Introduction

Inflammation-based prognostic indicators have been developed to predict the prognosis in patients with pancreatic cancer. Neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR) and C-Reactive Protein/Albumin Ratio (CAR) have shown to be predictors of poor outcomes in various malignancies.

Aims & Methods

The aim of our study was to assess the role of NLR, PLR and CAR in predicting prognosis and sarcopenia in pancreatic cancer. We conducted a retrospective single-center study, including all patients diagnosed with pancreatic adenocarcinoma between 2014 and 2022.
Patient characteristics, inflammatory markers, tumor status and outcomes were collected. Baseline NLR, PLR and CAR were calculated for each patients. Radiologic sarcopenia was assessed by calculating the total psoas area index.

Results

In total, thirty patients were enrolled with a mean age of 61 ± 11 years and a sex ratio M/F of 4. The tumor was predominantly located at the head (66.6%). Regarding staging, the tumor was resectable in 11 patients (36%), locally advanced in 10 patients (33%) and metastatic in 9 patients (9%). Mortality rates at 6 months, 12 months and 36 months were 50%, 63.3% and 75% respectively. All the analyzed ratios were not associated to tumor locations or stages (p>0.05). High NLR was significantly higher in patients with sarcopenia compared to those without sarcopenia (4.7 ± 2.6 vs 2.9 ± 1.62 respectively, p = 0.02), Difference not found with PLR or CAR. When analyzing the receiver operating characteristic (ROC) curve, AUC of NLR, PLR and CAR in predicting 12 months survival were [0.838 (95% CI: 0.683–0.992, p=0.004)], [0.831 (95% CI: 0.671–0.992, p=0.005)] and [0.781 (95% CI: 0.601–0.961, p=0.018)], respectively.

Conclusion

In our study, pre-therapeutic NLR was significantly correlated with sarcopenia and showed better prediction of 12 months survival and thus associated to a poor prognosis. It is a simple, easly to calculate and valuable tool for assessment of prognosis in pancreatic cancer.

Introduction

Despite advances in cancer treatment, pancreatic adenocarcinoma continues to have a poor prognosis and remains a leading cause of death among all tumors [1]. Most pancreatic adenocarcinoma patients are diagnosed in stages with metastasis. Moreover, pancreatic adenocarcinoma has a restricted ability to respond to immunotherapy due to low immune infiltration. Prior studies have demonstrated a correlation between tumor endothelial cells (TECs) and angiogenesis, which has been indicated to play a critical role in T-cell-mediated immunosurveillance [2]. Identifying the specific subpopulation of TEC that correlates with immune evasion in pancreatic adenocarcinoma remains a challenge. However, to clarify this subpopulation may bring novel hope to patients with pancreatic adenocarcinoma.

Aims & Methods

Our study aimed to identify the specific subpopulation of TECs associated with increased immunoevasion. Analysis based on the TCGA database, scRNA-seq, the online tool Kaplan Meier plotter, and spatial transcriptomics data were utilized in our study. Moreover, we cultured human umbilical vein endothelial cells (HUVECs) and performed siRNA transfection and Western blotting to confirm the function of RASIP1 in endothelial cells.

Results

Genes related to patient survival in pancreatic adenocarcinoma were identified by the analysis from the Cancer Genome Atlas (TCGA). On the other hand, based on scRNA-seq data, the genes exclusively expressed in endothelial cells were identified. Our analysis revealed that RASIP1 (Ras Interacting Protein 1) was specifically expressed in endothelial cells and linked to improved pancreatic cancer patient survival. Subsequently, the gene ontology analysis of RASIP1-positive TECs was performed based on the scRNA-seq data, and it turned out that RASIP1-positive TECs were related to leukocyte transendothelial migration. Intriguingly, based on the analysis of the TCGA database and spatial transcriptomics data, it was revealed that RASIP1-positive TECs are correlated with increased T cell infiltration and elevated T-effector and IFN-gamma gene signatures in pancreatic adenocarcinoma. These findings suggest that RASIP1-positive TECs may contribute to the "hot" tumor phenotype by promoting T cell infiltration. To further investigate the relationship between RASIP1 and leukocyte transendothelial migration, we knocked down RASIP1 in human umbilical vein endothelial cells (HUVECs) and found a decrease in the expression of the key adhesion molecule intercellular cell adhesion molecule (ICAM). Moreover, the potential related signaling pathway was also investigated, including Erk and p38. Intriguingly, the survival analysis based on patients with pan-cancer immunotherapy also indicated the expression of RASIP1 was proportional to the survival time of cancer patients receiving immunotherapy. In the patients with high expression of RASIP1, longer survival was shown, with a p-value of 0.014 and HR of 0.74. And particularly, in patients treated with PD-L1, significantly longer survival was shown in the group with high expression of RASIP1, with a p-value of 0.01 and HR of 0.74

