Human P2X7 receptor variants Gly150Arg and Arg276His polymorphisms have differential effects on risk association and cellular functions in pancreatic cancer.

BACKGROUND: The purinergic P2X7 receptor (P2X7R) plays an important role in the crosstalk between pancreatic stellate cells (PSCs) and cancer cells, thus promoting progression of pancreatic ductal adenocarcinoma (PDAC). Single nucleotide polymorphisms (SNPs) in the P2X7R have been reported for several cancers, but have not been explored in PDAC. MATERIALS AND METHODS: Blood samples from PDAC patients and controls were genotyped for 11 non-synonymous SNPs in P2X7R and a risk analysis was performed. Relevant P2X7R-SNP GFP variants were expressed in PSCs and cancer cells and their function was assayed in the following tests. Responses in Ca2+ were studied with Fura-2 and dye uptake with YO-PRO-1. Cell migration was monitored by fluorescence microscopy. Released cytokines were measured with MSD assay. RESULTS: Risk analysis showed that two SNPs 474G>A and 853G>A (rs28360447, rs7958316), that lead to the Gly150Arg and Arg276His variants, had a significant but opposite risk association with PDAC development, protecting against and predisposing to the disease, respectively. In vitro experiments performed on cancer cells and PSCs expressing the Gly150Arg variant showed reduced intracellular Ca2+ response, fluorescent dye uptake, and cell migration, while the Arg276His variant reduced dye uptake but displayed WT-like Ca2+ responses. As predicted, P2X7R was involved in cytokine release (IL-6, IL-1ß, IL-8, TNF-α), but the P2X7R inhibitors displayed varied effects. CONCLUSION: In conclusion, we provide evidence for the P2X7R SNPs association with PDAC and propose that they could be considered as potential biomarkers.

Levels of type XVII collagen (BP180) ectodomain are elevated in circulation from patients with multiple cancer types and is prognostic for patients with metastatic colorectal cancer.

BACKGROUND: Collagens are the major components of the extracellular matrix (ECM) and are known to contribute to tumor progression and metastasis. There are 28 different types of collagens each with unique functions in maintaining tissue structure and function. Type XVII collagen (BP180) is a type II transmembrane protein that provides stable adhesion between epithelial cells and the underlying basement membrane. Aberrant expression and ectodomain shedding of type XVII collagen have been associated with epithelial damage, tumor invasiveness, and metastasis in multiple tumor types and may consequently be used as a potential (non-invasive) biomarker in cancer and treatment target. METHOD: An ELISA targeting the type XVII collagen ectodomain (PRO-C17) was developed for use in serum. PRO-C17 was measured in a cohort of patients with 11 different cancer types (n = 214) and compared to healthy controls (n = 23) (cohort 1). Based on the findings from cohort 1, PRO-C17 and its association with survival was explored in patients with metastatic colorectal cancer (mCRC) treated with bevacizumab in combination with chemotherapy (n = 212) (cohort 2). RESULTS: PRO-C17 was robust and specific towards the ectodomain of type XVII collagen. In cohort 1, PRO-C17 levels were elevated (p < 0.05) in serum from patients with CRC, kidney, ovarian, bladder, breast, and head and neck cancer compared to healthy controls. PRO-C17 was especially good at discriminating between CRC patients and healthy controls with an AUROC of 0.904. In cohort 2, patients with mCRC and high levels (tertile 3) of PRO-C17 had shorter overall survival (OS) with a median OS of 390 days compared to 539 days for patients with low levels of PRO-C17. When evaluated by multivariate Cox regression analysis, high PRO-C17 was predictive for poor OS independent of risk factors and the tumor fibrosis biomarker PRO-C3. CONCLUSION: PRO-C17 measures the ectodomain of type XVII collagen in serum and is a promising non-invasive biomarker that can aid in understanding tumor heterogeneity as well as elaborate on the role of collagen XVII in tumor progression. Moreover, the findings in the study proposes PRO-C17 as novel biomarker of epithelial damage in specific cancer types including CRC.

