Pancreatic Adenocarcinoma Patients Research Articles

In-Depth Endogenous Phosphopeptidomics of Serum with Zirconium(IV)-Grafted Mesoporous Silica Enrichment.

Detection of endogenous peptides, especially those with modifications (such as phosphorylation) in biofluids, can serve as an indicator of intracellular pathophysiology. Although great progress has been made in phosphoproteomics in recent years, endogenous phosphopeptidomics has largely lagged behind. One main hurdle in endogenous phosphopeptidomics analysis is the coexistence of proteins and highly abundant nonmodified peptides in complex matrices. In this study, we developed an approach using zirconium(IV)-grafted mesoporous beads to enrich phosphopeptides, followed by analysis with a high resolution nanoRPLC-MS/MS system. The bifunctional material was first tested with digests of standard phosphoproteins and HeLa cell lysates, with excellent enrichment performance achieved. Given the size exclusion nature, the beads were directly applied for endogenous phosphopeptidomic analysis of serum samples from pancreatic ductal adenocarcinoma (PDAC) patients and controls. In total, 329 endogenous phosphopeptides (containing 113 high confidence sites) were identified across samples, by far the largest endogenous phosphopeptide data set cataloged to date. In addition, the method was readily applied for phosphoproteomics of the same set of samples, with 172 phosphopeptides identified and significant changes in dozens of phosphopeptides observed. Given the simplicity and robustness of the proposed method, we envision that it can be readily used for comprehensive phosphorylation studies of serum and other biofluid samples.

Identification of Chronic Pancreatitis Associated microRNAs and Genes for the Diagnosis of Pancreatic Cancer.

The timely identification of both malignant and nonmalignant pancreatic lesions has the potential to significantly enhance prognosis and implement risk management strategies across various levels. microRNAs (miRs) and their corresponding targets play a crucial role in the development of pancreatic lesions and can serve as valuable diagnostic and therapeutic targets. The objective of our study was to investigate potential diagnostic markers that can effectively differentiate between malignant and nonmalignant pancreatic lesions. Gene Expression Omnibus (GEO) database with GSE24279 dataset was utilized to screen differentially expressed miRNAs (DEMs). We utilized the TargetScanHuman database to predict the target genes associated with hsa-miR-150-3p, hsa-miR-150-5p, and hsa-miR-214-3p. Furthermore, a cohort comprising healthy individuals (n = 52), chronic pancreatitis (CP; n = 34), and pancreatic adenocarcinoma (PAAD; n = 53) patients was recruited to ascertain the levels of plasma markers. We identified 3 miRNAs (hsa-miR-150-3p, hsa-miR-150-5p, and hsa-miR-214-3p) and 2 proteins (PCDH1 and AMN) as potential diagnostic markers for distinguishing between CP and PAAD. The area under the curve (AUC) values for all markers exceeded .800. Notably, a combination of plasma PCDH1 and AMN demonstrated excellent diagnostic performance (AUC = .921; 95% CI: .866-.977; sensitivity = .792; specificity = .941) in discriminating between CP and PAAD. In addition, the model of hsa-miR-150-3p, hsa-miR-150-5p, and hsa-miR-214-3p yielded an AUC of .928, sensitivity of .830, and specificity of .912, respectively. Plasma levels of miRNAs (hsa-miR-150-3p, hsa-miR-150-5p, and hsa-miR-214-3p) and their corresponding targets (PCDH1 and AMN) hold promise as potential biomarkers for predicting PAAD in patients with CP.

Clinical impact of high-quality testing for peritoneal lavage cytology in pancreatic cancer

In pancreatic ductal adenocarcinoma (PDAC) patients, the importance of peritoneal lavage cytology, which indicates unresectability, remains controversial. This study sought to determine whether positive peritoneal lavage cytology (CY+) precludes pancreatectomy. Furthermore, we propose a novel liquid biopsy using peritoneal lavage fluid to detect viable peritoneal tumor cells (v-PTCs) with TelomeScan F35, a telomerase-specific replication-selective adenovirus engineered to express green fluorescent protein. Resectable cytologically or histologically proven PDAC patients (n = 53) were enrolled. CY was conducted immediately following laparotomy. The resulting fluid was examined by conventional cytology (conv-CY; Papanicolaou staining and MOC-31 immunostaining) and by the novel technique (Telo-CY; using TelomeScan F35). Of them, 5 and 12 were conv-CY+ and Telo-CY+, respectively. All underwent pancreatectomy. The two double-CY+ (conv-CY+ and Telo-CY+) patients showed early peritoneal recurrence (P-rec) postoperatively, despite adjuvant chemotherapy. None of the three conv-CY+ Telo-CY− patients exhibited P-rec. Six of the 10 Telo-CY+ conv-CY− patients (60%) relapsed with P-rec. Of the remaining 38 double-CY− [conv-CY−, Telo-CY−, conv-CY± (Class III)] patients, 3 (8.3%) exhibited P-rec. Although conv-CY+ status predicted poor prognosis and a higher risk of P-rec, Telo-CY was more sensitive for detecting v-PTC. Staging laparoscopy and performing conv-CY and Telo-CY are needed to confirm the indication for pancreatectomy.

