNA sequencing (15 samples, 10 patients, 5 matched pairs) and proteomics (16 samples, 6 matched pairs) from fresh-frozen tissue, as well as spatial transcriptomics with 10x Visium (14 samples, 8 patients, 6 matched pairs) and 10x Xenium in-situ molecular imaging (6 samples, 2 matched pairs). Using spatial transcriptomics data, we identified regional subsets present in different proportions and distributions between primary and recurrent glioblastoma. These clusters were characterized with gene set and gene ontology analysis, identifying regions enriched with known glioma subtypes and cellular activity. Spatial-based cluster genes were consistent with significantly differentially expressed proteins. We used single-nucleus RNAseq and in-situ molecular imaging data from matched tissue samples to further interrogate relevant cell populations and location in the tumor micro-environment. We identified regions with enriched extra cellular matrix interaction and remodeling signatures to be highly expressed in recurrent glioblastoma. Primary glioblastoma was enriched with oligodendrocyte-like regions with high cell communication and extracellular vesicle signatures. This study will further our understanding of the changes in the mechanisms at play in the complex ecosystem of glioblastoma as it recurs.
Citation Format: Jean R. Clemenceau, Paola Suarez-Meade, Alfredo Quiñones-Hinojosa, Tae Hyun Hwang. Spatial and multi-omic interrogation of the primary and recurrent glioblastoma microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5482.
Abstract 549: PDZK1 sensitizes TNBC cells to erlotinib via promoting c-Cbl-mediated EGFR degradation and inhibiting EGFR phosphorylation
Authors: Zheng Junfang , Ma Yuanzhen , Fang Zhiyu , Qi Yijun 
Published: March 2024

Abstract
Background: Triple-negative breast cancer (TNBC) is lack of effective targeted therapies. Epidermal growth factor receptor (EGFR) is a potential therapeutic target for TNBC. However, TNBC is resistant to EGFR-targeted drugs in clinical trials, and the tolerance mechanism remains unclear. This study aimed to elucidate the possible resistance mechanism for TNBC to EGFR-targeted therapy and how sensitize TNBC cells to erlotinib.
Methods: PDZ domain containing 1 (PDZK1) correlated with TNBC development was screened out by bioinformatics analysis and its correlation with resistance to erlotinib was identified by western blotting (WB). Its interaction with EGFR and structural basis were determined by GST-Pull down and coimmunoprecipitation. Its regulation on EGFR signaling activation and expression level were detected by WB in PDZK1 overexpression and knockdown TNBC cells, xenograft tissues and correlation analyses. Correlation was performed by gene set enrichment analysis (GSEA) and immunohistochemistry of tissue microarray followed by Pearson analysis. TNBC-suppressing effects of PDZK1 and its sensitizing effects on erlotinib were investigated using cell viability assay, colony-forming assay, wound healing assay, Matrigel invasion assay and xenograft model in combination with PDZK1 wild type/mutant transfection and rescue experiment.
Results: Here we found that PDZK1 was correlated with TNBC development and its level was downregulated in erlotinib-resistant TNBC cells, suggesting that PDZK1 downregulation was related to erlotinib resistance in TNBC. PDZK1 bound with EGFR. Through the interaction, PDZK1 promoted EGFR degradation by enhancing the binding of EGFR with c-Cbl and inhibited EGFR phosphorylation by hindering EGFR dimerization. PDZK1 was specifically downregulated in TNBC tissues and was correlated with poor prognosis of TNBC. Functional assays in vitro and in vivo showed that PDZK1 suppressed TNBC development. Restoring EGFR expression and kinase inhibitor treatment reversed the malignancy caused by PDZK1 overexpression and knockdown, respectively. PDZK1 wild type, but not mutant overexpression enhanced the inhibitory effect of erlotinib on EGFR signaling and TNBC malignancy in vitro and in vivo.
Conclusion: PDZK1 is a significant prognostic factor for TNBC and a potential molecular therapeutic target for TNBC to reverse the tolerance of TNBC cells to erlotinib.
Citation Format: Junfang Zheng, Yuanzhen Ma, Zhiyu Fang, Yijun Qi. PDZK1 sensitizes TNBC cells to erlotinib via promoting c-Cbl-mediated EGFR degradation and inhibiting EGFR phosphorylation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 549.
Abstract 5514: Crosstalk between prostatic epithelial cells and the tumor microenvironment drives cancer progression and metastasis
Authors: Yanushko Darya , Falcon Beatriz German , Keime Céline , Ye Tao , Thibault-Carpentier Christelle , Metzger Daniel , Laverny Gilles 
Published: March 2024