Plasma YKL-40 is associated with prognosis in patients with metastatic pancreatic cancer receiving immune checkpoint inhibitors in combination with radiotherapy.

Background: YKL-40, also known as chitinase-3-like protein 1 (CHI3L1), is a secreted glycoprotein produced by various cell types including stromal, immune, and cancer cells. It contributes to cancer progression through tumor-promoting inflammation and has been shown to inhibit the cytotoxicity of T and NK lymphocytes. In vivo studies have demonstrated synergistic anti-cancer effects of blocking YKL-40 in combination with immune checkpoint inhibitors (ICIs). Biomarkers for the prediction of the response to ICIs are highly needed. We investigated the association between plasma YKL-40 and clinical benefit and survival in patients with metastatic pancreatic cancer (mPC) receiving ICIs and stereotactic body radiotherapy (SBRT). Methods: Blood samples were collected from 84 patients with mPC who participated in the randomized phase II CheckPAC study, in which patients received nivolumab with or without ipilimumab combined with a single fraction of SBRT. Plasma YKL-40 was measured using a commercial ELISA kit. Results: Elevated baseline plasma YKL-40 was an independent predictor of shorter overall survival (OS) (HR 2.19, 95% CI 1.21-3.95). A ≥ 40% decrease in plasma YKL-40 during treatment was associated with longer progression-free survival (p = 0.009) and OS (p = 0.0028). There was no correlation between plasma YKL-40 and the tumor burden marker CA19-9 at baseline or during treatment. Conclusion: This study contributes new knowledge regarding YKL-40 as a predictor of clinical benefit from ICIs and radiotherapy. These exploratory results warrant further investigation of YKL-40 as a biomarker for patients treated with immunotherapies. Clinical trial registration: Clinicaltrials.gov, identifier NCT02866383.

High serum levels of the C-propetide of type V collagen (PRO-C5) are prognostic for short overall survival in patients with pancreatic ductal adenocarcinoma.

Introduction: Pancreatic ductal adenocarcinoma (PDAC) is characterized by a pronounced fibrotic tumor microenvironment, which impairs treatment response. Type I and V collagens are responsible for the densely packed fibrils in the tumor fibrosis environment. While the role of the major type I collagen in cancer is well described, less is known about the minor type V collagen. Quantifying collagen propeptides in serum has been shown to have prognostic and predictive value. In this study, we evaluated the clinical utility of measuring the propeptide of type V collagen (PRO-C5) in serum from a discovery cohort and a validation cohort of patients with PDAC as well as in non-pancreatic solid tumor types to explore the relevance of the PRO-C5 biomarker in cancer. Methods: Serum PRO-C5 was measured in three cohorts: a discovery cohort (19 healthy controls, 12 patients with chronic pancreatitis and 33 patients with PDAC (stage I-IV)), a validation cohort (800 patients with PDAC (stage I-IV)), and a non-pancreatic solid tumor type cohort of 33 healthy controls and 200 patients with 10 different non-pancreatic solid tumor types. The levels of serum PRO-C5 in patients with cancer were compared to levels in healthy controls. The association between PRO-C5 levels and overall survival (OS) was evaluated in patients with PDAC after adjusting for established prognostic factors. Results: PRO-C5 was significantly increased in serum from patients with PDAC compared to healthy controls (p < 0.001). High PRO-C5 levels were significantly associated with short OS in both the discovery- and the validation cohort, especially in early stages of PDAC (validation cohort stage II, HR = 2.0, 95%CI1.2-3.4). The association was independent of other prognostic parameters including stage, performance status and CA19-9. Furthermore, serum levels of PRO-C5 were significantly increased in serum from patients with other non-pancreatic solid tumor types compared to healthy controls. Conclusion: High levels of serum PRO-C5 is prognostic for short OS in patients with PDAC and may provide clinical value in many other tumor types beyond PDAC. This underlines the importance of type V collagen in tumor fibrosis. PRO-C5 could have the potential to be used in several aspects within drug discovery, patient stratification and drug efficacy.