Clinicopathological Characteristics, Prognosis, and Correlated Tumor Cell Function of Tropomodulin-3 in Pancreatic Adenocarcinoma.

Pancreatic adenocarcinoma (PAAD) is a frequent malignant tumor with a high mortality rate. Searching for novel biomarkers that can influence its prognosis may help patients. It has been shown that tropomodulin-3 (TMOD3) may influence tumor progression, but its role in pancreatic cancer is not clear. We aimed to explore the expression and prognostic value of TMOD3 in PAAD. We used bioinformatics analysis to analyze the relationship between TMOD3 expression and clinicopathological features and prognosis and verified it with clinical data from tissue microarray. We also conducted in vitro cell experiments to explore the effects of TMOD3 on the function of PAAD cells. TMOD3 expression was found to be significantly higher in PAAD tissues than in matched paracancerous tissues (P < 0.05). Meanwhile, high TMOD3 expression was associated with significantly poorer overall survival (P < 0.05). Analysis of relevant clinicopathological characteristics data obtained from TCGA showed that high TMOD3 expression correlated with age, TNM stage, N stage, and M stage (P < 0.05). Analysis of correlation data obtained from tissue microarrays showed that high TMOD3 expression was associated with lymph node invasion, nerve invasion, macrovascular invasion, and TNM stage (P < 0.05). In addition, siRNA knockdown of TMOD3 significantly reduced the migration and invasion of PAAD cells. Our study shows that TMOD3 may be associated with the progression of PAAD cells, and that it is an independent risk factor for poor pathological features and prognosis of PAAD. It may be helpful as a prognostic indicator of clinical outcomes in PAAD patients.

The survival effect of neoadjuvant therapy and neoadjuvant plus adjuvant therapy on pancreatic ductal adenocarcinoma patients with different TNM stages: a propensity score matching analysis based on the SEER database

ABSTRACT Background Adjuvant therapy (AT) and neoadjuvant therapy (NAT) are standard treatments for pancreatic ductal adenocarcinoma (PDAC) depending on the status of the disease. However, whether AT improves survival after NAT and radical resection in all TNM stages remains unclear. Research design and methods We utilized the Surveillance, Epidemiology, and End Results (SEER) database (2010–2019) for PDAC patients who underwent radical surgery and applied Pearson’s chi-square test, multivariate and univariate Cox regression, Kaplan-Meier plot, Log-rank tests, and propensity score matching (PSM) for analysis. Results Given PSM after enrolling 13,868 PDAC patients, significant differences in survival were identified between AT and neoadjuvant therapy plus adjuvant therapy (NATAT) (p = 0.023) as well as between NAT and NATAT (p < 0.001). According to the AJCC 8th TNM stage, a survival advantage associated with NATAT was exclusively observed in stage III and IV disease, except for T4N0M0. Some stage IV patients receiving NATAT exhibited comparable survival to their counterparts without metastasis. Conclusions In this retrospective cohort study, we demonstrated that patients harboring tumors in late TNM stages, including N2 resectable PDAC, might have better survival from NATAT, and that certain patients with M1 disease might still benefit from comprehensive systemic therapy and radical resection.

Early onset pancreatic cancer-exploring contemporary treatment and outcomes using real-world data.

Pancreatic cancer incidence is increasing in younger populations. Differences between early onset pancreatic cancer (EOPC) and later onset pancreatic cancer (LOPC), and how these should inform management warrant exploration in the contemporary setting. A prospectively collected multi-site dataset on consecutive pancreatic adenocarcinoma patients was interrogated. Patient, tumour, treatment, and outcome data were extracted for EOPC (≤50 years old) vs LOPC (>50 years old). Of 1683 patients diagnosed between 2016 and 2022, 112 (6.7%) were EOPC. EOPC more frequently had the tail of pancreas tumours, earlier stage disease, surgical resection, and trended towards increased receipt of chemotherapy in the curative setting compared to LOPC. EOPC more frequently received 1st line chemotherapy, 2nd line chemotherapy, and chemoradiotherapy than LOPC in the palliative setting. Recurrence-free survival was improved for the tail of pancreas EOPC vs LOPC in the resected setting; overall survival was superior for EOPC compared to LOPC across the resected, locally advanced unresectable and metastatic settings. EOPC remains a small proportion of pancreatic cancer diagnoses. The more favourable outcomes in EOPC suggest these younger patients are overall deriving benefits from increased treatment in the curative setting and increased therapy in the palliative setting.

