bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A068: Lfng-expressing centroacinar cell is a prime cell-of-origin for p53-deficient pancreatic cancer
Authors: Xu Keli , Chung Wen-Cheng , Zhang Shubing , Atfi Azeddine
Published: September 2024

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with poor understanding of etiology. Studies using mouse models showed that both acinar and ductal cells of the pancreas can be transformed by combination of oncogenic Kras and p53 mutations to form PDAC. How the transforming capacities of pancreatic cells are constrained, or whether a unique subset of cells serve as a prime target for oncogenic transformation, remains obscure. We found that expression of a Notch modulator, Lunatic Fringe (Lfng), is confined to a limited number of cells with centroacinar location and morphology in the adult mouse pancreas. These cells are selectively targeted by KrasG12D coupled with p53 loss-of-function (but not gain-of-function) mutations, leading to rapid pancreatic tumor formation, whereas the elimination of Lfng completely blocks the initiation of these tumors. We showed that p53 constrains the progenitor-like property of Lfng-expressing cells and that Lfng is required for hyperproliferation of these cells when p53 is inactivated. We identified Notch3 as a functional Notch receptor for PDAC initiation and progression in this context. Lfng expression is undetectable in normal acinar and ductal cells but upregulated in acinar- as well as ductal-derived precursor lesions, and its deletion hinders both the initiation and progression of these lesions. Collectively, Lfng marks a centroacinar subpopulation that is restrained by p53 and is uniquely susceptible to oncogenic transformation with p53 loss. Furthermore, Lfng functions as an oncogene in all three exocrine lineages in the adult pancreas. Genetic alterations of LFNG, predominantly in the forms of mRNA upregulation and gene amplification, were detected in 9% of PDAC patients and were associated with shortened overall survival. Consistent with its expression in mouse pancreas, LFNG immunostaining was noted in a subset of centroacinar cells in normal adjacent pancreatic tissues from human patients, and its expression in the adenocarcinoma specimens was positively correlated to tumor grade. Thus, LFNG may play an important role in human PDAC, in parallel with the one shown in the murine models. This study offers new insights into the pancreatic cancer cell-of-origin, initiation, and progression, which may lead to the identification of a therapeutic target for the disease.

Citation Format: Keli Xu, Wen-Cheng Chung, Shubing Zhang, Azeddine Atfi. Lfng-expressing centroacinar cell is a prime cell-of-origin for p53-deficient pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A068.
Abstract A070: Time-Dependent Oxidation and Aggregation of Vimentin upon Binding by β-lap Induced Immunogenic Cell Death
Authors: Qian Feng , Dou Lei , Wang Kaixin , Xia Yi , Chen Zhidong , Zu Yumeng , Xi Shuang
Published: September 2024

β-lapachone (β-lap) has been demonstrated as an NAD(P)H:quinone oxidoreductase1 (NQO1) bioactivatable compound. It initiated a futile redox cycle catalyzed by NQO1, resulting in reactive oxygen species (ROS) generation, with consumption of the reductive NAD(P)H. After a 4-h pulse of sufficiently lethal dose of β-lap, high levels of ROS diffused into nuclei to induce tumor-selective DNA damage and programmed necrosis in cancers overexpressing NQO1. Recently, our experiments showed that prolonged low dose β-lap administration exerted significant and NQO1 independent antitumor efficacy in mouse models with no overt signs of toxicity. Thus, we synthesized a photoactive β-lap probe to identify the molecular target of β-lap. Vimentin, a major constituent of the intermediate filament family of proteins, was identified as the direct binding target of β-lap. Vimentin was highly upregulated in multiple cancers including PDAC, and was a binding partner of β-lap with a dissociation constant at micromolar levels. Ligand binding assays in Panc1 cells employing β-lap as a specific competitor showed that vimentin was bound by β-lap-probe in a β-lap-competitive manner, and further confirmed that the in vivobinding target of β-lap was vimentin. Moreover, it was manifested that endogenous vimentin mRNA levels directly correlated with sensitivity to prolonged exposure of β-lap in a panel of pancreatic cancer cell lines, and knockout of vimentin in Panc1 cells markedly decreased lethality after prolonged exposure of β-lap. Mechanistically, we revealed that β-lap bound to vimentin and induced disulfide bonds that participated in disulfide cross-linking between a pair of vimentin dimers in a concentration and time dependent manner, forming vimentin oligomers of approximately 150 kDa. Prolonged exposure of β-lap promoted the disruption of filament architecture and vimentin degradation, ultimately resulting in cell death. In addition, the cell death by targeting vimentin exhibited a more inflammatory condition and enhanced DC activation. Together, our findings unveiled an unknown molecular target for β-lap and provided valuable insights into its mechanism of action. Targeting vimentin also presented a new strategy that inhibited the tumor growth and synergized with immunotherapy to disrupt immune evasion and reduce immune suppression against pancreatic cancers.