Abstract
Prostate cancer is a heterogeneous disease with a slow progression and a highly variable clinical outcome. Recent advances in the clinical handling of localized prostate cancers resulted in an almost 100% 10-year survival, but no curative treatment is currently available for metastatic disease. Thus, understanding the molecular mechanisms of progression of primary tumors to metastasis is crucial to develop new therapeutic strategies. While tumorigenesis initiates in prostatic epithelial cells, various cell types, including cancer associated fibroblasts (CAFs), immune and endothelial cells produce extra-cellular matrix and secrete signaling molecules that might contribute to tumor progression. However, the complex communication network driving tumor progression is poorly characterized. As the tumor suppressors PTEN and TP53 are among the most frequently altered genes in prostate cancer, we generated genetically engineered mice with Pten and Trp53 selective inactivation in prostatic epithelial cells at adulthood. Their analysis revealed that they develop precancerous lesions that progress to invasive and metastatic tumors. Spatial transcriptomic and single-cell analyses of Pten- and Trp53-deficient prostatic tumors identified the presence of a cancer cell population exhibiting cell plasticity driven by Jak/Stat3 signaling and induced by CAF-produced IL6. In addition, these tumors exhibit a hypoxic core and an immune infiltrate dominated by neutrophils. Inhibition of the hypoxic signaling reduced immune infiltration in primary tumors and their metastatic potential, but did not induce apoptosis of Pten- and Trp53- deficient epithelial cells, in contrast to those of Pten-deficient tumors. These results underline the importance of the tumor genomic background for therapeutic intervention. Taken together, our work highlights the key role of the tumor microenvironment in driving prostate cancer progression and uncovers new vulnerabilities of this disease.
Citation Format: Darya Yanushko, Beatriz German Falcon, Céline Keime, Tao Ye, Christelle Thibault-Carpentier, Daniel Metzger, Gilles Laverny. Crosstalk between prostatic epithelial cells and the tumor microenvironment drives cancer progression and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5514.
Abstract 5520: β2 adrenergic receptor signaling promotes colorectal cancer liver metastasis
Authors: Zhang Yixuan , Chen Jintian , Chan Thomas Ting Hei , Wang Huanyu , Lu Jiahuan , Li Jingqing , Zhou Jingying 
Published: March 2024

Abstract
Liver is the most common site for cancer metastases, which may be attributed to the hepatic immunosuppressive microenvironment. Recent studies have highlighted the immunomodulatory and tumor-supportive roles of neurotransmitter-receptor signals in various cancers. Given that various neurotransmitter receptors are expressed in the liver, we aimed to investigate their possible involvement in the metastatic-prone liver microenvironment. Single-cell RNA sequencing analysis identified ADRB2, which encodes β2-adrenergic receptor (β2-AR), as the top-upregulated neurotransmitter receptor gene in colorectal cancer (CRC) and liver metastasis in patients with CRC liver metastasis (CRLM). Co-immunofluorescence (co-IF) staining confirmed the colocalization of β2-AR with myeloid cell marker CD11b instead of T cell marker CD3 and NK cell marker CD56 in patients with CRLM. Consistently, we observed the upregulation of CD11b/β2-AR double-positive cells in the liver tissues from mice bearing CRC or CRLM compared to naïve mice through co-IF staining. High-dimensional flow cytometry analysis further revealed that β2-AR was mainly expressed in macrophages and myeloid-derived suppressor cells (MDSCs). Treatment of CRC-tumor bearing mice with pan β-AR inhibitor propranolol and β2-AR specific inhibitor showed a potential to reduce liver metastatic incidence without suppressing primary CRC. Mechanistically, β2-AR activation may upregulate the expression of S100a8/9 to enhance the immunosuppressive activity of macrophages and MDSCs, thereby promoting CRC liver metastasis. Taken together, our findings reveal the potential of targeting β2-AR signaling for the prevention and treatment of liver metastasis.
This project is supported by the Hong Kong University Grants Committee through the General Research Fund (14104820, 14107622, 24110323) and the Strategic Seed Funding for Collaborative Research Scheme (SSFCRS) from the Chinese University of Hong Kong.
Conflict of interest: The authors declare no conflict of interest.
Citation Format: Yixuan Zhang, Jintian Chen, Thomas Ting Hei Chan, Huanyu Wang, Jiahuan Lu, Jingqing Li, Jingying Zhou. β2 adrenergic receptor signaling promotes colorectal cancer liver metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5520.
Abstract 5607: A highly efficient donor system for CRISPR knock-in editing
Authors: Wu Lipeng , Yin Dezhong , Su Hua , Fellows Justin , Fu Mao , Heller Julian , Park Brian , Juminaga Alex 
Published: March 2024