Clinical Utility of the Combined Use of CA19-9 and DUPAN-2 in Pancreatic Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) patients with normal carbohydrate antigen (CA) 19-9 levels can have early-stage cancer or advanced cancer without elevation of CA19-9 level; estimating their malignant potential is difficult. This study investigated the clinical utility of the combined use of preoperative CA 19-9 and Duke pancreatic monoclonal antigen type 2 (DUPAN-2) levels in patients with PDAC. Patients who underwent curative-intent surgery for PDAC between November 2005 and December 2021 were investigated. Eligible patients were classified into four groups based on these two markers. Among patients with normal CA19-9 levels, those with normal and high DUPAN-2 levels were classified into normal/normal (N/N) and normal/high (N/H) groups, respectively. Among patients with high CA19-9 levels, those with normal and high DUPAN-2 levels were classified into high/normal (H/N) and high/high (H/H) groups, respectively. Survival rates were compared between the groups. Among 521 patients, the N/N, N/H, H/N, and H/H groups accounted for 25.0%, 10.6%, 35.1%, and 29.4% of patients, respectively. The proportions of resectable PDAC in the N/N and H/N groups (71.5% and 66.7%) were significantly higher than those in the N/H and H/H groups (49.1% and 54.9%) (P < 0.01). The 5-year survival rates in the N/N, N/H, H/N, and H/H groups were 66.0%, 31.1%, 34.9%, and 29.7%, respectively; the rate in the N/N group was significantly better than those in the other three groups (P < 0.0001, P < 0.0001, and P < 0.0001, respectively). Only patients with normal CA19-9 and DUPNA-2 values should be diagnosed with early-stage PDAC.

The inverted U-shaped relationship between epinephrine and pancreatic ductal adenocarcinoma patients' survival with compensation of lymphocyte.

The relationship between epinephrine and cancer can be dose-dependent in invivo study. Whether it is the same in human body still needs verification. We used frozen human pancreatic ductal adenocarcinoma (PDAC) tissues to detect epinephrine content and analyzed its relationship with survival using the K-M method and Cox regression. Disturbance of blood cell count and C-reactive protein and identification of related potent intermediary factors were also analyzed. K-M plot and Cox regression all showed the inverted U-shaped relationship between epinephrine and PDAC survival. Lymphocyte adjustment can increase the HRs of epinephrine for PDAC death by >10%. Epinephrine played an anti-tumor or pro-tumor effect depending on the specific concentration. Circulating lymphocyte count was elevated and might acted as a compensation pathway to reduce the pro-tumor effect of epinephrine to PDAC.

Highly sensitive serum volatolomic biomarkers for pancreatic cancer diagnosis

The discovery of new diagnostic tools for the early detection of diseases with poor prognosis such as pancreatic adenocarcinoma (PAC) is of high importance. The results from a control-case study (20 PAC patients, 19 healthy controls) for the search of new biomarkers of pancreatic cancer based in differences in the serum volatolome are presented in this work. Volatolomics were performed following a non-targeted HS-SPME-GC/MS approach, and a total of 433 volatile organic compounds (VOCs) was detected in the human serum samples. Of these, 125 VOC indexes showed a significant variation when controls and patients were compared (p-value < 0.05). Bonferroni corrected p-values < 0.05 were found for 40 features. PCA analysis showed the control-PAC discrimination capability of VOCs in serum, and PLS-DA was performed to select the best candidate biomarkers for the diagnosis of PAC. For the 40 selected VOCs, calculated areas under the curve (AUC) ranged from 0.98 to 0.85, and 11 of them were successfully validated using an independent set of samples (5 PAC patients, 5 healthy controls). Four of the proposed PAC biomarkers were identified as toluene, 2-ethyl-1-hexanol, pentylbenzene, and butoxymethylbenzene. Combinations of the identified PAC biomarkers were tested and showed AUC > 0.90, with the more promising candidate being butoxymethylbenzene (AUC = 0.98).

Development of Clinical Radiomics-Based Models to Predict Survival Outcome in Pancreatic Ductal Adenocarcinoma: A Multicenter Retrospective Study.

This multicenter retrospective study aims to identify reliable clinical and radiomic features to build machine learning models that predict progression-free survival (PFS) and overall survival (OS) in pancreatic ductal adenocarcinoma (PDAC) patients. Between 2010 and 2020 pre-treatment contrast-enhanced CT scans of 287 pathology-confirmed PDAC patients from two sites of the Hopital Universitaire de Bruxelles (HUB) and from 47 hospitals within the HUB network were retrospectively analysed. Demographic, clinical, and survival data were also collected. Gross tumour volume (GTV) and non-tumoral pancreas (RPV) were semi-manually segmented and radiomics features were extracted. Patients from two HUB sites comprised the training dataset, while those from the remaining 47 hospitals of the HUB network constituted the testing dataset. A three-step method was used for feature selection. Based on the GradientBoostingSurvivalAnalysis classifier, different machine learning models were trained and tested to predict OS and PFS. Model performances were assessed using the C-index and Kaplan-Meier curves. SHAP analysis was applied to allow for post hoc interpretability. A total of 107 radiomics features were extracted from each of the GTV and RPV. Fourteen subgroups of features were selected: clinical, GTV, RPV, clinical & GTV, clinical & GTV & RPV, GTV-volume and RPV-volume both for OS and PFS. Subsequently, 14 Gradient Boosting Survival Analysis models were trained and tested. In the testing dataset, the clinical & GTV model demonstrated the highest performance for OS (C-index: 0.72) among all other models, while for PFS, the clinical model exhibited a superior performance (C-index: 0.70). An integrated approach, combining clinical and radiomics features, excels in predicting OS, whereas clinical features demonstrate strong performance in PFS prediction.