Conclusion

All in all, our study revealed a potential novel type of endothelial cell in pancreatic adenocarcinoma, which correlated with increased leukocyte transendothelial migration and may provide a potentially effective therapeutic mechanism.

References

1. Ullman NA, Burchard PR, Dunne RF, Linehan DC. Immunologic Strategies in Pancreatic Cancer: Making Cold Tumors Hot. J Clin Oncol. 2022 Aug 20;40(24):2789-2805.
2. Nagl L, Horvath L, Pircher A, Wolf D. Tumor Endothelial Cells (TECs) as Potential Immune Directors of the Tumor Microenvironment - New Findings and Future Perspectives. Front Cell Dev Biol. 2020 Aug 19;8:766.

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is in the tenth place of newly diagnosed case of all tumors and one of the most lethal cancer related malignancies in developed countries [1]. Surgical reduction still is the main treatment method, but often not effective due to the advanced stage. Immunotherapy is a promising treatment for many cancers. However, the efforts and attempts to use immunotherapy in the treatment of PDAC are extensive and real breakthroughs have not been achieved [2]. Peripheral blood mononuclear cells (PBMC) as immune cells are crucial for the body's response against cancer [3]. One of the main immune checkpoint member is programmed cell death protein 1 (PD1). It is known that PD1 expression in T cells is strongly regulated by aryl hydrocarbon receptor (AHR) [4]. Moreover, there are immunological barriers to treatment challenge in cancer like insufficient immune function activation and excess immune suppression [5]. Previously we analyzed immune response disbalance regards of AHR expression in PDAC patients PBMCs. Results showed significant changes of lower AHR gene expression patients' group. Next, we decided to check PBMCs functions in healthy participants after indirect interaction with pancreatic cancer cells. We hypothesized that AHR expression differences in PBMCs could regulate anti-cancer response.

Aims & Methods

The aim of this study was to analyze PBMCs functions and targets expression of healthy participants after indirect interaction with pancreatic cancer cells. PBMCs (n=11) were isolated from peripheral blood by Ficoll-Paque gradient centrifugation. Analysis was done at 3 points. The first point of study was prepared immediately after PBMCs isolation. Second and third points were prepared after 20h of PBMCs -/+ co-cultivation with BxPC3 cells. Phagocytic activity analysis was prepared after 3-hour PBMCs activation with LPS. ROS production was measured effecting with TBHP. NO production was measured after activation with LPS + L-Arginine. Simultaneously PBMCs metabolic activity was measured by resazurin assay. Monocytes and lymphocytes differentiation was evaluated by FACS analysis. Gene expressions were analyzed by RT-PCR method. Statistical analysis was performed using GraphPad software and the Mann – Whitney test.

Results

The results showed that AHR and IL1b, IL6, IL10 mRNA were increased, only IL6 was significantly upregulated after PBMC co-cultivation with BxPC3 cells. While PD1 and IL4 mRNA expression were decreased. The expression of AHR strongly negatively correlated with expression of PD1 in healthy volunteers PBMCs’ after co-cultivation with cancer cells. M1 type monocytes and NK cells decreased after co-cultivation with PDAC cells. Moreover, M0 type monocytes and cytotoxic T lymphocytes were increased. Phagocytic activity and NO production were more intensive in PBMC cells after co-cultivation. ROS production did not show significant changes.

