Overall Survival In Pancreatic Ductal Adenocarcinoma Research Articles

Abstract B070: TGFB2 mRNA levels prognostically interact with Interferon-alpha receptor activation of IRF9 and IFI27, and makers for tumor-associated macrophages impacting overall survival in PDAC

Abstract Background: The treatment of pancreatic ductal adenocarcinoma (PDAC) is an unmet challenge, with the median overall survival rate remaining less than a year, even with the use of FOLFIRINOX-based therapies. Methods: In this study, we analyzed archived macrophage- associated mRNA expression using datasets deposited in the UCSC Xena web platform to compare normal pancreatic tissue and PDAC tumor samples. We also utilized TCGA datasets deposited in cBioportal (https://www.cbioportal.org/study/summary?id=paad_tcga_pan_can_atlas) and KMplotter (http://kmplot.com/analysis/index.php?p=service&cancer=pancancer_rnaseq) to relate mRNA expression levels to overall survival outcomes. The prognostic impact of TGFB2 mRNA expression levels was determined by comparing patients expressing high versus low levels of TGFB2 (50th percentile cut-off) in low macrophage TME. Results: The TGFB2 gene exhibited low mRNA expression levels in normal tissue, with less than one TPM, while, in tumor tissue, TGFB2 expression levels exhibited a 7.9-fold increase in mRNA expression relative to normal tissue (P < 0.0001). Interferon Alpha Inducible Protein 27 (IFI27), the downstream product of Interferon regulatory factor 9 (IRF9) and Signal transducer and activator of transcription 1 (STAT1) transcriptional activation by Interferon alpha/beta receptor 1 (IFNAR1) exhibited a significant 66.3-fold increase in expression in tumor tissue compared to normal tissue (P < 0.0001). The statistical contrast between the expression of IRF9 mRNA also displayed a significant (P<0.0001) 4.2-fold increase in mRNA expression levels. These increased levels of mRNA expression were found to be prognostically significant, whereby patients with high expression levels of either TGFB2, IRF9, or IFI27 showed median OS times ranging from 16 to 20 months (P<0.01 compared to 72 months for patients with low levels of expression for both TGFB2 and either IRF9 or IFI27). In TME with low macrophage levels, patients with high levels of TGFB2 mRNA exhibited significantly shorter OS outcomes than patients with low TGFB2 mRNA levels (Median OS of 72.2 versus 15.3 months, P<0.0001). Furthermore, expression of TGFB2 mRNA and TAM markers exhibited TGFB2 prognostic impact independent of 17 TAM markers suggested by multivariate Cox regression models. Conclusions: The results of our study suggest that a combination of pharmacological tools can be used in treating PDAC patients, targeting both TGFB2 and the components of the type-I interferon signaling pathway to improve OS outcomes. TGFB2 expression impacts OS independent of TAM markers, but the expression of TAM markers impacts the prognostic effect of TGFB2 mRNA expression. Citation Format: Sanjive Qazi, Wen-Han Chang, Cynthia Lee, Vuong Trieu. TGFB2 mRNA levels prognostically interact with Interferon-alpha receptor activation of IRF9 and IFI27, and makers for tumor-associated macrophages impacting overall survival in PDAC [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 B070.

Abstract C010: Single-cell profiling unleashes KRAS-driven redrafting of the tumor microenvironment before cancer onset

Abstract Pancreatic cancer is a highly lethal and aggressive disease, commonly characterized by a unique tumor microenvironment, where mutant KRAS signaling plays a pivotal role in its pathophysiology. To deepen our understanding of KRAS-dependent signaling in earliest tumor development, oncogenic KRAS (KRASG12D) expression was induced in human stem cell-derived pancreatic ductal-like organoids (PDLOs), followed by time-resolved single-cell RNA and bulk ATAC sequencing. This unique precancerous state model facilitated a comprehensive characterization of early oncogenic events at cellular resolution. Furthermore, a machine learning approach was applied to accurately predict KRAS-dependent cell identities. Within seven days, mutated KRAS triggers distinct gene signature programs related to extracellular matrix remodeling and inflammation. Although derived from dysplastic yet non-cancerous states, these gene signatures negatively correlated with overall survival in PDAC (pancreatic ductal adenocarcinoma) patients. The KRAS-dependent signatures retrieved with the machine learning approach allowed the grouping of 24 published PDAC cases, showing differences in niche composition and communication patterns. Various co-culture systems demonstrated that KRASG12D-expressing PDLOs could activate pancreatic stellate cells, inducing a cancer- associated fibroblast (CAF)-like state. Simultaneously, these KRAS-activated organoids were protected from T cell infiltration in vitro. Furthermore, in silico approaches were employed to reconstruct a virtual pancreatic cancer space, demonstrating that our PDLOs transcriptionally bridge healthy and malignant ductal states. This also revealed potential KRAS-driven pathways mediating CAF activation and T cell shielding preceding cancer onset. In summary, our human PDLO model enables us to explore the transition from normal to cancerous states, addressing gaps in our understanding of tumor development and tumor niche preparation. Citation Format: Chantal Allgoewer, Markus Breunig, Meike Hohwieler, Medhanie Mulaw, Alexander Kleger. Single-cell profiling unleashes KRAS-driven redrafting of the tumor microenvironment before cancer onset [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 C010.