Citation Format: Feng Qian, Lei Dou, Kaixin Wang, Yi Xia, Zhidong Chen, Yumeng Zu, Shuang Xi. Time-Dependent Oxidation and Aggregation of Vimentin upon Binding by β-lap Induced Immunogenic Cell Death [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A070.
Abstract A072: Decoding human kinome specificities through a computational data-driven approach and unveiling kinase activity in pancreatic ductal adenocarcinoma
Authors: Cai Li , Johnson Jared L , Peng Chen , Yaron Tomer M , Cantley Lewis C , DePinho Ronald A
Published: September 2024

Phosphorylation is a crucial post-translational modification involved in virtually all biological processes; its dysregulation is a critical factor in the development and progression of pancreatic ductal adenocarcinoma (PDAC). A large family of kinases drives the phosphorylation of specific substrates, and understanding kinase substrate specificities is central to deciphering kinase activities in PDAC. Recent experimental approaches using positional scanning peptide arrays (PSPA) have determined position-specific scoring matrices (PSSMs) for many human kinases. Inherent limitations and the resource-intensive nature of this empirical method prompted development of a bottom-up computational data-driven method (CDDM) to generate PSSMs of 289 human kinases from large kinase-substrate datasets. Our results align with previous experimental findings and proved effective in identifying kinase-substrate interactions and predicting kinase functions. Moreover, we applied the data-driven PSSMs in analyzing the phosphoproteomics data of PDAC from Clinical Proteomic Tumor Analysis Consortium (CPTAC). We generated kinase activity profiles for each patient and identified subtypes of PDAC with different kinase activities. These clusters were further correlated with key genomic alterations and hallmark cancer processes. Together, we anticipate that such comprehensive knowledge of the kinome specificities will enhance our interpretation of phosphoproteomics data from clinical samples, elucidate signaling pathways under various conditions, and identify key kinases implicated in PDAC.

Citation Format: Li Cai, Jared L Johnson, Chen Peng, Tomer M Yaron, Lewis C Cantley, Ronald A DePinho. Decoding human kinome specificities through a computational data-driven approach and unveiling kinase activity in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A072.
Abstract A074: Integrated clinical and single-cell profiling analysis reveals cellular states associated with metastatic organotropism and survival in patients with pancreatic ductal adenocarcinoma
Authors: Hajkarim Morteza Chalabi , May Michael , Amin Amit Dipak , Jamison Jacob , Tagore Somnath , Caprio Lindsay , Walsh Zachary , Shah Parin , Abuzaid Sinan , Michel Alissa , Dajiang Sun , Wong Winston , Shaikh Neha , Ramaradj Priyanka , Bakir Basil , Kluger Michael D , Chabot John , Hibshoosh Hanina , Rustgi Anil K , Raufi Alexander , Manji Gulam A , Izar Benjamin
Published: September 2024

Some patients with localized pancreatic ductal adenocarcinoma (PDAC) undergo resection with curative intent, but most experience disease recurrence and succumb to their disease. Through examination of 743 PDAC patients in a single center we found that the site of first recurrence is a major predictor for survival. Patients with liver-metastatic recurrence (LIV-MR) had significantly shorter disease free and overall survival compared to those with recurrence to the lung (LUN-MR). We performed single-nucleus RNA-seq (snRNA-seq) and whole-genome sequencing (WGS) of primary PDAC with either LIV-MR or LUN-MR and found that malignant cells adopt transcriptional programs that mimicked that of their ultimate metastatic target organ. We validated this observation in independent data sets of human adult and fetal liver and lung tissues, and two additional patient cohorts who underwent molecular profiling of their actual PDAC liver and lung metastatic lesions. We reconstructed two different trajectories of malignant transformation in PDAC and found that the LIV-MR and LUN-MR are associated with these, suggesting that subsequent metastatic organotropism may be imprinted early in tumorigenesis. Furthermore, primary PDAC tumors recurring with LIV-MR compared to LUN-MR were depleted of infiltrating T cells and enriched with M2-like macrophages, highlighting potential contributions to subsequent outcomes. Taken together, this work provides evidence for pre-existing cellular adaptations that may determine metastatic organotropism in the context of cancer recurrence. These insights may be useful for prognostication and development of therapeutic strategies at the time of diagnosis of resectable PDAC.

Citation Format: Morteza Chalabi Hajkarim, Michael May, Amit Dipak Amin, Jacob Jamison, Somnath Tagore, Lindsay Caprio, Zachary Walsh, Parin Shah, Sinan Abuzaid, Alissa Michel, Sun Dajiang, Winston Wong, Neha Shaikh, Priyanka Ramaradj, Basil Bakir, Michael D Kluger, John Chabot, Hanina Hibshoosh, Anil K Rustgi, Alexander Raufi, Gulam A Manji, Benjamin Izar. Integrated clinical and single-cell profiling analysis reveals cellular states associated with metastatic organotropism and survival in patients with pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A074.
Abstract A075: Establishing the interaction mechanism of oncoprotein, Mucin1, and extracellular vesicle marker protein, Alix, in pancreatic cancer
Authors: Hoagstrom Kristine V , Huang Ying , Borgstahl Gloria E , Hollingsworth Michael A
Published: September 2024