Conclusion

This study showed negative PD1 gene expression correlation with AHR expression after indirect interactions with cancer cells in healthy participants PBMCs. Also, PBMCs co-cultivation with cancer cells initiated strong functional response and higher expression of inflammatory cytokines. These results show us different AHR relations with immunity response in healthy and cancer patients. Meanwhile, this knowledge lets us understand the importance of AHR modulation as an additional PDAC treatment strategy.

References

1. Siegel, R. L., Miller, K. D., Wagle, N. S., & Jemal, A. (2023). Cancer statistics, 2023. CA: A Cancer Journal for Clinicians, 73(1), 17-48. doi:10.3322/caac.21763
2. Mundry, C. S., Eberle, K. C., Singh, P. K., Hollingsworth, M. A., & Mehla, K. (2020). Local and systemic immunosuppression in pancreatic cancer: Targeting the stalwarts in tumor’s arsenal. Biochimica Et Biophysica Acta. Reviews on Cancer, 1874(1), 188387. doi:10.1016/j.bbcan.2020.188387
3. Griffin JF, Poruk KE, Wolfgang CL. Pancreatic cancer surgery: past, present, and future. Chin J Cancer Res. 2015 Aug;27(4):332-48
4. Liu, Y., Liang, X., Dong, W., Fang, Y., Lv, J., Zhang, T., . . . Huang, B. (2018). Tumor-repopulating cells induce PD1 expression in CD8+ T cells by transferring kynurenine and AhR activation. Cancer Cell, 33(3), 480-494.e7. doi: 10.1016/j.ccell.2018.02.005
5. Rojas, L. A., & Balachandran, V. P. (2021). Scaling the immune incline in PDAC. Nature Reviews. Gastroenterology & Hepatology, 18(7), 453-454. doi:10.1038/s41575-021-00475-9

Disclosure

The authors have no conflicts of interest to declare

Introduction

Pancreatic cancer (PC) is one of the fatal malignant tumors, with a 5-year survival rate is approximately 6%, and is expected to become the second leading cause of cancer deaths by 2030. The poor prognosis of PC is associated with the tumour microenvironment, characterized by excessive fibrosis and extracellular matrix deposition, known as desmoplasia. PC’s highly desmoplastic property not only increases the malignancy of the tumor but also inhibits the penetration and diffusion of anti-cancer drugs, related to drug resistance. Chauhan et al. demonstrated that the angiotensin inhibitor (ASI), losartan reduced stromal collagen and hyaluronan production, thus improving the efficacy of chemotherapy in PC models. Several retrospective studies have reported that the ASI usage group showed higher overall survival (OS) and progression-free survival compared to the non-ASI usage group in PC patients. However, most of the previous studies on ASI in PC patients were based on retrospective data from single institutions. Recently, real-world big data study could explain this subject.

Aims & Methods

In this study, we aimed to conduct whether ASI use in PC patients improve OS using National Health Information Database of the National Health Insurance Service, which represents the entire Korean population. We assessed 209,723 participants for eligibility, diagnosed with pancreatic cancer between 2002 and 2020. A total of 64,219 PC patients were enrolled and divided into 3 groups [chronic ASI users with hypertension (HTN), n=13,239; non-ASI users with HTN, n=5,963; non-HTN group, n=45,017]. We performed propensity score matching to balance the baseline characteristics of the 3 groups and to reduce potential confounders using a logistic regression model with adjustment for the following: age; sex; region of residence (urban or rural); household income; underlying disease of HTN, diabetes mellitus, cardiovascular disease, cerebrovascular disease, chronic kidney disease or chronic pancreatitis; Charlson Comorbidity Index (0, 1 or ≥2); concurrent use of aspirin, metformin, statin; PC surgery type; radiotherapy; and chemotherapy.

Results

Multivariate analysis using Cox regression analysis for matched cohort presented ASI usage, female, younger, history of chronic pancreatitis, CCI ≥2, concurrent use of statin, and receiving distal pancreatectomy were significantly associated with longer OS. In addition, Kaplan-Meier curve for OS of 3 groups according to ASI usage showed that chronic ASI users presented significantly longer OS than non-ASI users and non-HTN group [Hazard ratio, chronic ASI users (control) versus non-ASI users 1.447; chronic ASI users (control) versus non-HTN group 1.518; P value < 0.001].