Abstract B079: Disassembly of embryonic keratin filaments promotes pancreatic cancer metastases

Abstract Keratin 17 (K17), an intermediate filament protein typically expressed during embryonic development, is a hallmark of one of the most aggressive pancreatic ductal adenocarcinoma (PDAC) subtypes. Its expression is also associated with poor patient survival and resistance to first-line therapies. However, the mechanistic roles of K17 in malignancy remain largely unexplored. Here, we demonstrate that K17 expression and disassembly enhance tumor growth, increase metastatic potential, and shorten overall survival. Using the Clinical Proteomic Tumor Analysis Consortium (CPTAC) stratified by K17 phosphorylation status, we found that increased K17 phosphorylation correlated with worse survival. Mass spectrometry of untreated PDAC patient samples identified a phosphorylation hotspot at serines 10-13, which is conserved across species and type I keratin proteins. We found that K17 phosphorylation, mediated by the PKC/MEK/RSK pathway, led to increased disassembly and nuclear translocation in pancreatic cancer cells. Mice with phosphomimetic mutations at the serine hotspot showed increased metastases and decreased survival compared to wild-type and phosphorylation-resistant K17. Lastly, detergent-soluble nuclear K17 promoted metastasis- related genes in both patient and murine tumors. These findings suggest that disassembly of K17 mediated by phosphorylation at serines 10-13 is sufficient to promote metastases and decrease overall survival in PDAC. This opens the door for further investigation into K17 phosphorylation as a novel therapeutic target to enhance PDAC patient survival, especially in those with more aggressive, chemoresistant subtypes. Citation Format: Deanne E Yugawa, Ryan Kawalerski, Mariana Torrente Gonçalves, Chun-Hao Pan, Robert Tseng, Lucia Roa-Pena, Cindy V Leiton, Luke A Torre-Healy, Taryn Boyle, Sumedha Chowdhury, Natasha T Snider, Kenneth R Shroyer, Luisa F Escobar-Hoyos. Disassembly of embryonic keratin filaments promotes pancreatic cancer metastases [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 B079.

Analysis of PD-L1 promoter methylation combined with immunogenic context in pancreatic ductal adenocarcinoma

Despite the successful application of programmed cell death ligand 1 (PD-L1)-blocking strategies in some types of cancers and well-established prognostic indicators in pancreatic ductal adenocarcinoma (PDAC), the biological and clinical implications of the methylation status of PD-L1/PD-L2 in PDAC remain largely unknown. Therefore, this study aimed to explore the biological role of PD-L1/PD-L2 methylation and its association with clinicopathological features, clinical outcomes, and the immune microenvironment by analyzing the data on PD-L1/PD-L2 methylation and mRNA expression in PDAC cohorts obtained from the Cancer Genome Atlas and International Cancer Genome Consortium. The correlation between PD-L1 promoter methylation and PD-L1 expression and survival was further validated in an independent validation cohort (Peking Union Medical College Hospital [PUMCH] cohort) using pyrosequencing and immunohistochemistry. These results demonstrated that hypomethylation of the PD-L1 promoter was strongly associated with upregulated PD-L1 expression and shorter overall survival in PDAC. Multivariate Cox regression analyses revealed that the PD-L1 promoter methylation was an independent prognostic factor. PD-L1 promoter hypomethylation and high expression were related to aggressive clinical phenotypes. Moreover, both PD-L1 and PD-L2 methylation correlated with immune cell infiltration and the expression of immune checkpoint genes. PD-L1 promoter methylation status was further validated as an independent prognostic biomarker in patients with PDAC using the PUMCH cohort. The prognostic significance of PD-L1 promoter methylation was more discriminative in tumors with perineural/lymphovascular invasion and distant metastasis than in those without perineural/lymphovascular invasion and distant metastasis. In summary, the methylation status of the PD-L1 promoter is a promising biomarker for survival outcomes, immune infiltration, and the potential immune benefits of immunotherapy in PDAC.