Pancreatic ductal adenocarcinoma (PDAC) remains an aggressive disease with devastating mortality rates largely due to a poor understanding of its progression mechanism. Small extracellular vesicles (sEVs) are used by cancer cells to aid in their progression, and tumor-derived EVs (TEVs) show a distinct protein cargo composition potentially contributing to this rapid progression and metastasis of PDAC. To investigate this phenomenon, we isolated sEVs from conditioned media of pancreatic cancer cell lines using ultracentrifugation and characterized them by transmission electron microscopy (TEM), nanoparticle tracking analysis, and immunoblotting. These sEVs were then used in several studies to investigate critical TEV cargo sorting machinery. For instance, PDAC-derived sEVs have selective enrichment of the oncogenic transmembrane protein Mucin1 (MUC1). Preliminary MUC1KO studies from our lab revealed that MUC1 potentially regulates sEV formation, where we saw a clear impact on sEV cargo in the absence of MUC1. Additional studies found that the cytoplasmic adaptor protein ALG-2-Interacting Protein X (Alix), a protein known to play a critical role during TEV formation, regulates MUC1’s presence in sEVs. From this discovery, we then identified a novel MUC1-Alix interaction, which was confirmed using several techniques, including co-immunoprecipitation (CoIP). To define the Alix-interacting domain of MUC1, we utilized in silico modeling and localized surface plasmon resonance (LSPR) of MUC1.CT and Alix to quantify the interaction. This revealed the cytoplasmic tail of MUC1 (MUC1.CT) as the Alix-interacting domain. Moving forward, our central hypothesis for this study is that this MUC1.CT-Alix interaction utilizes a conserved binding motif that targets Alix to the endosomal membrane and contributes to the formation of MUC1+ sEVs. While a distinct sEV biogenesis pathway employed by cancer cells has not yet been defined, our studies will provide insight into the potential sEV sorting mechanism of MUC1 and other cancer biomarkers and advance the understanding of PDAC progression.

Citation Format: Kristine V Hoagstrom, Ying Huang, Gloria E Borgstahl, Michael A Hollingsworth. Establishing the interaction mechanism of oncoprotein, Mucin1, and extracellular vesicle marker protein, Alix, in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A075.
Abstract A077: The role of RhoV in basal-like pancreatic cancer
Authors: Wu Shang , Feng Jia , Gao Selina , Wang Ting , Li Zebang , Liu Yifei , Huang Chiung-Kuei , Lu Shaolei
Published: September 2024

Background Pancreatic ductal adenocarcinoma is a devastating disease lacking targeted therapy. Basal-like pancreatic cancer is a tumor subtype with a worse clinical outcome than other subtypes. These cancers are characterized by the expression of basal keratins and mesenchymal markers. However, the genes that drive the basal-like phenotype are poorly understood. Ras homolog family member V (RhoV) is one of the Rho GTPases, and it has been demonstrated to drive the progression of lung and breast cancers. The current study aims to reveal the role of RhoV in basal-like pancreatic cancer. Design RhoV, p63, and CK5/6 were stained in sequential sections by immunohistochemistry. Expression of RhoV in pancreatic cancer and cell lines was reanalyzed based on the data from ProteinAtlas and Broad Insitute DepMap Portal. Basal cell line, BxPC3, and non-basal cell line, Panc1 were selected for functional evaluation based on their differentiated RhoV expression. RhoV gene was overexpressed and knocked down using the CRISPR/gRNA-based Lentiviral system, followed by sphere formation, colony formation, migration, and invasion assays. Results RhoV, p63, and CK5/6 were expressed in the same histologic location of basal pancreatic cancer. The gene expression level of RhoV was co-expressed with p63 (a classic basal marker) in four pancreatic cancer cell lines (broadinstitute.org). Knockout of RhoV by three runs of gRNAs in BxPC3 cells all significantly attenuated tumor sphere formation with less colony formation as compared to mock knockout. Overexpression of RhoV in non-basal-like cells, Panc 1, significantly increased the size of sphere formation, numbers of colony formation, and p38 activation compared to EV control. Cell migration and invasion assays showed the same RhoV-dependent migration and invasion in corresponding cells. Panc1 cells with RhoV knockout significantly generated smaller tumor xenografts in immunodeficient mice and RhoV overexpression led to larger tumor sizes. Conclusion Finding the driver of the basal-like phenotype helps to reveal the pathophysiology of the basal-like subtype of pancreatic cancer and facilitates the identification of new targets for therapeutic purposes. Our preliminary results indicated that RhoV, a small G protein from the Rho family of GTPases, played essential roles in tumor proliferation and progression, by activating the p38 signaling pathway. Further study will be focused on revealing Rhov's downstream pathways and exploring the role of RhoV in apoptosis/cell cycle, cell metabolism, autophagy, etc. in basel-like pancreatic cancer.