Combining sCD163 with CA 19-9 Increases the Predictiveness of Pancreatic Ductal Adenocarcinoma.

The objective of this study was to evaluate the diagnostic and prognostic potential of soluble CD163 (sCD163) in patients with pancreatic ductal adenocarcinoma (PDAC). Preoperative serum samples from 255 patients with PDAC were analyzed for sCD163 using a commercially available enzyme-linked immunosorbent assay. The diagnostic value of sCD163 was evaluated using receiver operating characteristic (ROC) curves. The prognostic significance of sCD163 was evaluated by Cox regression analysis and Kaplan-Meier survival curves. sCD163 was significantly increased in patients with PDAC, across all stages, compared to healthy subjects (stage 1: p value = 0.033; stage 2-4: p value ≤ 0.0001). ROC curves showed that sCD163 combined with CA 19-9 had the highest diagnostic potential compared to sCD163 and CA 19-9 alone both in patients with local PDAC and patients with advanced PDAC. Univariate and multivariate analysis showed no association between sCD163 and overall survival. This study found elevated levels of circulating sCD163 in patients with PDAC, regardless of stage, compared to healthy subjects. This suggests that sCD163 may have a clinical value as a novel diagnostic biomarker in PDAC.

Prognostic value of circulating proteins in patients undergoing surgery for pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death. Less than 20% of patients are diagnosed with resectable disease. Identifying truly resectable disease is challenging because 20%-40% of the patients subjected to resection are found to have advanced disease during surgery. The aim of our study was to identify panels of circulating proteins that could be used to distinguish patients with unresectable PDAC from patients with resectable PDAC and to identify prognostic signatures for both groups. METHODS: We measured 92 circulating immuno-oncology-related proteins using the proximity extension assay from Olink Proteomics in 273 patients eligible for surgery for PDAC. Two bioinformaticians worked independently of one another on the same data. LASSO and Ridge regression were used in the statistical analyses. RESULTS: One protein index for determining resectability had an AUC value of 0.66. Several indices for prognosis had AUC values between 0.50 and 0.75 and were therefore not better than existing prognostic markers. DISCUSSION: Our study did not reveal any new high-performing protein panels that could be used to identify patients with inoperable PDAC before surgery. The panel of 92 proteins investigated has previously been found to be applicable for diagnostic use in patients with PDAC, but it does not seem to warrant further investigation regarding resectability in the subgroup of patients with PDAC referred to surgery.

Circulating Protein Biomarkers for Prognostic Use in Patients with Advanced Pancreatic Ductal Adenocarcinoma Undergoing Chemotherapy.

Patients with advanced pancreatic ductal adenocarcinoma (PDAC) have a dismal prognosis. We aimed to find a prognostic protein signature for overall survival (OS) in patients with advanced PDAC, and to explore whether early changes in circulating-protein levels could predict survival. We investigated 92 proteins using the Olink Immuno-Oncology panel in serum samples from 363 patients with advanced PDAC. Protein panels for several survival cut-offs were developed independently by two bioinformaticians using LASSO and Ridge regression models. Two panels of proteins discriminated patients with OS < 90 days from those with OS > 2 years. Index I (CSF-1, IL-6, PDCD1, TNFRSF12A, TRAIL, TWEAK, and CA19-9) had AUCs of 0.99 (95% CI: 0.98−1) (discovery cohort) and 0.89 (0.74−1) (replication cohort). For Index II (CXCL13, IL-6, PDCD1, and TNFRSF12A), the corresponding AUCs were 0.97 (0.93−1) and 0.82 (0.68−0.96). Four proteins (ANGPT2, IL-6, IL-10, and TNFRSF12A) were associated with survival across all treatment groups. Longitudinal samples revealed several changes, including four proteins that were also part of the prognostic signatures (CSF-1, CXCL13, IL-6, TNFRSF12A). This study identified two circulating-protein indices with the potential to identify patients with advanced PDAC with very short OS and with long OS.