Concurrent inhibition of pBADS99 synergistically improves MEK inhibitor efficacy in KRASG12D-mutant pancreatic ductal adenocarcinoma

Therapeutic targeting of KRAS-mutant pancreatic ductal adenocarcinoma (PDAC) has remained a significant challenge in clinical oncology. Direct targeting of KRAS has proven difficult, and inhibition of the KRAS effectors have shown limited success due to compensatory activation of survival pathways. Being a core downstream effector of the KRAS-driven p44/42 MAPK and PI3K/AKT pathways governing intrinsic apoptosis, BAD phosphorylation emerges as a promising therapeutic target. Herein, a positive association of the pBADS99/BAD ratio with higher disease stage and worse overall survival of PDAC was observed. Homology-directed repair of BAD to BADS99A or small molecule inhibition of BADS99 phosphorylation by NCK significantly reduced PDAC cell viability by promoting cell cycle arrest and apoptosis. NCK also abrogated the growth of preformed colonies of PDAC cells in 3D culture. Furthermore, high-throughput screening with an oncology drug library to identify potential combinations revealed a strong synergistic effect between NCK and MEK inhibitors in PDAC cells harboring either wild-type or mutant-KRAS. Mechanistically, both mutant-KRAS and MEK inhibition increased the phosphorylation of BADS99 in PDAC cells, an effect abrogated by NCK. Combined pBADS99-MEK inhibition demonstrated strong synergy in reducing cell viability, enhancing apoptosis, and achieving xenograft stasis in KRAS-mutant PDAC. In conclusion, the inhibition of BADS99 phosphorylation enhances the efficacy of MEK inhibition, and their combined inhibition represents a mechanistically based and potentially effective therapeutic strategy for the treatment of KRAS-mutant PDAC.

Tomatidine targets ATF4-dependent signaling and induces ferroptosis to limit pancreatic cancer progression

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with high metastasis and therapeutic resistance. Activating transcription factor 4 (ATF4), a master regulator of cellular stress, is exploited by cancer cells to survive. Prior research and data reported provide evidence that high ATF4 expression correlates with worse overall survival in PDAC. Tomatidine, a natural steroidal alkaloid, is associated with inhibition of ATF4 signaling in multiple diseases. Here, we discovered that invitro and invivo tomatidine treatment of PDAC cells inhibits tumor growth. Tomatidine inhibited nuclear translocation of ATF4 and reduced the transcriptional binding of ATF4 with downstream promoters. Tomatidine enhanced gemcitabine chemosensitivity in 3D ECM-hydrogels and invivo. Tomatidine treatment was associated with induction of ferroptosis signaling validated by increased lipid peroxidation, mitochondrial biogenesis, and decreased GPX4 expression in PDAC cells. This study highlights a possible therapeutic approach utilizing a plant-derived metabolite, tomatidine, to target ATF4 activity in PDAC.

NUAK1 governs centrosome replication in pancreatic cancer via MYPT1/PP1β and GSK3β-dependent regulation of PLK4.

The AMP-activated protein kinase (AMPK)-related kinase NUAK1 (NUAK family SNF1-like kinase 1) has emerged as a potential vulnerability in MYC-dependent cancer but the biological roles of NUAK1 in different settings are poorly characterised, and the spectrum of cancer types that exhibit a requirement for NUAK1 is unknown. Unlike canonical oncogenes, NUAK1 is rarely mutated in cancer and appears to function as an obligate facilitator rather than a cancer driver per se. Although numerous groups have developed small-molecule NUAK inhibitors, the circumstances that would trigger their use and the unwanted toxicities that may arise as a consequence of on-target activity are thus undetermined. Reasoning that MYC is a key effector of RAS pathway signalling and the GTPase KRAS is almost uniformly mutated in pancreatic ductal adenocarcinoma (PDAC), we investigated whether this cancer type exhibits a functional requirement for NUAK1. Here, we show that high NUAK1 expression is associated with reduced overall survival in PDAC and that inhibition or depletion of NUAK1 suppresses growth of PDAC cells in culture. We identify a previously unknown role for NUAK1 in regulating accurate centrosome duplication and show that loss of NUAK1 triggers genomic instability. The latter activity is conserved in primary fibroblasts, raising the possibility of undesirable genotoxic effects of NUAK1 inhibition.