Citation Format: Shang Wu, Jia Feng, Selina Gao, Ting Wang, Zebang Li, Yifei Liu, Chiung-Kuei Huang, Shaolei Lu. The role of RhoV in basal-like pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A077.
Abstract A078: Functional screen for mediators of onco-mRNA translation specificity in pancreatic cancer
Authors: Kovalski Joanna R , Sarioglu Goksu , Subramanyam Vishvak , Hernandez Grace , Rademaker Gilles , Oses-Prieto Juan A , Slota Macey , Wen Kwun Wah , Kim Grace E , Burlingame Alma L , Goodarzi Hani , Perera Rushika M , Ruggero Davide
Published: September 2024

Oncogenic protein dosage is tightly regulated to enable cancer cells to adapt and survive. In healthy cells, translation initiation of onco-mRNA transcripts encoding oncogenes and growth factors is tightly regulated to maintain accurate protein expression levels. However, this process is broken in cancer cells, which rely on the maintenance of high dosage of oncogenic proteins for their survival and to drive cancer development. Whether this is regulated at the level of translational control and the key factors in cis and trans remain unknown. The Myc oncogene is a central paradigm of an exquisitely regulated oncogene and a major driver of pancreatic ductal adenocarcinoma (PDAC). While there has been a tremendous effort to target Myc in cancer, clinically relevant Myc inhibition remains a challenge. Therefore, identifying selective promoters of MYC translation may present a novel therapeutic opportunity. Employing a genome-wide CRISPRi screen for modulators of MYC mRNA translation initiation in PDAC cells, we identified regulators of selective MYC translation and validated four RNA binding proteins (RBPs), including epitranscriptome modifiers. We focused on a top hit, a little studied RBP, RBM42, which is highly expressed in PDAC and predicts poor patient survival. We found that RBM42 binds and selectively regulates the translation of MYC and a precise, yet vital suite of pro-oncogenic transcripts, including JUN and EGFR. Mechanistically, employing immunoprecipitation-mass spectrometry analysis, we found that RMB42 is a novel ribosome-associated protein (RAP). Through RNA structure determination and mutagenesis analysis, we showed that RBM42 directly binds and remodels the MYC 5’UTR RNA structure, facilitating the formation of the translation pre-initiation complex and increasing MYC translation. Importantly, we demonstrated that RBM42 is necessary for human PDAC cell growth and fitness and PDAC tumorigenesis in xenograft mouse models in a Myc-dependent manner in vivo. In PDAC patient samples, RBM42 expression is correlated with Myc protein levels and transcriptional activity. This work transforms our understanding of the translational code in cancer and offers a new therapeutic opening to target the expression of oncogenes.

Citation Format: Joanna R Kovalski, Goksu Sarioglu, Vishvak Subramanyam, Grace Hernandez, Gilles Rademaker, Juan A Oses-Prieto, Macey Slota, Kwun Wah Wen, Grace E Kim, Alma L Burlingame, Hani Goodarzi, Rushika M Perera, Davide Ruggero. Functional screen for mediators of onco-mRNA translation specificity in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A078.
Abstract A081: Single-cell transcriptomic analysis of organoids derived from normal pancreata reveals epithelial cell heterogeneity and expression of PanIN-associated genes
Authors: Bray Alexander W , Carpenter Eileen S , Frankel Timothy L , Pasca di Magliano Marina
Published: September 2024

Pancreatic intraepithelial neoplasia (PanIN) lesions are the microscopic pre-malignant precursor to pancreatic ductal adenocarcinoma (PDAC). Animal models have demonstrated that PanINs arise from acinar cells which have undergone acinar-to-ductal metaplasia (ADM), but the exact molecular mechanisms which govern this transformation remain poorly understood. Novel in vitro models of PanIN genesis are necessary to better characterize pancreatic carcinogenesis and identify clinically actionable biomarkers of early disease. Organoids cultured from normal human pancreata have generated promise in this regard, but the cellular identity and malignant potential of their component cells have not been comprehensively characterized. We performed single-cell RNA sequencing (10X Genomics) on 6 normal pancreatic organoid lines maintained in culture for 3 weeks as well as their parent pancreata obtained from donors with no history of PDAC or other exocrine pancreatic pathology. Integration of whole pancreatic and organoid data using the R package Seurat generated a UMAP with clearly defined “ductal” and “acinar” clusters based on marker gene expression. The majority of organoid cells (47.3-81.5%) colocalized to the “ductal” cluster, and expressed SOX9, KRT19, and PDX1. However, a subset of cells (1.0-33.3%) present in each organoid sample aligned with the “acinar” cluster and expressed PRSS1, PRSS3, REG3A, MUC1, and SPINK1. In accordance with prior reports, organoid cells did not meaningfully express the acinar/endocrine markers PTF1A, CPA1, CHGA, or INS. Further examination of the “acinar” organoid cells revealed heightened expression of CLDN18, TFF1, TFF2, CTSE, CA2, and LYZ, genes associated with PanIN in prior spatial RNA sequencing studies. Other genes significantly enriched in this “acinar” organoid cluster include MUC5AC, a known PanIN marker, OLFM4, a tumor suppressor implicated in intestinal carcinogenesis, and the chemokine CXCL17, which is expressed in cells undergoing ADM. Overall, this data suggests that organoids derived from normal pancreata represent a heterogeneous population of epithelial cells with both acinar and ductal features. Furthermore, these organoids may represent an in vitro model of ADM and/or early PanIN formation, though additional functional mechanistic studies are required for confirmation.