Fibrotic activity quantified in serum by measurements of type III collagen pro-peptides can be used for prognosis across different solid tumor types.

Due to activation of fibroblast into cancer-associated fibroblasts, there is often an increased deposition of extracellular matrix and fibrillar collagens, e.g. type III collagen, in the tumor microenvironment (TME) that leads to tumor fibrosis (desmoplasia). Tumor fibrosis is closely associated with treatment response and poor prognosis for patients with solid tumors. To assure that the best possible treatment option is provided for patients, there is medical need for identifying patients with high (or low) fibrotic activity in the TME. Measuring unique collagen fragments such as the pro-peptides released into the bloodstream during fibrillar collagen deposition in the TME can provide a non-invasive measure of the fibrotic activity. Based on data from 8 previously published cohorts, this review provides insight into the prognostic value of quantifying tumor fibrosis by measuring the pro-peptide of type III collagen in serum of a total of 1692 patients with different solid tumor types and discusses the importance of tumor fibrosis for understanding prognosis and for potentially guiding future drug development efforts that aim at overcoming the poor outcome associated with a fibrotic TME.

Collagen Biomarkers Quantify Fibroblast Activity In Vitro and Predict Survival in Patients with Pancreatic Ductal Adenocarcinoma.

The use of novel tools to understand tumour-fibrosis in pancreatic ductal adenocarcinoma (PDAC) and novel anti-fibrotic treatments are highly needed. We established a pseudo-3D in vitro model including humane pancreatic fibroblasts (PFs) and pancreatic cancer-associated fibroblasts (CAFs) in combination with clinical collagen biomarkers, as a translational anti-fibrotic drug screening tool. Furthermore, we investigated the prognostic potential of serum collagen biomarkers in 810 patients with PDAC. PFs and CAFs were cultured in Ficoll-media. Cells were treated w/wo TGF-ß1 and the anti-fibrotic compound ALK5i. Biomarkers measuring the formation of type III (PRO-C3) and VI (PRO-C6) collagens were measured by ELISA in supernatant at days 3, 6, 9, and 12. PRO-C3 and PRO-C6, and their association with overall survival (OS), were evaluated in serum with PDAC (n = 810). PRO-C3 and PRO-C6 were upregulated in CAFs compared to PFs (p < 0.0001.). TGF-ß1 increased PRO-C3 in both PFs and CAFs (p < 0.0001). The anti-fibrotic compound ALK5i inhibited both PRO-C3 and PRO-C6 (p < 0.0001). High serum levels of PRO-C3 and PRO-C6 in patients with PDAC were associated with short OS (PRO-C3: HR = 1.48, 95%CI: 1.29-1.71, p < 0.0001 and PRO-C6: HR = 1.31, 95%CI: 1.14-1.50, p = 0.0002). PRO-C3 and PRO-C6 have the potential to be used both pre-clinically and clinically as a measure of tumor fibrosis and CAF activity.

Inflammatory Biomarker Score Identifies Patients with Six-Fold Increased Risk of One-Year Mortality after Pancreatic Cancer.