Single-cell and bulk RNA sequencing identifies T cell marker genes score to predict the prognosis of pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is one of the lethal malignancies, with limited biomarkers identified to predict its prognosis and treatment response of immune checkpoint blockade (ICB). This study aimed to explore the predictive ability of T cell marker genes score (TMGS) to predict their overall survival (OS) and treatment response to ICB by integrating single-cell RNA sequencing (scRNA-seq) and bulk RNA-seq data. Multi-omics data of PDAC were applied in this study. The uniform manifold approximation and projection (UMAP) was utilized for dimensionality reduction and cluster identification. The non-negative matrix factorization (NMF) algorithm was applied to molecular subtypes clustering. The Least Absolute Shrinkage and Selection Operator (LASSO)-Cox regression was adopted for TMGS construction. The prognosis, biological characteristics, mutation profile, and immune function status between different groups were compared. Two molecular subtypes were identified via NMF: proliferative PDAC (C1) and immune PDAC (C2). Distinct prognoses and biological characteristics were observed between them. TMGS was developed based on 10 T cell marker genes (TMGs) through LASSO-Cox regression. TMGS is an independent prognostic factor of OS in PDAC. Enrichment analysis indicated that cell cycle and cell proliferation-related pathways are significantly enriched in the high-TMGS group. Besides, high-TMGS is related to more frequent KRAS, TP53, and CDKN2A germline mutations than the low-TMGS group. Furthermore, high-TMGS is significantly associated with attenuated antitumor immunity and reduced immune cell infiltration compared to the low-TMGS group. However, high TMGS is correlated to higher tumor mutation burden (TMB), a low expression level of inhibitory immune checkpoint molecules, and a low immune dysfunction score, thus having a higher ICB response rate. On the contrary, low TMGS is related to a favorable response rate to chemotherapeutic agents and targeted therapy. By combining scRNA-seq and bulk RNA-seq data, we identified a novel biomarker, TMGS, which has remarkable performance in predicting the prognosis and guiding the treatment pattern for patients with PDAC.

Prognostic value of diffuse reduction of spleen density on postoperative survival of pancreatic ductal adenocarcinoma: A retrospective study.

It is difficult to predict the prognosis of patients with pancreatic ductal adenocarcinoma (PDAC) before radical operation. The purpose of this study was to explore the connection between the diffuse reduction of spleen density on computed tomography (DROSD) and the postoperative prognosis of patients with PDAC. A total of 160 patients with PDAC who underwent radical surgery in the First Affiliated Hospital of Wenzhou Medical University were enrolled. Cox regression analysis was used to cast the overall survival (OS) and evaluate the prognostic factors. Nomogram was used to forecast the possibility of 1-year, 3-year, and 5-year OS. The prediction accuracy and clinical net benefit are performed by concordance index (C-index), calibration curve, time-dependent receiver operating characteristics (tdROC), and decision curve analysis. In multivariable Cox analysis, DROSD is independently related to OS. Advanced age, TNM stage, neutrophil/lymphocyte ratio, and severe complications were also independent prognostic factors. The calibration curves of nomogram showed optimal agreement between prediction and observation. The C-index of nomogram is 0.662 (95%CI, 0.606-0.754). The area under tdROC curve for a 3-year OS of nomogram is 0.770. DROSD is an independent risk factor for an OS of PDAC. We developed a nomogram that combined imaging features, clinicopathological factors, and systemic inflammatory response to provide a personalized risk assessment for patients with PDAC.

Development and external validation of a radiomics combined with clinical nomogram for preoperative prediction prognosis of resectable pancreatic ductal adenocarcinoma patients.

To develop and externally validate a prognosis nomogram based on contrast-enhanced computed tomography (CECT) combined clinical for preoperative prognosis prediction of patients with pancreatic ductal adenocarcinoma (PDAC). 184 patients from Center A with histopathologically confirmed PDAC who underwent CECT were included and allocated to training cohort (n=111) and internal validation cohort (n=28). The radiomic score (Rad - score) for predicting overall survival (OS) was constructed by using the least absolute shrinkage and selection operator (LASSO). Univariate and multivariable Cox regression analysis was used to construct clinic-pathologic features. Finally, a radiomics nomogram incorporating the Rad - score and clinical features was established. External validation was performed using Center B dataset (n = 45). The validation of nomogram was evaluated by calibration curve, Harrell's concordance index (C-index) and decision curve analysis (DCA). The Kaplan-Meier (K-M) method was used for OS analysis. Univariate and multivariate analysis indicated that Rad - score, preoperative CA 19-9 and postoperative American Joint Committee on Cancer (AJCC) TNM stage were significant prognostic factors. The nomogram based on Rad - score and preoperative CA19-9 was found to exhibit excellent prediction ability: in the training cohort, C-index was superior to that of the preoperative CA19-9 (0.713 vs 0.616, P< 0.001) and AJCC TNM stage (0.713 vs 0.614, P< 0.001); the C-index was also had good performance in the validation cohort compared with CA19-9 (internal validation cohort: 0.694 vs 0.555, P< 0.001; external validation cohort: 0.684 vs 0.607, P< 0.001) and AJCC TNM stage (internal validation cohort: 0.694 vs 0.563, P< 0.001; external validation cohort: 0.684 vs 0.596, P< 0.001). The calibration plot and DCA showed excellent predictive accuracy in the validation cohort. We established a well-designed nomogram to accurately predict OS of PDAC preoperatively. The nomogram showed a satisfactory prediction effect and was worthy of further evaluation in the future.

Integrated analysis of senescence-associated genes in pancreatic ductal adenocarcinoma.