Citation Format: Alexander W Bray, Eileen S Carpenter, Timothy L Frankel, Marina Pasca di Magliano. Single-cell transcriptomic analysis of organoids derived from normal pancreata reveals epithelial cell heterogeneity and expression of PanIN-associated genes [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A081.
Abstract A082: Defining the lysosome proteome during pancreatic cancer tumor evolution and metastasis
Authors: Nguyen Thuy T.P , Hernandez Grace A. , Rademaker Gilles , Luy Matthew , Li Alexander , Qiu Longhui , Paulo Joao A. , Perera Rushika M. , Mancias Joseph D.
Published: September 2024

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a five-year survival rate of only 13%, characterized by rapid proliferation, high metastatic potential, and resistance to conventional therapies. PDAC tumors are heavily reliant on nutrient-scavenging pathways such as autophagy and the lysosome – a degradative organelle that plays an essential role in the digestion and recycling of diverse cellular material. Despite their importance, the specific function of lysosomes over the course of tumor evolution and metastasis in PDAC tumors remains poorly understood. To understand the function of the lysosome in PDAC, we utilize a biochemical method known as LysoIP-based proteomics, which isolates intact lysosomes for subsequent analysis using mass spectrometry-based proteomics. This approach allows for a comprehensive analysis of lysosomal composition that can uncover unique features and functions of PDAC lysosomes that promote cellular adaptation to stress in support of tumor growth. A limitation of lysosome isolation from 2D cultured cells is that it does not recapitulate lysosomal features associated with tumors growing in vivo. To address this limitation, we generated a genetically engineered mouse model of PDAC incorporating conditional expression of ‘LysoTag’ (TMEM192-mRFP-3xHA) that enables capture and profiling of lysosomes at different stages of tumor evolution, including early-stage, late-stage, and metastatic disease. Employing this strategy, we successfully captured lysosomal proteins from primary tumors (pancreas), as well as lung and liver metastases. Our preliminary analysis revealed an enrichment of resident lysosomal membrane and luminal proteins in the datasets, confirming the successful isolation of intact lysosomes from these complex tissues. Additionally, we identified non-resident lysosomal proteins (termed cargo) that are enriched and co-evolved with advancing tumor stages. Notably, our results reveal distinct differences in the lysosomal proteome of tumor cells across different tissues. These findings emphasize the critical role of lysosomal function in tumor progression and suggest that targeting lysosomal components could be a promising therapeutic strategy for treating metastatic PDAC.

Citation Format: Thuy T.P Nguyen, Grace A. Hernandez, Gilles Rademaker, Matthew Luy, Alexander Li, Longhui Qiu, Joao A. Paulo, Rushika M. Perera, Joseph D. Mancias. Defining the lysosome proteome during pancreatic cancer tumor evolution and metastasis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A082.
Abstract A083: Genomic Analysis of Structural Variations in Pancreatic Cancer Using Long-Read Sequencing
Authors: Du Yongxing , Zheng Xiaohao , Duan Yunjie , Wang Chengfeng
Published: September 2024

Objective: Structural variations (SVs) constitute the greatest source of genomic alterations leading to the oncogenesis of many cancers, including the deadliest pancreatic cancer (PC). However, SVs in PC remain largely undefined due to technological limitations of the traditional short-read sequencing. This study aims to establish a comprehensive signature of genomic SVs in PC. Methods: PacBio whole genome long-read sequencing was employed to investigate the genomic landscape of SVs in eight tissues from four pancreatic cancer patients. A variety of SVs callers, including pbsv, Sniffles, cuteSV, and svim, were comprehensively adopted to improve the accuracy of SVs detection. Many undefined characteristics of germline and somatic SVs were revealed by comparing with multiple previously published databases. Besides, the impacts of structural variations on 3D chromatin organization were analyzed by integrating multi-omics data including Hi-C, RNA seq and ChIP seq. Results: A total of 26,844 and 27,028 non-redundant SVs were identified in human pancreatic cancer and matched para-cancerous tissues, respectively. Notably, among these SVs, 23.97% were novel and had not been previously reported. Germline SVs were further characterized and found enriched in 25 well-known susceptible genes, such as TP63, PVT1, and ABO. Additionally, we proposed a panel of germline SVs with predicted highest pathogenicity, involving STOX1, DSPP, and WRN. Totally, 616 somatic SVs were identified and presented high burdens in patients with lymph node metastasis. We observed that somatic DELs and INSs were significantly more prevalent in repetitive regions than in non-repetitive regions. Interestingly, DELs showed a decrease in the proportion occurring in simple repeat regions but an increase in the proportion occurring in LINE and SINE regions compared to INSs. Moreover, it was observed that deletions within topologically associated domains were associated with local enhanced interactions and disrupted chromatin loops, which might influence H3K27ac modification of histone contributing to enhancer or silencer hijacking. Conclusion: This study provides insights into the genomic signature of SVs in human pancreatic cancer. These findings might shed new light on the complexity of SVs and their pathogenesis by interplaying with chromatin organization.