We examined whether elevated plasma C-reactive protein (CRP), carbohydrate antigen (CA) 19-9, interleukin-6 (IL-6) and YKL-40, individually or combined, can identify poor survivors among patients with pancreatic ductal adenocarcinoma (PDAC). We measured CRP, CA 19-9, IL-6 and YKL-40 in 993 patients at the time of PDAC diagnosis. The biomarker score was the sum of biomarker categories, coded 0, 1 and 2 for low, intermediate and high plasma concentrations, respectively. High vs. low levels of CRP, CA 19-9 and IL-6 were each independently associated with a two-fold increased risk of one-year mortality. CRP performed best in patients with advanced and CA 19-9 in patients with low cancer stages. YKL-40 was not associated with mortality and, therefore, was not included in the biomarker score. Compared to the biomarker score = 0, the multifactorially adjusted hazard ratios for one-year mortality were 1.56 (95% confidence interval: 0.99-2.44) for score = 1, 2.22 (1.41-3.49) for score = 2, 3.44 (2.20-5.38) for score = 3, 5.13 (3.21-8.17) for score = 4 and 6.32 (3.84-10.41) for score = 5-6 (p-value for trend = 3 × 10-31). This score performed better than any single biomarker or combination of biomarkers when examined in similarly sized or other categories. In conclusion, a combination score of elevated CRP, CA 19-9 and IL-6 identified patients with six-fold higher one-year mortality.

Lack of association of CD44-rs353630 and CHI3L2-rs684559 with pancreatic ductal adenocarcinoma survival.

Although pancreatic ductal adenocarcinoma (PDAC) survival is poor, there are differences in patients' response to the treatments. Detection of predictive biomarkers explaining these differences is of the utmost importance. In a recent study two genetic markers (CD44-rs353630 and CHI3L2-rs684559) were reported to be associated with survival after PDAC resection. We attempted to replicate the associations in 1856 PDAC patients (685 resected with stage I/II) from the PANcreatic Disease ReseArch (PANDoRA) consortium. We also analysed the combined effect of the two genotypes in order to compare our results with what was previously reported. Additional stratified analyses considering TNM stage of the disease and whether the patients received surgery were also performed. We observed no statistically significant associations, except for the heterozygous carriers of CD44-rs353630, who were associated with worse OS (HR = 5.01; 95% CI 1.58-15.88; p = 0.006) among patients with stage I disease. This association is in the opposite direction of those reported previously, suggesting that data obtained in such small subgroups are hardly replicable and should be considered cautiously. The two polymorphisms combined did not show any statistically significant association. Our results suggest that the effect of CD44-rs353630 and CHI3L2-rs684559 cannot be generalized to all PDAC patients.

Circulating Protein Biomarkers for Use in Pancreatic Ductal Adenocarcinoma Identification.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal solid tumors. Most patients are diagnosed at an advanced stage where curative surgery is not an option. The aim of this study was to identify a panel of circulating proteins that could distinguish patients with PDAC from non-PDAC individuals. EXPERIMENTAL DESIGN: We investigated 92 proteins known to be involved in inflammation, development, and progression of PDAC using the Olink immuno-oncology panel in serum samples from 701 patients with PDAC (stage I-IV), 102 patients with nonmalignant pancreatic diseases, and 180 healthy blood donors. Patients were included prospectively between 2008 and 2018. Plasma carbohydrate antigen 19-9 (CA19-9) was measured in all samples. The protein panels with the best diagnostic performances were developed by two bioinformaticians working independently, using LASSO and Ridge regression models. RESULTS: Two panels of proteins (index I, containing 9 proteins + CA19-9, and index II, containing 23 proteins + CA19-9) were identified. Index I was able to discriminate patients with PDAC from all patients with non-PDAC, with a ROC AUC value of 0.92 [95% confidence interval (CI), 0.89-0.96] in the discovery cohort and 0.92 (95% CI, 0.87-0.97) in the replication cohort. For index II, the AUC value was 0.96 (95% CI, 0.95-0.98) in the discovery cohort and 0.93 (95% CI, 0.90-0.96) in the replication cohort. All nine serum proteins of index I were found in index II. CONCLUSIONS: This study identified two circulating protein indices with the potential to discriminate between individuals with and without PDAC.

Noninvasive prognostic biomarker potential of quantifying the propeptides of Type XI collagen alpha-1 chain (PRO-C11) in patients with pancreatic ductal adenocarcinoma.