Background: Cellular senescence plays a critical role in the occurrence and development, and immune modulation of cancer. This research primarily investigated the role of senescence-associated genes (SAGs) in the survival and tumor microenvironment of pancreatic ductal adenocarcinoma (PDAC). Methods: From the Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) database, the gene expression profiles and clinical data of PDAC samples were downloaded. SAGs in the TCGA cohort were used to build a novel prognostic model and validated in the ICGC cohort. The relationship of signature with the immune landscape, tumor mutational burden (TMB), as well as the sensitivity of different therapies, was explored. Moreover, a nomogram was developed to predict the overall survival of PDAC patients. Results: A prognostic signature was constructed on basis of three SAGs, and patients in the low-risk score group had a longer survival time. The accuracy of the signature to distinguish different score groups was confirmed through principal component analysis (PCA) and the Receiver operator curves curve. The mRNA expression of the three signature genes was also verified in normal pancreatic and PDAC cell lines by RT-qPCR. The signature could independently predict the prognosis of PDAC patients and had broad applicability. Meanwhile, the nomogram predicted that 1- and 3-years survival rates were in good agreement with the observed overall survival rates. Low-risk patients had lower tumor mutational burden, and low-TMB patients had a better prognosis. Low- and high-risk patients exhibit distinct immune cell infiltration and immune checkpoint changes. By further analyzing the risk score, patients in the low-risk group were more responsive to immunotherapy and a variety of commonly used chemotherapeutic drugs. Conclusion: The prognostic signature can well predict the prognosis and assess the possibility of immunotherapy in personalized PDAC treatment.

Abstract 1565: Targeting stromal-specific p38 MAPK signaling to stifle inflammatory reprogramming of cancer-associated fibroblasts in pancreatic cancer

Abstract Introduction: Major contributors to therapeutic resistance in pancreatic ductal adenocarcinoma (PDAC) are the desmoplastic stroma that acts as a barrier to drug delivery and effector immune cell infiltration. In our efforts to identify mediators of therapeutic resistance, we identified the pro-inflammatory cytokine interleukin-1α (IL-1) to be critical in the activation and reprogramming of neighboring cancer-associated fibroblasts (CAFs) within the stroma to a pro-inflammatory phenotype to promote myeloid cell chemotaxis. Our mechanistic studies have identified the p38 MAPK pathway as a novel mediator of IL-1-induced CAF activation from quiescent stellate cells into the inflammatory fibroblast. Our central hypothesis is that disruption of the IL-1/p38 MAPK signaling cascade in pancreatic stellate cells (PSCs) and CAFs can improve therapeutic resistance by remodeling the fibrotic stromal landscape and the overall immune microenvironment in PDAC tumors. Methods: Inhibition of phosphorylated p38 MAPK was achieved pharmacologically with Pexmetinib and genetically with an shRNA lentiviral system in CAF and PSC cell lines. Inflammatory PSC/CAF activation was determined by qPCR, immunofluorescence, and vitamin A assay. Ptf1acre/+;LSL-KrasG12D/+; Tgfbr2flox/flox (PKT) mice were treated with the p38 inhibitor, pexmetinib (30mg/kg, daily PO), or vehicle control for 2.5 weeks prior to sacrifice. For survival studies, PKT mice were treated with vehicle, gemcitabine (20 μg/twice weekly), pexmetinib, or combination until moribund. Results: Inhibition of p38 MAPK in PSCs prevented activation into an inflammatory fibroblast in vitro when stimulated with IL-1 or tumor cell cocultures. p38 MAPK inhibition in vivo resulted in signification reduction of PDGFR+ CAFs in a PDAC GEMM. Additionally, p38 MAPK inhibition resulted in a significant reduction of circulating myeloid cells (CD11b+) and intratumoral monocytic MDSCs (Ly6C+). p38 inhibition, in combination with chemotherapy, significantly improves overall survival in a PDAC GEMM. Conclusions: These findings provide important mechanistic data to explore p38 MAPK inhibition to target the fibrotic stroma and reduce immunosuppressive myeloid levels in tumors and provide compelling preclinical evidence to combine pexmetinib with chemotherapy to improve overall survival in PDAC. Citation Format: Samara Singh, Austin R. Dosch, Siddharth Mehra, Iago de Castro Silva, Anna Bianchi, Vanessa Tonin Garrido, Nilesh Deshpande, Zhiqun Zhou, Jashodeep Datta, Nagaraj Nagathihalli, Nipun Merchant. Targeting stromal-specific p38 MAPK signaling to stifle inflammatory reprogramming of cancer-associated fibroblasts in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1565.