Abstract
CRISPR is a potent tool widely used for genome editing. While CRISPR knock-out, achieved through in-del mutations via cellular repair mechanisms, has proven remarkably effective, the knock-in system for exogenous fragment insertion encounters challenges due to limited specificity and efficiency. Two primary methods for exogenous fragment insertion, namely NHEJ (Non-Homologous End Joining) and HR (Homologous Recombination), exist. HR allows for the construction of DNA insertions with precise junctions but is comparatively less efficient than NHEJ. To address these limitations, our system focuses on enhancing the efficiency of HR-based genome editing. Primarily, we have optimized the Lenti All-in-one CRISPR system to augment the delivery and targeting efficiency of the Cas9-gRNA complex. The system has been successfully employed to knock out Glutamine Synthetase (GLUL) in HEK293T cells, exhibiting superior efficiency compared to commercially available tools. Secondly, we have implemented a pseudo-lenti system to amplify the entry capacity of donor DNA with homology arms into target cells. This technique has shown an improved HR effect in the experiment to tag a GFP at the C terminals of CDC25A. Lastly, we leverage proximity effects by fusing Cas9 with a capture protein that can grasp a bait sequence attached to the donor DNA, thereby facilitating its transfer to the target sites. Our system has demonstrated exceptional efficacy as a CRISPR knock-out platform and a powerful tool for DNA tagging functionality on target genes.
Citation Format: Lipeng Wu, Dezhong Yin, Hua Su, Justin Fellows, Mao Fu, Julian Heller, Brian Park, Alex Juminaga. A highly efficient donor system for CRISPR knock-in editing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5607.
Abstract 5632: Advancing transcriptome insights during RNA-Seq library construction of FFPE samples in a novel workflow
Authors: Jiang Cissy , Dreskin Elizabeth Jordan , Olcott Angelica , Lingelbach Linda , Perike Srikanth 
Published: March 2024

Abstract
Formalin-fixed, paraffin-embedded (FFPE) tumor tissue has significant clinical benefits in oncology research and is considered vital for studying tumor gene expression through RNA sequencing (RNA-Seq). However, RNA extracted from FFPE samples tends to be severely degraded, making it suboptimal for constructing high-quality RNA-Seq libraries. The RNA-Seq workflow can further impair data quality, as traditional library preparation and pre-library ribodepletion workflows may result in the loss of valuable information due to rare transcripts being not captured or washed away. Moreover, small, degraded fragments are a challenging template for ligases, and the resulting adapter addition may be inadequate. Here we assessed a novel workflow employing a proprietary enzyme to overcome the limitations and biases of cDNA synthesis using reverse transcriptase (RT) and ligases. This innovative RNA-Seq library preparation approach, coupled with a post-library ribodepletion strategy, aims to capture a more complete transcriptome profile from RNA extracted from FFPE breast cancer samples, and can also be used for evaluation of gene regulation through intronic and intergenic bases. The study also compared two commercially available kits, RecoverAll and PureLink, for FFPE RNA extraction, evaluating quality metrics such as RNA integrity number (RIN), DV200, recovery of small RNAs during NGS library construction, and transcriptome coverage. A parallel fresh frozen RNA sample, extracted using a phenol-based reagent, served as a benchmark. Our findings demonstrate that the novel workflow yields a richer dataset, encompassing a wider range of transcriptomic elements, including both small and long RNAs from FFPE samples. Performing ribodepletion after the library preparation procedure led to increased detection of small RNAs compared to pre-library ribodepletion. The robust analytical performance of this streamlined library preparation protocol enables its suitability for RNA-Seq analysis of archived FFPE specimens, offering substantial utility for oncology research.
Citation Format: Cissy Jiang, Elizabeth Jordan Dreskin, Angelica Olcott, Linda Lingelbach, Srikanth Perike. Advancing transcriptome insights during RNA-Seq library construction of FFPE samples in a novel workflow [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5632.