Type XI collagen has been associated with tumor fibrosis and aggressiveness in patients with pancreatic ductal adenocarcinoma (PDAC). The propeptide on Type XI collagen is released into the circulation after proteolytic processing at either amino acid 253 or 511. This allows for a noninvasive biomarker approach to quantify Type XI collagen production. We developed two ELISA-based biomarkers, targeting the two enzymatic cleavage sites (PRO-C11-253 and PRO-C11-511). In a discovery cohort including serum from patients with PDAC (n = 39, Stages 1-4), chronic pancreatitis (CP, n = 12) and healthy controls (n = 20), PRO-C11-511, but not PRO-C11-253, was significantly upregulated in patients with PDAC and CP compared to healthy controls. Furthermore, PRO-C11-511 levels >75th percentile were associated with poor overall survival (OS) (HR, 95% CI: 3.40, 1.48-7.83). The PRO-C11-511 biomarker potential was validated in serum from 686 patients with PDAC. Again, high levels of PRO-C11-511 (>75th percentile) were associated with poor OS (HR, 95% CI: 1.68, 1.40-2.02). Furthermore, PRO-C11-511 remained significant after adjusting for clinical risk factors (HR, 95% CI: 1.50, 1.22-1.86). In conclusion, quantifying serum levels of Type XI collagen with PRO-C11-511 predicts poor OS in patients with PDAC. This supports that Type XI collagen is important for PDAC biology and that PRO-C11-511 has prognostic noninvasive biomarker potential for patients with PDAC.

Prognostic value of blood-based fibrosis biomarkers in patients with metastatic colorectal cancer receiving chemotherapy and bevacizumab.

A desmoplastic colorectal cancer stroma, characterized by excess turnover of the cancer-associated fibroblast derived collagens type III and VI, can lead to reduced drug-uptake and poor treatment response. We investigated the association between biomarkers of collagen type III and VI and overall survival (OS) in patients with metastatic colorectal cancer (mCRC). Serum samples were collected from 252 patients with mCRC prior to treatment with bevacizumab and chemotherapy. Serum concentrations of biomarkers reflecting formation of collagen type III (PRO-C3) and VI (PRO-C6) and degradation of collagen type VI (C6M and C6Mα3) were determined by ELISA. The biomarkers were evaluated for associations with OS, individually, combined, and after adjusting for carcinoembryonic antigen (CEA), lactate dehydrogenase (LDH) and performance status (PS). High baseline levels (> median) of each collagen biomarker were significantly associated with shorter OS (PRO-C3: HR = 2.0, 95%CI = 1.54-2.63; PRO-C6: HR = 1.6, 95%CI = 1.24-2.11; C6M: HR = 1.4, 95%CI = 1.05-1.78; C6Mα3: HR = 1.6, 95%CI = 1.16-2.07). PRO-C3 and PRO-C6 remained significant after adjustment for CEA, LDH and PS. Weak correlations were seen between the collagen biomarkers (r = 0.03-0.59) and combining all improved prognostic capacity (HR = 3.6, 95%CI = 2.30-5.76). Collagen biomarkers were predictive of shorter OS in patients with mCRC. This supports that collagen- and CAF biology is important in CRC.

Pre-treatment serum vitamin D deficiency is associated with increased inflammatory biomarkers and short overall survival in patients with pancreatic cancer.