Abstract 1328: Deciphering the role of CREB1 in shaping the tumor immune landscape of pancreatic ductal adenocarcinoma

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) remains a major therapeutic challenge with hallmarks including oncogenic mutations, desmoplastic architecture, and an immunosuppressive tumor microenvironment (TME). Previously, we have identified Cyclic AMP Response Element Binding protein 1 (CREB) as an oncogenic transcriptional factor downstream of KRAS that promotes disease aggressiveness, poor survival, and immune exclusion. Based on these, we sought to determine the impact of tumor intrinsic CREB deletion in shaping the tumor immune microenvironment. Methods: We have generated a novel genetically engineered mouse model (GEMM) of pancreas-specific CREB deletion (CREBfl/fl) in LSL-KrasG12D/+; Trp53 R172H/+; Pdx1Cre/+ (KPC) mice that phenocopy human PDAC disease. CRISPR/CAS9-based genomic editing was utilized to ablate CREB (CREBKO) in KPC tumor cells. RNA-sequencing analysis was performed in KPC CREB wild type (CREBEV) vs. CREBKO tumor cells to identify CREB-mediated transcriptomic changes. Syngeneic orthotopic tumor implantation of these cells was performed into the pancreata of mice. High dimensional immunophenotyping was accomplished to assess changes in the immune subsets with CREB deletion in murine tumors. Additionally, these tissues were processed for immunohistochemical and qPCR-based analysis to assess changes in fibroinflammatory and immune mediators. Results: Pancreas-specific CREB deletion in the KPC GEMM led to a significant reduction in the primary tumor burden, liver metastases and improved overall survival compared to wild-type KPC. Additionally, CREB deletion significantly remodeled the tumor stroma, as evidenced by the reduction in the expression of fibroinflammatory and immunosuppressive markers. In assessing the immune repercussions of CREB deletion in pancreatic tumors, we observed a decreased infiltration of CD11b+ myeloid-derived suppressor cells (MDSCs) and granulocytic-PMN MDSCs (CD11b+ Ly6Ghigh Ly6Clow F4/80-), with a concomitant increase in the antigen-presenting M1-like macrophages (F4/80+ MHC-II high CD86 high). Also, CREB ablation in these tumors further facilitated increased infiltration of activated effector CD8+ T cells resulted in enhanced anti-tumor immune response within the PDAC TME. Mechanistically, RNA transcriptomic and secretome analysis in CREBKO Vs. the wild type KPC tumor cells identified several differentially expressed immunomodulatory soluble mediators responsible for shaping CREB dependent immunogenic landscape in PDAC. Conclusion: Overall, depleting CREB reshapes the tumor immune landscape to reduce innate immunosuppressive myeloid infiltration and reinvigorate the antitumor T cell immune responses to improve overall survival in PDAC. Citation Format: Siddharth Mehra, Vanessa Garrido, Samara Singh, Iago D Silva, Luis Alberto Nivelo, Anna Bianchi, Shrey Modi, Zhiqun Zhou, Austin Dosch, Nilesh Deshpande, Supriya Srinivasan, Christine Rafie, Ifeanyichukwu Ogobuiro, Xi Chen, Alejandro Villarino, Jashodeep Datta, Nipun Merchant, Nagaraj Nagathihalli. Deciphering the role of CREB1 in shaping the tumor immune landscape of pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1328.

Abstract 1506: APOBEC3A a causative agent in cancer-related chromosomal instability and STING driven metastasis

Abstract Chromosomal instability (CIN), a hallmark of cancer, has been associated with a highly metastatic phenotype in numerous cancers. Mechanisms by which these genomic events occur during the lifespan of a neoplasm have yet to be fully delineated. In our study we identified a novel function for APOBEC3A (A3A), a primate-specific cytidine deaminase upregulated in multiple cancers including Pancreatic ductal adenocarcinoma (PDAC), in the initiation of CIN.We demonstrate that A3A is not routinely detected in non-diseased pancreata; however, expression was detected in the epithelial and stromal compartments from patients with chronic pancreatitis and in precursor lesions of PDAC and is further significantly increased in invasive neoplasia. Strikingly, we found that A3A expression but not A3A-induced mutagenesis, was associated with a significantly reduced overall survival in PDAC. By employing a variety of genetic tools, we identified a novel function for A3A in initiating CIN, underscoring its potential role in driving the observed early metastatic propensity in PDAC. Using a series of in vivo and in vitro models, we demonstrate that A3A-induced CIN leads to aggressive cancer, featuring enhanced, STING-dependent, distant organ seeding and metastatic growth. As a consequence of aberrant A3A function and CIN-mediated upregulation of STING, NfkB and Stat3 pathways were activated. More importantly, all these effects were independent of the deaminase domain, underscoring a novel role of A3A beyond its established canonical function. Remarkably, in a novel autochthonous murine model of PDA expressing human A3A, we identified numerous copy number changes, homologous to those altered in A3A overexpressing patient tumors, including deletions in DNA repair pathway genes. While we showed that A3A-expressing PDAC cells do not fully phenocopy a BRCA mutation-like phenotype, A3A-expressing PDAC cells are exceptionally sensitive to PARP inhibition and DNA cross linking agents, recapitulating a state of “chemical BRCAness”. The combined effect of deletions in these genes has yet to be ascertained, and we hypothesize that these losses further add to ongoing DNA damage overall in PDAC. In summary, our study shows, that A3A drives metastasis and specific deletions through a deaminase-independent initiation of CIN, with potential implications for targeted treatment strategies in pancreatic cancer. Citation Format: Sonja Maria Woermann, Amy Zhang, Fredrik I. Thege, Chris Gates, Reuben S. Harris, Susan R. Ross, Faiyaz Notta, Anirban Maitra, Andrew D. Rhim. APOBEC3A a causative agent in cancer-related chromosomal instability and STING driven metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1506.