Conclusion

We found that the use of ASI after PC diagnosis is associated with longer survival in PC patients. Additional well-designed randomized controlled studies are needed to confirm these findings in the future.

References

1. Rahib, L., B. D. Smith, R. Aizenberg, A. B. Rosenzweig, J. M. Fleshman and L. M. Matrisian (2014). "Projecting cancer incidence and deaths to 2030: the unexpected burden of thyroid, liver, and pancreas cancers in the United States" Cancer Res 74(11): 2913-2921.
2. Encarnación-Rosado, J. and A. C. Kimmelman (2021). "Harnessing metabolic dependencies in pancreatic cancers." Nature Reviews Gastroenterology & Hepatology 18(7): 482-492.
3. Thomas, D. and P. Radhakrishnan (2019). "Tumor-stromal crosstalk in pancreatic cancer and tissue fibrosis." Molecular cancer 18: 1-15.
4. Chauhan, V. P., J. D. Martin, H. Liu, D. A. Lacorre, S. R. Jain, S. V. Kozin, T. Stylianopoulos, A. S. Mousa, X. Han, P. Adstamongkonkul, Z. Popović, P. Huang, M. G. Bawendi, Y. Boucher and R. K. Jain (2013). "Angiotensin inhibition enhances drug delivery and potentiates chemotherapy by decompressing tumour blood vessels." Nat Commun 4: 2516.
5. Pinter, M. and R. K. Jain (2017). "Targeting the renin-angiotensin system to improve cancer treatment: Implications for immunotherapy." Sci Transl Med 9(410).

Introduction

Pancreatic cancer (PC) is emerging as one the most lethal types of cancer with only a 10% five-year survival rate [1]. Pancreatic ductal adenocarcinoma (PDAC) being the most common among PC. Low survival rate is attributed to the fact that most incidences of PDACs are diagnosed in advanced or metastatic stages [2]. In such cases, gemcitabine remains one the first-line drugs used for treatment [2] but the efficacy of the treatment can fall short due to PDAC tumour heterogeneity and its acquired resistance to said chemotherapy [3]. The aryl hydrocarbon receptor (AHR) is a transcription factor that is commonly upregulated in pancreatic ductal adenocarcinoma [4]. AHR can be linked to Human antigen R (ELAVL1) whose expression in cell cytoplasm is positively associated with PDAC response to gemcitabine [5]. AHR can block ELAVL1 from shuttling from nucleus to cytoplasm [6] where it stabilizes its target messenger RNAs (mRNAs) and increases protein expression of said mRNAs. Among those target mRNAs are those induced by gemcitabine treatment. Frequently with upregulated AHR expression, ELAVL1 would be sequestered in the nucleus which would lead to increased chemoresistance, however the relationship between AHR and ELAVL1 in PDAC is still poorly understood.

Aims & Methods

The aim of the study was to investigate how AHR and ELAVL1 genes and proteins interacts with each other and change their cellular localization in PDAC cells.
Two PDAC cell lines were used (BxPC-3, Su.86.86). AHR and ELAVL1 genes were silenced for 24-hours by lipofectamine mediated siRNA transfection. Cellular localization change of AHR and ELAVL1 proteins was illucidated by fluorescent microscopy. After silencing of AHR or ELAVL1 genes, RNA and proteins were extracted and real-time polymerase chain reaction (RT-PCR) and Western Blot (WB) analysis were performed. Direct binding between ELAVL1 protein and AHR mRNA was investigated by immunoprecipitation assay followed by RT-PCR and WB assays.

Results

The results of gene silencing showed that silencing of AHR can increase ELAVL1 mRNA and protein and silencing of ELAVL1 can decrease AHR mRNA and protein expression. The results of immunoprecipitation assay showed that ELAVL1 protein binds AHR mRNA resulting in mRNA stabilization. This shows a direct link between these two molecules. Moreover, silencing of AHR resulted in increased ELAVL1 protein concentration in cytoplasm which indicates that AHR blocks ELAVL1 shuttling from nucleus to cytoplasm. Silencing of ELAVL1 did not alter the cellular localization of AHR protein.