BACKGROUND: Vitamin D deficiency and inflammation are associated with increased mortality. We investigated the relationship between pre-treatment serum vitamin D levels, inflammatory biomarkers (IL-6, YKL-40 and CRP) and overall survival (OS) in pancreatic ductal adenocarcinoma (PDAC) patients. METHODS: Pre-treatment serum vitamin D, IL-6, YKL-40 and CRP levels were determined in 1,267 patients with PDAC enrolled from July 2008 to September 2018 in the prospective BIOPAC study (NCT03311776). The patients were grouped according to vitamin D levels: sufficient >50 nmol/L, insufficient 25-50 nmol/L and deficient <25 nmol/L. RESULTS: Across all tumour stages, vitamin D-deficient patients had the highest median levels of IL-6 (8.3 pg/mL, range 0.7-91), YKL-40 (177 ng/ml, range 25-5279) and CRP (15.5 mg/L, range 0.8-384). The resected stage I and II patients with vitamin D deficiencies had a shorter median OS, 18.3 months (95% CI, 12.1-31.5 months) than those with sufficient levels, 29.7 months (95% CI, 22.3-36.1 months), and the hazard ratio for death was 1.55 (95% CI, 1.04-2.31; p = 0.03). In advanced PDAC, there was no significant difference in OS between the vitamin D groups. CONCLUSIONS: Vitamin D deficiency was associated with increased inflammatory biomarkers in all PDAC stages. The resected stage I and II patients with sufficient vitamin D levels had a higher OS than those with a vitamin D deficiency. However, there was no correlation between vitamin D levels and survival in advanced PDAC. Future studies need to investigate vitamin D supplementation effects on survival in PDAC.

Serum IL6 as a Prognostic Biomarker and IL6R as a Therapeutic Target in Biliary Tract Cancers.

PURPOSE: Biliary tract cancer (BTC) is a heterogeneous group of rare gastrointestinal malignancies with dismal prognosis often associated with inflammation. We assessed the prognostic value of IL6 and YKL-40 compared with CA19-9 before and during palliative chemotherapy. We also investigated in mice whether IL6R inhibition in combination with gemcitabine could prolong chemosensitivity. EXPERIMENTAL DESIGN: A total of 452 Danish participants with advanced (locally advanced and metastatic) BTC were included from six clinical trials (February 2004 to March 2017). Serum CA19-9, IL6, and YKL-40 were measured before and during palliative treatment. Associations between candidate biomarkers and progression-free survival (PFS) and overall survival (OS) were analyzed by univariate and multivariate Cox regression. Effects of inhibiting IL6R and YKL-40 were assessed in vitro, and of IL6R inhibition in vivo. RESULTS: High pretreatment levels of CA19-9, IL6, and YKL-40, and increasing levels during treatment, were associated with short PFS and OS in patients with advanced BTC. IL6 provided independent prognostic information, independent of tumor location and in patients with normal serum CA19-9. ROC analyses showed that IL6 and YKL-40 were predictive of very short OS (OS < 6 months), whereas CA19-9 was best to predict OS > 1.5 years. Treatment with anti-IL6R and gemcitabine significantly diminished tumor growth when compared with gemcitabine monotherapy in an in vivo transplant model of BTC. CONCLUSIONS: Serum IL6 and YKL-40 are potential new prognostic biomarkers in BTC. IL6 provides independent prognostic information and may be superior to CA19-9 in certain contexts. Moreover, anti-IL6R should be considered as a new treatment option to sustain gemcitabine response in patients with BTC.

Collagen Density Modulates the Immunosuppressive Functions of Macrophages.

Tumor-associated macrophages (TAMs) support tumor growth by suppressing the activity of tumor-infiltrating T cells. Consistently, TAMs are considered a major limitation for the efficacy of cancer immunotherapy. However, the molecular reason behind the acquisition of an immunosuppressive TAM phenotype is not fully clarified. During tumor growth, the extracellular matrix (ECM) is degraded and substituted with a tumor-specific collagen-rich ECM. The collagen density of this tumor ECM has been associated with poor patient prognosis but the reason for this is not well understood. In this study, we investigated whether the collagen density could modulate the immunosuppressive activity of TAMs. The murine macrophage cell line RAW 264.7 was three-dimensionally cultured in collagen matrices of low and high collagen densities mimicking healthy and tumor tissue, respectively. Collagen density did not affect proliferation or viability of the macrophages. However, whole-transcriptome analysis revealed a striking response to the surrounding collagen density, including the regulation of immune regulatory genes and genes encoding chemokines. These transcriptional changes were shown to be similar in murine bone marrow-derived macrophages and TAMs isolated from murine tumors. Strikingly, coculture assays with primary T cells showed that macrophages cultured in high-density collagen were less efficient at attracting cytotoxic T cells and capable of inhibiting T cell proliferation more than macrophages cultured in low-density collagen. Our study demonstrates that a high collagen density can instruct macrophages to acquire an immunosuppressive phenotype. This mechanism could reduce the efficacy of immunotherapy and explain the link between high collagen density and poor prognosis.