Overexpression of integrin alpha 2 (ITGA2) correlates with poor survival in patients with pancreatic ductal adenocarcinoma

AimsDue to the known malignant potential and the poor overall prognosis of pancreatic ductal adenocarcinoma (PDAC), the identification of new biomarkers is of utmost importance. It has been reported that...

Progranulin mediates immune evasion of pancreatic ductal adenocarcinoma through regulation of MHCI expression

Immune evasion is indispensable for cancer initiation and progression, although its underlying mechanisms in pancreatic ductal adenocarcinoma (PDAC) are not fully known. Here, we characterize the function of tumor-derived PGRN in promoting immune evasion in primary PDAC. Tumor- but not macrophage-derived PGRN is associated with poor overall survival in PDAC. Multiplex immunohistochemistry shows low MHC class I (MHCI) expression and lack of CD8+ T cell infiltration in PGRN-high tumors. Inhibition of PGRN abrogates autophagy-dependent MHCI degradation and restores MHCI expression on PDAC cells. Antibody-based blockade of PGRN in a PDAC mouse model remarkably decelerates tumor initiation and progression. Notably, tumors expressing LCMV-gp33 as a model antigen are sensitized to gp33-TCR transgenic T cell-mediated cytotoxicity upon PGRN blockade. Overall, our study shows a crucial function of tumor-derived PGRN in regulating immunogenicity of primary PDAC.

Modulated Electro-Hyperthermic (mEHT) Treatment in the Therapy of Inoperable Pancreatic Cancer Patients-A Single-Center Case-Control Study.

Our present oncological treatment arsenal has limited treatment options for pancreatic ductal adenocarcinoma (PDAC). Extended reviews have shown the benefits of hyperthermia for PDAC, supporting the perspectives with the improvements of the treatment possibilities. METHODS: A retrospective single-center case-control study was conducted with the inclusion of 78 inoperable PDAC patients. Age-, sex-, chemotherapy-, stage-, and ascites formation-matched patients were assigned to two equal groups based on the application of modulated electro-hyperthermia (mEHT). The EHY2030 mEHT device was used. RESULTS: A trend in favor of mEHT was found in overall survival (p = 0.1420). To further evaluate the potential beneficial effects of mEHT, the presence of distant metastasis or ascites in the patients’ oncological history was investigated. Of note, mEHT treatment had a favorable effect on patients’ overall survival in metastatic disease (p = 0.0154), while less abdominal fluid responded to the mEHT treatment in a more efficient way (p ≤ 0.0138). CONCLUSION: mEHT treatment was associated with improved overall survival in PDAC in our single-center retrospective case-control study. The outcome measures encourage us to design a randomized prospective clinical study to further confirm the efficiency of mEHT in this patient cohort.

N6-methylandenosine-related lncRNAs play an important role in the prognosis and immune microenvironment of pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive, fatal tumor. N6-methylandenosine (m6A) methylation is the major epigenetic modification of RNA including lncRNAs. The roles of m6A-related lncRNAs in PDAC have not been fully clarified. This study aims to assess gene signatures and prognostic value of m6A-related lncRNAs in PDAC. The Cancer Genome Atlas (TCGA) dataset and the International Cancer Genome Consortium (ICGC) dataset were explored to identify m6A-related lncRNAs. Univariate, least absolute shrinkage and selection operator (LASSO) and multivariate Cox regression were performed to construct the m6A-related lncRNAs prognostic riskscore (m6A-LPR) model to predict the overall survival (OS) in the TCGA training cohort. Kaplan–Meier curve with log-rank test and receiver operating characteristic (ROC) curve were used to evaluate the prognostic value of the m6A-LPR. Furthermore, the robustness of the m6A-LPR was further validated in the ICGC cohort. Tumor immunity was evaluated using ESTIMATE and CIBERSORT algorithms. A total of 262 m6A-related lncRNAs were identified in two datasets. In the TCGA training cohort, 28 prognostic m6A-related lncRNAs were identified and the m6A-LPR including four m6A-related lncRNAs was constructed. The m6A-LPR was able to identify high-risk patients with significantly poorer OS and accurately predict OS in both the TCGA training cohort and the ICGC validation cohort. Analysis of tumor immunity revealed that high-risk groups had remarkably lower stromal, immune, and ESTIMATE scores. Moreover, high-risk groups were associated with significantly higher levels of plasma B cells and resting NK cells infiltration, and lower levels of infiltrating resting memory CD4 T cells, monocytes, and resting mast cells. Our study proposed a robust m6A-related prognostic signature of lncRNAs for predicting OS in PDAC, which provides some clues for further studies focusing on the mechanism process underlying m6A modification of lncRNAs.