Citation Format: Yongxing Du, Xiaohao Zheng, Yunjie Duan, Chengfeng Wang. Genomic Analysis of Structural Variations in Pancreatic Cancer Using Long-Read Sequencing [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A083.
Abstract A084: CDK12 inhibition drives mitotic catastrophe, triggering genome instability and innate immune response in pancreatic cancer
Authors: Iwadate Dosuke , Yamamoto Keisuke , Kato Hiroyuki , Tateishi Keisuke , Fujishiro Mitsuhiro
Published: September 2024

Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with limited therapies. In our screening using patient-derived organoids, we identified cyclin-dependent kinase 12 (CDK12) as a promising target in a subset of PDAC. Both pharmacological and genetic inhibition of CDK12 exerted potent antitumor activity in vitro and in vivo, inducing double-strand DNA breaks and apoptosis. CDK12 is a member of cyclin-dependent kinases that is involved in gene expression regulations at multiple levels. Recently, CDK12 has gained attention as a therapeutic target in several types of cancers including triple-negative breast cancer and Ewing sarcoma. However, little is known about the impact and possibility of CDK12 inhibition in pancreatic cancer treatment. As the lack of specific inhibitors that selectively target CDK12, studying CDK12 has long been challenging. To overcome this, we used CRISPR-Cas9 mediated genome editing to generate FKBP12F36V (dTAG) knock-in PDAC cell lines, enabling rapid elimination of endogenous CDK12 using a degrader against dTAG (dTAG-13). Quantitative-image-based cytometry and time lapse imaging showed that CDK12 ablation induces mitotic arrest, DNA damage and apoptosis (mitotic catastrophe). Specifically, loss of CDK12 kinase activity in the S phase was sufficient to slow DNA replication speed, leading to spindle assembly checkpoint activation and mitotic catastrophe. Importantly, cells that escaped mitotic death accumulate micronuclei, triggering immune response signaling that is mediated by the cGAS-STING-type I interferon pathway. Our results suggest that CDK12 is a promising target in a subset of PDAC patients, where CDK12 inhibition exerts antitumor activity both in cell autonomous and non-cell autonomous ways through inducing mitotic catastrophe as well as triggering immune response.

Citation Format: Dosuke Iwadate, Keisuke Yamamoto, Hiroyuki Kato, Keisuke Tateishi, Mitsuhiro Fujishiro. CDK12 inhibition drives mitotic catastrophe, triggering genome instability and innate immune response in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A084.
Abstract B002: Tumor-secreted PTHrP facilitates pancreatic cancer cachexia by regulation of de novo lipogenesis pathway
Authors: Bhalerao Nikita , Peura Jessica , Ogoti Yamini , Johnson Calvin , Chen Qingbo , Korobkina Ekaterina , Keller Faith , Wengyn Maximillan , Norgard Robert , Kremmer Richard , Watson Emma , Ruscetti Marcus , Guertin David , Pitarresi Jason R
Published: September 2024

Pancreatic cancer is associated with one of the highest and most severe forms of cancer- associated cachexia amongst solid tumors. In an effort to further our understanding of the molecular etiology of pancreatic cancer-associated cachexia, we have developed mouse models to study adipose tissue wasting during Pancreatic Ductal Adenocarcinoma (PDAC) progression. In these pursuits, we have identified a pro-cachectic factor, PTHrP, that is produced and secreted by PDAC tumor cells and directly signals to adipocytes to facilitate wasting. Genetic deletion and pharmacological inhibition of PTHrP significantly extended the survival of tumor-bearing animals and dramatically reduced cachectic phenotypes in these animals, manifesting as decreased adipose and muscle tissue wasting. Mechanistic studies have shown crosstalk between tumor-cell derived PTHrP and adipose tissue PTH1R that drives the cachectic phenotype. Finally, bulk RNA sequencing analysis in PTHrP-driven cachectic models point towards a role for reduced de novo lipogenesis and adipogenesis in cachectic adipose depots, suggesting potential rewiring of the metabolic profile of white adipose tissue in tumor-bearing mice with higher PTHrP expression.

Citation Format: Nikita Bhalerao, Jessica Peura, Yamini Ogoti, Calvin Johnson, Qingbo Chen, Ekaterina Korobkina, Faith Keller, Maximillan Wengyn, Robert Norgard, Richard Kremmer, Emma Watson, Marcus Ruscetti, David Guertin, Jason R Pitarresi. Tumor-secreted PTHrP facilitates pancreatic cancer cachexia by regulation of de novo lipogenesis pathway [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B002.
Abstract B007: Optimization of an Orthotopic Mouse Model of Pancreatic Cancer to Simulate Liver Metastasis and Cancer Cachexia
Authors: Wang David J , Spadafora Victoria , Pryce Benjamin R , Oles Alexander , Talbert Erin E , Ostrowski Micheal C , Guttridge Denis C
Published: September 2024