Identification of Novel Tumor Pyroptosis-Related Antigens and Pyroptosis Subtypes for Developing mRNA Vaccines in Pancreatic Adenocarcinoma.

As one of the important components of immunotherapies, mRNA vaccines have displayed promising clinical outcomes in solid tumors. Nonetheless, their efficacy remains unclear in pancreatic adenocarcinoma (PAAD). Given the interaction of pyroptosis with anticancer immunity, our study aims to identify pyroptosis-related antigens for mRNA vaccine development and discern eligible candidates for vaccination. Utilizing gene expression data from TCGA and ICGC, we integrated RNA-seq data and compared genetic alterations through cBioPortal. Differential gene expressions were integrated using GEPIA. Relationships between immune cell abundance and tumor antigens were analyzed and visualized via TIMER. WGCNA facilitated the clustering of pyroptosis-related genes, identification of hub genes, and pathway enrichment analyses. Pyroptosis landscape was depicted through graph learning-based dimensional reduction. Four overexpressed and mutant pyroptosis-related genes associated with poor prognosis were identified as potential antigens for mRNA vaccines in PAAD, including ANO6, PAK2, CHMP2B, and RAB5A. These genes displayed positive associations with antigen-presenting cells. PAAD patients were stratified into three pyroptosis subtypes. Notably, the PS3 subtype, characterized by a lower mutation count and TMB, exhibited "cold" immunological traits and superior survival compared to other subtypes. The pyroptosis landscape exhibited considerable heterogeneity among individuals. Furthermore, the turquoise module emerged as an independent prognostic indicator and patients with high expressions of hub genes might not be suitable candidates for mRNA vaccination. In PAAD, ANO6, PAK2, CHMP2B, and RAB5A are prospective pyroptosis-related antigens for mRNA vaccine development, which holds potential benefits for patients classified as PS3 and those with diminished hub gene expressions, providing insights into personalized mRNA vaccine strategies.

Abstract 1585: Tenascin C in pancreatic cancer-associated fibroblasts enhances epithelial mesenchymal transition and is associated with resistance to immune checkpoint inhibitor

Abstract Backgrounds: Tenascin C (TNC) is an extracellular matrix glycoprotein that is abundantly expressed in cancer stroma and its overexpression correlates with tumor progression in various types of cancer including pancreatic adenocarcinoma (PAAD). However, the role of TNC in regards to response to immune checkpoint inhibitor (ICI) still remains to be identified. In this study, we aimed to investigate the potential involvement of TNC in the response to ICI among PAAD patients. Materials and Methods: Transcriptomic profiles were downloaded by TCGA and GTEx database and TNC mRNA levels were compared between tumors and normal tissues. These transcriptomic data were deconvoluted using CIBERSORTx to identify specific enriched pathways in cancer cells and cancer-associated fibroblasts (CAFs) in PAAD patients. Bioinformatic programs were used to predict paracrine communications between cancer cells and cancer-associated fibroblasts (CAFs), and the Tumor Immune Dysfunction and Exclusion (TIDE) score was calculated to predict response to ICI treatment in PAAD patients. An independent immunotherapeutic cohort was applied to validate prediction values for immunotherapy. Results: TNC mRNA levels were significantly upregulated in tumors compared to normal tissues in eight cancer types including PAAD. TNC-high (TNC-H) PAAD patients (n = 138) had significant shorter overall survival than TNC-low (TNC-L) patients (n = 40) (p = 0.0125, Log-rank test). Using CIBERSORTx, TNC was estimated to be predominantly expressed in CAFs in PAAD. In pathway analysis, both cancer cells and CAFs had significant enrichments of the epithelial mesenchymal transition pathway in TNC-H patients. Ligand-receptor interaction analysis showed that TNC produced in CAFs had the highest interaction potentials to bind to integrin families such as ITGαV and ITGβ3 in cancer cells. In an independent immunotherapy data cohort, patients with TNC-H and ITGαV-high or ITGβ3-high expression were associated with poor response to ICI treatment and had shorter overall survival than those with both low expressions. Conclusions: In conclusion, these findings suggest that TNC-high CAFs can play a crucial role in tumor progression and resistance to ICI therapy in PAAD patients, and targeting TNC and its interactions with cancer cells may provide a potential strategy for improving the efficacy of ICI therapy in PAAD. Citation Format: Satoru Furuhashi, Yoshifumi Morita, Akio Matsumoto, Shinya Ida, Ryuta Muraki, Ryo Kitajima, Makoto Takeda, Hirotoshi Kikuchi, Yoshihiro Hiramatsu, Hiroya Takeuchi. Tenascin C in pancreatic cancer-associated fibroblasts enhances epithelial mesenchymal transition and is associated with resistance to immune checkpoint inhibitor [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 1585.