Conclusion

A direct relationship between AHR and ELAVL1 can be seen in PDAC. Moreover, by decreasing AHR expression it is possible to increase ELAVL1 shuttling from nucleus to cytoplasm which might contribute to PDAC cells responding better to chemotherapy.

References

[1] Hu JX, Zhao CF, Chen WB, Liu QC, Li QW, Lin YY, Gao F. Pancreatic cancer: A review of epidemiology, trend, and risk factors. World journal of gastroenterology. 2021 Jul 7;27(27):4298.
[2] Kamisawa T, Wood LD, Itoi T, Takaori K. Pancreatic cancer. The Lancet. 2016 Jul 2;388(10039):73-85.
[3] Juiz NA, Iovanna J, Dusetti N. Pancreatic cancer heterogeneity can be explained beyond the genome. Frontiers in oncology. 2019 Apr 5;9:246.
[4] Koliopanos A, Kleeff J, Xiao Y, Safe S, Zimmermann A, Büchler MW, Friess H. Increased arylhydrocarbon receptor expression offers a potential therapeutic target for pancreatic cancer. Oncogene. 2002 Sep;21(39):6059-70
[5] Richards, N. G., Rittenhouse, D. W., Freydin, B., Cozzitorto, J. A., Grenda, D., Rui, H., ... & Witkiewicz, A. K. (2010). HuR status is a powerful marker for prognosis and response to gemcitabine-based chemotherapy for resected pancreatic ductal adenocarcinoma patients. Annals of surgery, 252(3), 499-506.
[6] Zago, M., Sheridan, J. A., Nair, P., Rico de Souza, A., Gallouzi, I. E., Rousseau, S., ... & Baglole, C. J. (2013). Aryl hydrocarbon receptor-dependent retention of nuclear HuR suppresses cigarette smoke-induced cyclooxygenase-2 expression independent of DNA-binding. PLoS One, 8(9), e74953.

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is a highly metastatic disease refractory to all targeted and immune therapies. However, our understanding of PDAC microenvironment especially the metastatic microenvironment is very limited partly due to the inaccessibility to metastatic tumor tissues. Here, we aim to carry out the first single-cell transcriptome analysis of PDAC metastatic microenvironment in matched primary tumor and liver metastasis tissues.

Aims & Methods

We collected synchronously resected PDAC primary tumors and matched liver metastases from four PDAC patients and performed single-cell RNA sequencing. We performed comparative analysis on both cellular composition and functional phenotyping between primary and metastatic tumors. We also performed trajectory analysis and cellular interactome analysis in the metastasis microenvironment.

Results

Epithelial cancer cells exhibited distinct transcriptomic profile in liver metastasis with clearly defined evolutionary routes from primary tumor. We identified specific subtypes of stromal and immune cells critical to the formation of the pro-tumor microenvironment in metastatic lesions, including RGS5⁺ cancer-associated fibroblasts, CCL18⁺ lipid-associated macrophages, S100A8⁺ neutrophils and FOXP3⁺ regulatory T cells. We also identified several new biomarkers associated with PDAC metastasis and poor prognosis, such as LITAF, SPRR1B, KLK7 and LGALS9. Cellular interactome analysis further revealed that the lack of tumor-immune cell interaction in metastatic tissues contributes to the formation of the immunosuppressive microenvironment.

Conclusion

Our study provides a comprehensive characterization of the transcriptional landscape of PDAC liver metastasis, which deepens our understanding of the mechanisms underlying the immunosuppressive microenvironment in metastatic PDAC and serves as a valuable resource for the design of novel targeted therapies.

Disclosure

No

Introduction

The retroperitoneal location of the pancreas represents a challenge for the early identification of chronic pancreatitis (CP) and pancreas neoplasms (PN). Imaging and endoscopic ultrasound (EUS) with elastography are helpful resources with variable accuracy. EUS-guided shear wave elastography (EUS-SW) measures tissue elasticity by shear waves inside the organ through acoustic radiation force impulse. Its accuracy has not been assessed for CP and solid PN.