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease with a poor prognosis due to two key features. The first is metastasis at presentation, occurring in approximately 80% of PDAC patients, with 90% of these metastases occurring in the liver. The second is cachexia, which compromises treatment tolerance and reduces the quality of life for patients. Although various mouse models of PDAC exist, recapitulating both metastatic and cachectic features have been challenging. In this study, we optimized an orthotopic mouse model of PDAC by altering implantation sites, subcloning parental murine PDAC cells, adjusting the number of transplanted cells, and varying the age of recipient mice to study liver metastasis and cancer cachexia. These modifications significantly increased the latency of tumor development and the time course of the disease, concurrently increasing the rate of liver metastasis to approximately 70%. Furthermore, reliable cachexia endpoints were also achieved in both PDAC mice with and without metastases, mirroring the condition in patients. We also found that cachectic muscles from PDAC mice with metastasis exhibit a similar transcriptional profile to muscles derived from mice and patients without metastasis. Together, this model is likely to be advantageous in both advancing our understanding of the mechanisms of PDAC cachexia and in evaluating novel therapeutics.

Citation Format: David J Wang, Victoria Spadafora, Benjamin R Pryce, Alexander Oles, Erin E Talbert, Micheal C Ostrowski, Denis C Guttridge. Optimization of an Orthotopic Mouse Model of Pancreatic Cancer to Simulate Liver Metastasis and Cancer Cachexia [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B007.
Abstract B008: Twist1 drives pancreatic ductal adenocarcinoma-mediated muscle cachexia
Authors: Parajuli Parash , Atfi Azeddine
Published: September 2024

Cancer cachexia is a debilitating syndrome that affects the survival and quality of life of cancer patients. Up to 95% of pancreatic ductal adenocarcinoma (PDAC) patients experience muscle cachexia, yet the underlying mechanisms remain poorly understood. The transcription factor Twist1 plays essential roles in mesenchymal stem cell fate determination, and its deregulation contributes to cancer progression and metastasis. In this study, we report on the serendipitous finding that Twist1 functions in PDAC to drive muscle cachexia. We found that conditional overexpression of Twist1 in the pancreatic epithelium (pTwist1) during early development resulted in severe growth retardation, which was mainly characterized by muscle hypotrophy. This intriguing observation prompted us to inquire whether pTwist1 could function in the pancreatic epithelium to regulate muscle mass, and if so, whether this has any implication as it relates to PDAC-mediated muscle cachexia. We found that enforced expression of pTwist1 in healthy adult animals was sufficient to elicit the cardinal features of muscle cachexia, including progressive decline in body weight, physical activity, muscle force, and muscle mass. In the context of PDAC, enforced expression of pTwist1 in a KrasG12D background led to a dramatic acceleration of tumor progression, and this was associated with severe muscle cachexia. To the best of our knowledge, these findings provide the first evidence that the pro-malignant transcription factor Twist1 functions in PDAC to drive muscle cachexia, thus revealing new insights into the underlying mechanisms of muscle that could be harnessed to curb this debilitating syndrome.

Citation Format: Parash Parajuli, Azeddine Atfi. Twist1 drives pancreatic ductal adenocarcinoma-mediated muscle cachexia [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B008.
Abstract B014: NF-kB Regulates Innate Immunity in the Muscle Microenvironment to Control Distinct States of Wasting in Pancreatic Cancer-Induced Cachexia
Authors: Guttridge Denis C , Pryce Benjamin R , Oles Alexander , Talbert Erin E , Morgan Katherine A , Mahvi David A , Ostrowski Michael C , Zimmers Teresa A , Tidball James G , Wang David J
Published: September 2024

Cancer cachexia is a debilitating syndrome characterized by unintentional weight loss largely due to the depletion of adipose and skeletal muscle mass. Cachexia occurs in at least half of all patients with cancer, with increasing incidences in more advanced cases, and is estimated to be responsible for greater than 20% of all cancer related deaths. In pancreatic ductal adenocarcinoma (PDAC), which has a 5-year survival rate of 13%, the incidence of cachexia can be as high as 80% and is associated with poor disease outcomes. Mitigating the effects of cachexia has the potential to increase both quality of life and survival for patients with PDAC. However, there are currently no effective means of treating cachexia, thereby underscoring the need to further understand the pathology of this disease. Although cancer cachexia is classically characterized as a systemic inflammatory disorder, emerging evidence indicates that weight loss also associates with local tissue inflammation. We queried the regulation of this inflammation and its causality to cachexia by exploring skeletal muscle, whose atrophy strongly associates with poor outcomes. Using both a genetically engineered mouse model of PDAC- induced cachexia named KPP and patient samples, we show that cachectic muscle is marked by enhanced innate immunity. Accumulation of macrophages is regulated by an NF-kB activity that localizes to multiple cells within the muscle microenvironment. In addition, this accumulation of macrophages derives from an equal contribution of infiltrating monocytes and resident cells. Moreover, NF-kB activated cells and macrophages undergo a crosstalk; whereas NF-kB+ cells signal and recruit macrophages to inhibit muscle regeneration and promote atrophy, but interestingly at the same time can also protect myofibers, macrophages signal back to NF-kB+ cells to sustain an innate immune response in a feed-forward loop. Together, we propose that NF-kB functions in multiple cells in the muscle microenvironment to regulate innate immunity that both promotes and protects against muscle wasting in cancer.