Abstract 6402: Treatment resistance and survival prediction using a machine learning gene expression signature in pancreatic adenocarcinoma

Abstract Background: Pancreatic adenocarcinoma (PAAD) is highly aggressive and difficult to treat. The 5-year survival rate is only 12.5%. The Cancer Genome Atlas (TCGA) data has shown about 51% of PAAD patients didn’t achieve complete response after first course of treatment (CRAFCT); this study is to use TCGA data to develop survival-treatment associated gene signature to screen the patients, and provide novel therapeutic strategies to customize patient treatment. Methods: RNAseq data of 177 PAAD patients and their clinical data were downloaded from TCGA data portal and processed. The dataset was randomly split into training and testing data. RNAseq data analysis and survival analyses were performed on the training data to identify the survival-treatment associated genes, and a survival-treatment gene signature was derived and independently tested on the test data using signature score by cutoff, logistic regression, and random forest. Furthermore, the signature was validated by an independent dataset of 96 cases. Results: Among the 177 patients, 137 patients had the information of CRAFCT, and 122 patients had specific treatment information. Among those 122 patients, 99% received chemotherapy, 36% received radiation therapy. About half of the PAAD patients (51%) didn’t achieve CRAFCT. RNAseq data analysis identified 304 significantly expressed genes associated with the status of complete response Yes vs No, and survival analyses further selected 55 genes associated with both treatment and survival as a signature. This signature, using z-scaled scores and the cut-off of 0.3, was applied to the 99 patients of training data; the accuracy is 0.66, and the patients predicted to be Yes had a significantly better (p=0.005, HR=0.42) overall survival than those predicted No. To further test the signature, we applied it to the test data of 78 patients; the random forest model showed patients screened to be Yes had a significantly better (p=0.016, HR=0.43) overall survival than those screened No. The signature (using 40 of the 55 genes) was applied to an independent dataset of 96 cases to significantly screen the patients to be Yes vs No (resistance to treatment) with p=0.04 and HR=0.57, showing the 55-gene signature is valid and robust. Conclusion: A 55 survival-treatment associated gene signature was developed and validated robustly, and can potentially be applied as a novel strategy to screen patients for treatment response/resistance. These 55 genes could also be targeted to develop new drugs for those difficult to treat PAAD patients. Disclaimer: The contents of this publication are the sole responsibility of the author(s) and do not necessarily reflect the views, opinions or policies of USUHS, HJF, the DoD or the Departments of the Army, Navy or Air Force. Mention of trade names, commercial products, or organizations does not imply endorsement by the U.S. Government. Citation Format: Jianfang Liu, Xiaoying Lin, Craig D. Shriver, Hai Hu. Treatment resistance and survival prediction using a machine learning gene expression signature in pancreatic adenocarcinoma [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 6402.

Abstract 2990: Differential micro-RNA expression in circulating exosomes of pancreatic ductal adenocarcinoma patients

Abstract Pancreatic ductal Adenocarcinoma (PDAC) is a highly lethal solid tumor with 5-year survival rate of 5-8%. Extracellular vesicles like exosomes circulating in the blood are crucial mediators of intracellular communication, immunomodulation and tumor progression. The microRNAs being the major cargo of exosomes, are the primary carriers of intracellular information. The role of exosomal-miRNA (exoMIR’s) in regulating the pathogenesis of tumors is well-established in oncology but its potential is still unexplored. To the best of our knowledge no study has yet explored the contrasting exoMIR expression in cancer patients before and after their surgery. In this study, differential exoMIR’s profile of three pancreatic ductal adenocarcinoma patients at three stages: pre-surgery, post-surgery, and follow-up was established using small RNA sequencing on the Illumina platform. In blood samples taken before and after surgery total of 181, 201 and 201 microRNAs were dysregulated in circulating exosomes of three different patients viz. P_1, P_9 &amp; P_11 (Patient identifiers) respectively. Out of the total dysregulated microRNAs in post-op samples 76 were upregulated and 105 downregulated in P_1, 95 upregulated and 106 downregulated in P_9 &amp; 69 upregulated and 132 downregulated in P_11 as compared to their Pre surgery samples. Further, when exoMIR’s of post-op samples were compared to that of follow-up samples after 6 months of surgery their expression profile changed notably such that a total of 184, 157 &amp; 190 microRNAs dysregulated in follow-up of P_1, P_9 and P_11 respectively. Out of the total dysregulated microRNAs in follow up samples 93 were upregulated and 91 downregulated in P_1, 69 upregulated and 88 downregulated in P_9 &amp; 96 upregulated and 94 downregulated in P_11 as compared to their post-op samples. On Venn diagram analysis 27 microRNAs were found to be commonly dysregulated in all 3 patients before surgery and 22 common micro RNAs were again dysregulated even after 6 months of surgery. Bioinformatic analysis of these differentially expressed miRNAs and their respective protein targets revealed that their role in oncogenic molecular pathways is very prominent, thus highlighting their potential use in cancer therapeutics, diagnostics as well as prognosis. Citation Format: Rupinder Kaur, Sandeep Bhoriwal, Promod Kumar Gautam. Differential micro-RNA expression in circulating exosomes of pancreatic ductal adenocarcinoma patients [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 2990.