Role of tumor mutation burden-related signatures in the prognosis and immune microenvironment of pancreatic ductal adenocarcinoma

BackgroundHigh tumor mutation burden (TMB) has gradually become a sensitive biomarker for predicting the response to immunotherapy in many cancers, including lung, bladder and head and neck cancers. However, whether high TMB predicts the response to immunotherapy and prognosis in pancreatic ductal adenocarcinoma (PDAC) remained obscure. Hence, it is significant to investigate the role of genes related to TMB (TRGs) in PDAC.MethodsThe transcriptome and mutation data of PDAC was downloaded from The Cancer Genome Atlas-Pancreatic Adenocarcinoma (TCGA). Five independent external datasets of PDAC were chosen to validate parts of our results. qRT-PCR and immunohistochemical staining were also performed to promote the reliability of this study.ResultsThe median overall survival (OS) was significantly increased in TMB_low group compared with the counterpart with higher TMB score after tumor purity adjusted (P = 0.03). 718 differentially expressed TRGs were identified and functionally enriched in some oncogenic pathways. 67 TRGs were associated with OS in PDAC. A prognostic model for the OS was constructed and showed a high predictive accuracy (AUC = 0.849). We also found TMB score was associated with multiple immune components and signatures in tumor microenvironment. In addition, we identified a PDAC subgroup featured with TMBlowMicrosatellite instabilityhigh (MSIhigh) was associated with prolonged OS and a key molecule, ANKRD55, potentially mediating the survival benefits.ConclusionThis study analyzed the biological function, prognosis value, implications for mutation landscape and potential influence on immune microenvironment of TRGs in PDAC, which contributed to get aware of the role of TMB in PDAC. Future studies are expected to investigate how these TRGs regulate the initiation, development or repression of PDAC.

Deciphering the Prognostic Implications of the Components and Signatures in the Immune Microenvironment of Pancreatic Ductal Adenocarcinoma

Background: The treatment modalities for pancreatic ductal adenocarcinoma (PDAC) are limited and unsatisfactory. Although many novel drugs targeting the tumor microenvironment, such as immune checkpoint inhibitors, have shown promising efficacy for some tumors, few of them significantly prolong the survival of patients with PDAC due to insufficient knowledge on the tumor microenvironment.Methods: A single-cell RNA sequencing (scRNA-seq) dataset and seven PDAC cohorts with complete clinical and bulk sequencing data were collected for bioinformatics analysis. The relative proportions of each cell type were estimated using the gene set variation analysis (GSVA) algorithm based on the signatures identified by scRNA-seq or previous literature.Results: A meta-analysis of 883 PDAC patients showed that neutrophils are associated with worse overall survival (OS) for PDAC, while CD8+ T cells, CD4+ T cells, and B cells are related to prolonged OS for PDAC, with marginal statistical significance. Seventeen cell categories were identified by clustering analysis based on single-cell sequencing. Among them, CD8+ T cells and NKT cells were universally exhausted by expressing exhaustion-associated molecular markers. Interestingly, signatures of CD8+ T cells and NKT cells predicted prolonged OS for PDAC only in the presence of “targets” for pyroptosis and ferroptosis induction. Moreover, a specific state of T cells with overexpression of ribosome-related proteins was associated with a good prognosis. In addition, the hematopoietic stem cell (HSC)-like signature predicted prolonged OS in PDAC. Weighted gene co-expression network analysis identified 5 hub genes whose downregulation may mediate the observed survival benefits of the HSC-like signature. Moreover, trajectory analysis revealed that myeloid cells evolutionarily consisted of 7 states, and antigen-presenting molecules and complement-associated genes were lost along the pseudotime flow. Consensus clustering based on the differentially expressed genes between two states harboring the longest pseudotime span identified two PDAC groups with prognostic differences, and more infiltrated immune cells and activated immune signatures may account for the survival benefits.Conclusion: This study systematically investigated the prognostic implications of the components of the PDAC tumor microenvironment by integrating single-cell sequencing and bulk sequencing, and future studies are expected to develop novel targeted agents for PDAC treatment.