Citation Format: Denis C Guttridge, Benjamin R Pryce, Alexander Oles, Erin E Talbert, Katherine A Morgan, David A Mahvi, Michael C Ostrowski, Teresa A Zimmers, James G Tidball, David J Wang. NF-kB Regulates Innate Immunity in the Muscle Microenvironment to Control Distinct States of Wasting in Pancreatic Cancer-Induced Cachexia [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B014.
Abstract B015: Duodenal aspiration biopsy in diagnosis of pancreatic neoplasms
Authors: Kashintsev Aleksei , Proutski Vitali
Published: September 2024

Introduction Intraductal papillary mucinous neoplasms (IPMNs) are a subset of lesions with elevated malignant potential, warranting their inclusion in screening standards. Despite endoscopic ultrasound (EUS) with fine-needle aspiration biopsy (FNAB) serving as the standard for verification, the occurrence of erroneous conclusions remains significant. Consequently, analysis of pancreatic juice is considered as a potential adjunctive diagnostic method. However, the absence of methods for obtaining pancreatic juice in adequate volume and quality poses a barrier to the widespread adoption of this approach.Materials and MethodsA method of duodenal aspiration biopsy (DAB) was developed involving active aspiration from the lumen of duodenum, yielding large volume of the mix of pancreatic juice and bile uncontaminated by gastric juice. Patients diagnosed with pancreatic neoplasms utilizing EUS with FNAB, were enrolled and ALB was implemented as an adjunctive diagnostic procedure. Cytological analysis of the material obtained was performed along with measuring the level of carcinoembryonic antigen (CEA). The level >192 ng/mL was considered significant, according to the Fukuoka 2017 guidelines.ResultsIn patients with cystic neoplasms of the pancreas undergoing DAB, results of cytological examination were non-informative in certain cases. However, analysis of CEA levels in the pancreatic juice and bile mixture always demonstrated strong concordance with the degree of dysplasia. In some instances, elevated CEA values facilitated the adoption of a more aggressive treatment strategy, leading to the detection of carcinoma. Conversely, low CEA levels prompted the decision to opt for observation rather than radical surgery in other cases.ConclusionDiagnosing precancerous transformations of the pancreas remains a challenging task, characterized by a notable rate of false-positive or false-negative conclusions. The DAB method offers a promising solution, seamlessly integrating into routine clinical practice and offering supplementary data to enhance the verification of pancreatic cancer.

Citation Format: Aleksei Kashintsev, Vitali Proutski. Duodenal aspiration biopsy in diagnosis of pancreatic neoplasms [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B015.
Abstract B021: Development of an ultrasensitive sequencing platform for blood-based mutation profiling of pancreas cancer patients
Authors: Ramaker Ryne , Abbruzzese James , Counter Chris
Published: September 2024

Introduction: One barrier to improving Pancreatic Ductal Adenocarcinoma (PDAC) patient outcomes is the absence of effective, non-invasive biomarkers for disease screening, treatment monitoring, and identification of therapeutic targets. While sequencing-based approaches to detect mutant circulating tumor DNA (ctDNA) in blood have transformed clinical management in several cancer types, use of these ‘liquid biopsies’ has lagged behind in PDAC due to low shedding of PDAC tumor cells. Nevertheless, intact circulating tumor cells have been documented in even the earliest stages of PDAC and precede the limits of disease detection on imaging or gross histology in mouse models. This suggests that circulating tumor cells could server as a PDAC biomarker if the sensitivity of sequencing could be increased. Methods: Point mutations in the gene KRAS are found in up to 95% of PDAC patients and hence represent a potential biomarker target for this cancer. To detect these pathogenic mutations in KRAS, we will adapt Maximum Depth Sequencing (MDS) for exon 2 of this gene by testing various primers and reaction conditions. The sensitivity of the K-MDS assay was optimized for blood by spiking PDAC cells with known KRAS mutations into healthy control blood at various dilutions. Finally, to evaluate the clinical sensitivity, we have collected paired blood draws from 200 advanced PDAC patients in prospective fashion, one to be tested with a commercial ctDNA test and one by K-MDS. Results: We optimized MDS to sequence exon 2 of KRAS and successfully detected a single mutant template in >107recovered independently barcoded templates (e.g. 107 individual KRAS alleles). We then optimized the assay for human blood and demonstrated that K-MDS could reproducibly detect a KRAS mutation corresponding to PDAC cells spiked into healthy control whole blood at the lowest dilution tested, namely one cell per ml of blood. This corresponds to a sensitivity of one mutated fragment per one million KRAS fragments, or 1000x more sensitive than currently available commercial assays. We are currently performing K-MDS on prospective PDAC cohort, with completion of this study expected this fall. Conclusions: We developed a highly sensitive sequencing assay cable of detecting PDAC ctDNA in whole blood. With successful validation in PDAC blood samples, this assay could provide a powerful tool for non-invasive PDAC mutation profiling and disease monitoring.

Citation Format: Ryne Ramaker, James Abbruzzese, Chris Counter. Development of an ultrasensitive sequencing platform for blood-based mutation profiling of pancreas cancer patients [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B021.