Abstract 7643: Prognostic significance and immune infiltration patterns related to Claudin heterogeneity in pancreatic ductal adenocarcinoma patients

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) stands out for its aggressive nature, often manifesting through invasion, metastasis, and recurrence. These characteristics are partly attributable to the Epithelial-mesenchymal transition (EMT), a critical phenomenon that follows the disintegration of epithelial barriers, including tight junctions (TJs), where Claudin (CLDN) proteins are integral components. Objective: This study focused on elucidating the differential expression of claudins in PDAC and exploring their correlations with disease progression, invasiveness, and immune infiltration. Methodology: Utilizing UCSC TOIL RNA-seq, TCGA-PAAD genomic, and clinical data, we investigated the genomic alterations and differential expression of claudins in PDAC. Conducted survival analysis using log-rank tests. Assessed immune infiltration patterns using the EPIC deconvolution algorithm. Results: Of 177 PDAC samples examined, 23 (13%) exhibited at least one form of claudin gene alteration, including mutations, amplifications, or deep deletions. Remarkably, 10 claudins (CLDN1, 2, 4, 5, 7, 11, 12, 15, 18, and 23) were found to be significantly overexpressed in PDAC samples, meeting a log2FC threshold of ≥1 (p &amp;lt; 0.05). Further, we identified a direct correlation between mRNA expression and copy numbers, alongside an inverse relationship with methylation levels, particularly in CLDN1, CLDN12, CLDN15, CLDN23, and CLDN4. Intriguingly, claudin expression escalated from grades G1 to G3 but diminished in G4, with CLDN1, CLDN4, and CLDN7 showing significant variability (Kruskal-Wallis rank test p-values of 1.2E-5, 0.00064, and 0.023, respectively). Survival analysis indicated that high expression levels of CLDN1 and CLDN4 were linked to poorer patient outcomes, while higher levels of CLDN5 and CLDN15 correlated with improved survival (log-rank p-values of 0.007, 0.016, 0.0035, and 0.0012, respectively), findings consistent across both univariate and multivariate analyses. Furthermore, immune infiltration patterns, assessed via the EPIC deconvolution algorithm, varied significantly between high and low CLDN expression groups. Specifically, low CLDN1 expression was associated with increased presence of B-cells, CAF, and CD8+ T-cells but fewer endothelial cells, a pattern reversed in high CLDN1 expression cases. A similar trend was observed for CLDN4. Conclusion: Our study underscores the significant prognostic value of claudin proteins in PDAC, highlighting their potential as indicators of disease progression and survival and their influence on the tumor's immune environment. Citation Format: Ajaz A. Bhat, Shahabuddin Usmani, Hana Q. Sadida, Sheema Hashem, Tariq Masoodi, Ikhlak Ahmed, Rakesh Kumar, Mayank Singh, Punita Dhawan, Shahab Uddin, Muzafar A. Macha, Ammira S. Al-Shabeeb Akil. Prognostic significance and immune infiltration patterns related to Claudin heterogeneity in pancreatic ductal adenocarcinoma patients [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 7643.

Abstract 2542: Real-world validation of the PurIST classifier demonstrates enhanced therapy selection for pancreatic ductal adenocarcinoma (PDAC) patients

Abstract Background: PDAC is a particularly fatal condition lacking established biomarkers for the choice of first-line treatment (1L). The PurIST algorithm is a gene signature (Rashid et al. Clin. Cancer Res., 2020) that categorizes PDAC tumors into basal or classical molecular subtypes and has been previously validated in a patient cohort of advanced PDAC (N=258). In this post-hoc analysis study, we examine whether the PurIST subtypes can distinguish patients likely to respond to FOLFIRINOX (FFX) versus Gemcitabine + nab-paclitaxel (GnP). Methods: A cohort of de-identified PDAC patients was selected from the Tempus Oncology Database following an IRB-approved protocol. Inclusion criteria required no prior systemic treatment, a surgically unresectable/metastatic status, available RNA-sequencing data from primary or metastatic tissue, and the use of FFX or GnP as 1L systemic therapy with documented outcomes. RNA-sequencing was generated by the CAP/CLIA validated Tempus xR assay. The PurIST gene signature was applied to normalized gene expression values to classify samples into basal or classical subtypes. Univariate and multivariate Cox PH analyzes comparing overall survival (OS) with adjustment for delayed entry, between two common 1L treatment regimens, FFX and GnP, were performed for classical patients with an ECOG performance status of 0 or 1. Results: Application of the PurIST model to the PDAC cohort (N=258) resulted in 67% (N=173) labeled as classical, 23% (N=59) as basal, and the remainder as no-call. Amongst classical patients, 35% (N=61) had a known ECOG score of 0 or 1 and were retained for subsequent analysis. Within this subset, 37 were treated with FFX and 24 with GnP as 1L, with an overall median OS of 11.4 months (95% CI: 9.5-16.5) and a 12-month survival rate of 47.2%. A univariate Cox PH analysis showed that classical patients treated with FFX had a significantly longer OS time (HR (95% CI) = 2.32 (1.29-4.17)). This result remained significant in a multivariate Cox PH, adjusting for age and ECOG, (HR=2.5 (1.15-5.44)). Conclusion: In this real-world cohort of advanced PDAC patients, we showed that classical patients with low ECOG scores had superior OS with 1L FFX treatment compared to 1L GnP. These findings demonstrate the potential for PurIST to aid in optimal treatment selection for patients with advanced PDAC. Citation Format: Stephane Wenric, James M. Davison, John Guittar, Gregory M. Mayhew, Kirk Beebe, Alia Zander, Seung Won Hyun, Kyle Beauchamp, Michael V. Milburn, Vincent Chung, Tanios S. Bekaii-Saab, Charles M. Perou. Real-world validation of the PurIST classifier demonstrates enhanced therapy selection for pancreatic ductal adenocarcinoma (PDAC) patients [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 2542.