Aims & Methods

We aim to estimate the diagnostic accuracy of EUS-SW for CP and solid PN. Cases with a recently confirmed diagnosis of CP or PN who underwent EUS-SW (12/2020-11/2022) were included. Cases that required EUS for subepithelial lesion assessment comprised a third study group (controls, CG). Age, gender, body mass index (BMI), diabetes, fatty liver disease, alcohol, and tobacco consumption were recorded. An expert endoscopist (RVZ) performed EUS, assessed fatty pancreas score, strain ratio (SR), histogram (SH), and ten measurements of EUS-SW elasticity (SWE) and dispersion (SWD) per case. SWE and SWD variation were based on quotient among interquartile range (IQR) and median. In cases with <30% variation, the association of SWE and SWD was estimated with study groups, baseline and EUS data through multivariate ANOVA regression. SWE and SWD cut-off values were calculated with Youden’s index. Data was analysed in R v4.0.

Results

88 cases were enrolled (37 CG, 14 CP, 37 PN), median age of 62.5 (IQR 53-70), 47.7% female. The proportion of fatty pancreas score II and III was similar in CG (37.8% and 56.8%) and CP (57.1% and 42.9%); there was a significant difference in PN, with a score II, III, and IV of 24.3%, 40.5%, and 29.7%, respectively (p<.001). Median SR was also similar among CG (3.6; IQR 2.9 - 4.3) and CP (3.9; IQR 3.6 - 4.7) but significantly different compared with PN (7.6; IQR 5.3 - 11.1; p<.001). Median SWE and SWD were 10.5 kPa (7.3 - 16.6) and 1.9 [m/s]/kHz (1.6 - 2.4), with a <30% variation in 27 (30.7%) and 57 (64.8%), respectively (table 1). Study groups (CG/CP/PN) and diabetes were associated to a higher SWE and SWD (p<.05). Predicted SWE were significantly different among PN vs CG (-13.1; -24.4 to -1.7; p=.0242) (figure 1). An SWE ≥17.4 diagnosed PN with a sensitivity, specificity, positive and negative predictive values of 75% (19-99), 91% (72-99), 60% (27-99) and 95% (63-99); while an SWD ≥2.31 predicted PN with a diagnostic accuracy of 54% (25-81), 70% (55-83), 35% (21-66), 84% (60-91), respectively.

Conclusion

EUS-SW constitutes an objectively valuable measuring tool for CP and PN diagnostic workups. Larger multicentric diagnostic trials are needed to confirm these findings. NCT05095831.

References

1. Xia C, Yin H, Zhang K, Wang Z, Yang X, Huang H. The global research status and trends of the application of endoscopic ultrasonography in pancreatic tumors over the last decades: A bibliometric study. Front Oncol. 2022 Aug 12;12:980415.
2. Salah NY, Madkour SS, Soliman KS. Pancreatic shear wave elastography in children with type 1 diabetes: relation to diabetes duration, glycemic indices, fasting C-peptide and diabetic complications. Pediatr Radiol. 2022 Nov;52(12):2348-2358. doi: 10.1007/s00247-022-05363-1. Epub 2022 Apr 23.
3. Fujita Y, Kitago M, Abe T, et al. Evaluation of pancreatic fibrosis with acoustic radiation force impulse imaging and automated quantification of pancreatic tissue components. Pancreas. 2018;47:1277–1282.
4. Jesper D, Fiedler S, Klett D, Waldner MJ, Schellhaas B, Zundler S, Neurath MF, Pfeifer L. Shear Wave Dispersion Imaging for the Characterization of Focal Liver Lesions - A Pilot study. Ultraschall Med. 2022 Oct;43(5):507-513.
5. Korta Martiartu N, Nambiar S, Nascimento Kirchner I, Paverd C, Cester D, Frauenfelder T, Ruby L, Rominger MB. Sources of Variability in Shear Wave Speed and Dispersion Quantification with Ultrasound Elastography: A Phantom Study. Ultrasound Med Biol. 2021 Dec;47(12):3529-3542.

Disclosure

Carlos Robles-Medranda is a key opinion leader and consultant for Pentax Medical, Boston Scientific, Steris, Medtronic, Motus, Micro-tech, G-Tech Medical Supply, CREO Medical, EndoSound, and Mdconsgroup. The other authors declare no conflicts of interest.