Abstract 4588: Combination of MTA-cooperative PRMT5 inhibitor and direct mutant-selective KRAS inhibitors as a novel therapeutic approach for MTAP-deficient pancreatic cancer

Abstract KRAS is mutationally activated in 95% of pancreatic ductal adenocarcinoma (PDAC) patients. Direct KRAS inhibitors are under intense preclinical and clinical development, with two KRASG12C mutant-selective inhibitors (G12Ci) now approved. However, treatment-associated resistance to KRAS inhibitors has been reported in the clinic highlighting an urgent need to identify both additional therapeutic targets and novel combination treatment strategies with KRAS inhibitors. To that end, we performed a CRISPR/Cas9 loss-of-function screen using a library targeting ~2,500 druggable genes in KRASG12C/D-mutant PDAC cell lines. We identified multiple genes that were essential for PDAC cell growth and selected PRMT5, a protein arginine methyltransferase, to assess as a novel therapeutic target for pancreatic cancer. We found that suppression of PRMT5 activity using two distinct first-generation clinical candidate small molecule inhibitors (JNJ-64619178 and GSK3326595) demonstrated single agent activity and reduced PDAC cell growth. We further assessed a mechanistically distinct MTA-cooperative clinical candidate PRMT5 inhibitor, MRTX1719, that is selective for MTAP-deleted tumors, which occur in ~25% of PDAC patients. We determined that MRTX1719 exhibited low nanomolar GI50 activities in MTAP-deficient but not MTAP wild-type KRAS-mutant cell lines. Unexpectedly, PDAC cells treated with MRTX1719 exhibited a paradoxical activation of ERK, suggesting that combined inhibition of PRMT5 and KRAS might be therapeutically advantageous. In support of our hypothesis, we demonstrated that combination treatment with MRTX1719 and mutant selective KRAS inhibitors (G12Ci/MRTX849 and G12Di/MRTX1133) further sensitized KRASG12C/D-mutant PDAC cells to KRAS inhibition in short- and long-term growth assays as well as in vivo xenograft studies. Our ongoing studies are evaluating the consequences of co-targeting PRMT5 and KRAS on gene expression changes using RNA-Seq and cancer cell signaling pathways using RPPA analyses. In summary, our data support concurrent inhibition of PRMT5 and KRAS as a promising therapeutic strategy for MTAP-deficient KRAS-mutant pancreatic cancer. Citation Format: Kristina Drizyte-Miller, Wen-Hsuan Chang, Andrew M. Waters, Clint A. Stalnecker, Junning Wang, Andrew J. Aguirre, Adrienne D. Cox, Channing J. Der. Combination of MTA-cooperative PRMT5 inhibitor and direct mutant-selective KRAS inhibitors as a novel therapeutic approach for MTAP-deficient pancreatic cancer [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 4588.

Abstract 382: Pancreatic cancer-derived organoids alter muscle fiber type and increased energy consumption leading to cachexia

Abstract Cachexia, marked by continuous skeletal muscle mass loss and severe metabolic disturbances, presents a substantial challenge for pancreatic ductal adenocarcinoma (PDAC) patients. Despite its prevalence and the adverse impact on quality of life and survival rates, effective clinical treatments for cachexia remain elusive. This study aims to bridge this gap by employing innovative in vitro models to investigate the underlying mechanisms of cachexia and identify potential therapeutic targets. To enhance the relevance of cell culture models, modifications were introduced to replicate in vivo conditions, where the muscle is exposed to the ongoing kinetics of constant tumor secretion of active factors. Patient-derived PDAC cells were cultured in a dense 3D stiffness extracellular matrix (ECM) to mimic the hypoxic desmoplastic tumor microenvironment. The conditioned medium from prolonged cultured patient-derived organoids (PDOs) was then used to supplement C2C12-derived myotubes. The investigation focuses on dynamic adaptations in muscle fiber type and metabolism when exposed to tumor paracrine factors. Metabolomic investigations of PDO-conditioned medium (PDO CM) on myotubes revealed distinct metabolic differences compared to non-treatment controls. Discriminatory metabolites included those associated with glycolysis, lipid, amino acid, and fatty acid metabolism. PDO CM induced a significant shift in muscle fiber type from slow-twitch to fast-twitch phenotypes, accompanied by de novel expression of embryonic and neonatal Myosin Heavy Chain (MyHC) isoforms. Fast fiber (F59) staining also confirmed the protein expression. Concurrently, PGC-1a, a master regulator of mitochondrial biogenesis and oxidative metabolism, significantly decreased, along with reduced ATP and Mito tracker staining, indicating a metabolic switch under the influence of tumor paracrine factors. Metabolome results on C2C12 myotubes treated with PDO CM also revealed significantly changed ABC transporter-related pathways, an adaptive process required to optimize substrate use under the influence of tumor secretome. However, the genes involved in the proteasome degradation process, such as MAFbx and MuRF1, did not show altered expression. The establishment of an in vitro cachexia model, focusing on changes in energy metabolism and utilizing muscle fiber type alterations as markers, will deepen our understanding of the mechanisms of cancer-associated cachexia. The insights gained may pave the way for identifying potential therapeutic targets to mitigate muscle wasting in PDAC patients and improve overall outcomes. Citation Format: Bo Han, Shuqing Zhao, zhi Yang, Jose Trevino, Steven Grossman, Heinz-Josef Lenz, Ba Xuan Hoang. Pancreatic cancer-derived organoids alter muscle fiber type and increased energy consumption leading to cachexia [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 382.

Abstract 7465: Leukemia inhibitory factor creates an immune suppressive tumor microenvironment and fosters tumor growth in KRAS-driven pancreatic ductal adenocarcinoma

Abstract Immune checkpoint blockade revolutionized cancer treatments by activating anti-tumor immunity. Pancreatic ductal adenocarcinoma (PDA) patients have not benefited from these advances due to an immune suppressive tumor microenvironment. Identifying PDA cell-encoded factors that drive immune evasion is necessary to unlock new immunologically active treatment options in PDA. We have recently demonstrated that the cytokine Leukemia inhibitory factor (LIF) is highly upregulated in PDA compared to the normal healthy pancreas. LIF is known for its immune suppressive qualities, but data on LIF in PDA is currently limited. We generated a conditional Lif flox (L) allele and crossed it into the well-established KrasLSL-G12D/+, Trp53flox/flox, Pdx1-Cre (KPC) mouse model. LKPC mice (n=29) had longer overall survival (69 days) than control KPC mice (n=29) (59 days), (p&amp;lt;0.05). LKPC mice harbored significantly smaller tumors with an average tumor weight of 0.6g compared to KPC control animals with an average tumor weight of 1.0g (p&amp;lt;0.005). Immunohistochemical evaluation of tumor proliferation measured by Ki-67 positivity in LKPC and KPC tumors at endpoint showed comparable results. Tumor-derived cell lines from KPC and LKPC mice showed no growth differences in vitro nor when reimplanted subcutaneously (sc) in immunodeficient mice. In contrast, we observed a significant decrease in tumor growth and weight in LKPC sc tumors (p&amp;lt;0.05) compared to the KPC sc tumors in syngeneic mice. LKPC sc tumors had significantly fewer neutrophils and more macrophages, particularly MHC II-high macrophages, known for their anti-tumor qualities when compared to KPC sc tumors using flow cytometry. A trend towards more CD8+ T-cells in sc LKPC tumors was also observed. Our genetically engineered mouse model confirmed this finding by immunohistochemical evaluation of tumors at endpoint, which demonstrated a significant increase of CD8+ T-cells in the LKPC animals compared to the KPC controls. In summary, we find that cancer-cell-expressed LIF augments tumor growth and shortens overall survival by changing the immune tumor microenvironment, nominating leukemia inhibitory factor depletion as a promising therapeutic immune target in KRAS-driven pancreatic cancer. Citation Format: Lisa Miller-Phillips, Imene Boukhalfa Hanafi, Alexis J. Combes, Shabana Bano, Lisiena Hysenaj, Jeroen P. Roose, Adam Olshen, Ashley Kiemen, Sarah Umetsu, Man-Tzu Wang, Eric A. Collisson. Leukemia inhibitory factor creates an immune suppressive tumor microenvironment and fosters tumor growth in KRAS-driven pancreatic ductal adenocarcinoma [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 7465.

Abstract 4901: High-resolution spatial transcriptomics identifies candidate genes facilitating perineural invasion

Abstract Perineural invasion (PNI), or the invasion of cancer to the space surrounding nerves, is associated with poor prognosis, but the underlying molecular factors that drive its initiation and progression are poorly understood. To explore this systematically, we applied a 6000-plex spatial transcriptomic panel with subcellular resolution to curated microarrays from 5 pancreatic ductal adenocarcinoma (PDAC) patients spanning different stages of tumor-nerve involvement. With this preliminary cohort, we explored malignant cell signatures associated with invasion and adaptation to the intraneural environment. Strikingly, we observed enrichment of a classical-like malignant transcriptional program marked by genes involved in glucose and lipid metabolism in nerve-invading malignant cells. Despite the known association of chemoresistance with serine dependence, these cells also exhibited upregulation of genes involved in serine biosynthesis, suggesting that intraneural invasion may offer an orthogonal path of treatment evasion. Overall, our findings indicate that nerve-invasive cancer cells adopt a unique transcriptomic signature within the neuronal microenvironment. Further studies will aim to uncover the relevance of these signatures to survival within the nerve environment and treatment resistance. Citation Format: Deniz Guney Olgun, Peter L. Wang, Ryan Zhao, Dennis H. Gong, William L. Hwang. High-resolution spatial transcriptomics identifies candidate genes facilitating perineural invasion [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 4901.