Prognostic impact of Charlson's Age-Comorbidity Index and other risk factors in patients with pancreatic cancer.

OBJECTIVES: Few studies have evaluated the impact of risk factors and comorbidity on overall survival (OS) in patients with pancreatic ductal adenocarcinoma (PDAC). The aim was to investigate the prognostic importance of Charlson's age-comorbidity index (CACI) and other risk factors on prognosis in a clinical real-world cohort of PDAC patients. METHODS: A total of 1,159 patients with PDAC who had received at least one cycle of adjuvant or palliative chemotherapy were included from the Danish BIOPAC study. We analysed OS according to CACI, tobacco smoking, alcohol intake, performance status (PS), BMI and diabetes. Hazard ratios (HRs) and 95% confidence intervals (CIs) were estimated for OS using Cox proportional hazards regression. RESULTS: At the end of follow-up, 994 (86%) patients had died. The median OS was 298 days for all patients (range 3-3010) and shortest in patients with stage IV. No association with short OS was seen for CACI > 2, diabetes, alcohol abuse, tobacco smoking, hypertension, and high BMI. Multivariate analysis showed that stage (IV vs. I: HR = 9.05, 95% CI 5.17-15.84), PS (2 vs. 0: HR = 3.67, 2.92-4.61) and treatment with angiotensin-converting enzyme inhibitors (yes vs. no: HR = 1.31, 1.06-1.61) were independent negative prognostic factors. CONCLUSIONS: We found that CACI, diabetes, tobacco smoking, alcohol abuse, hypertension, and high BMI were not associated with OS in a real-world cohort of patients with PDAC treated with chemotherapy. Only stage and PS were prognostic parameters.

Prognostic Value of Combined Detection of Serum IL6, YKL-40, and C-reactive Protein in Patients with Unresectable Pancreatic Cancer.

BACKGROUND: IL6 and YKL-40 (also known as chitinase 3-like 1 protein, CHI3L1) are produced by pancreatic cancer cells and macrophages and activate inflammation. C-reactive protein (CRP) is synthesized mainly in hepatic cells and primarily stimulated by IL6. The aim of this study was to determine the prognostic value of combined detection of serum IL6, YKL-40, and CRP in patients with pancreatic cancer receiving palliative chemotherapy. METHODS: In all, 592 patients with unresectable pancreatic cancer from five hospitals in Denmark were included in the BIOPAC biomarker study between 2008 and 2017. Pretreatment and longitudinal serum values of IL6 and YKL-40 were determined. Baseline CRP and CA19-9 values were available for the whole cohort. Patients were dichotomized as low versus high using cutoffs for IL6 of >4.92 pg/mL, YKL-40 of >95% age-corrected percentile, and CRP of >10 mg/L. The main outcome was overall survival. RESULTS: Combined elevations of serum IL6, YKL-40, and CRP were associated with worse survival in contrast to an isolated high concentration of a single marker. Serum IL6, YKL-40, and CRP were higher in patients with advanced stage disease and in patients with poor performance status. Higher IL6 and YKL-40 levels at the time of tumor progression and serum IL6 measured over time were associated with shorter overall survival. CONCLUSIONS: Combined high baseline serum levels of IL6, YKL-40, and CRP are associated with poor survival. IMPACT: Assessment of systemic inflammation via measurements of IL6, YKL-40, and CRP may be important for pancreatic cancer prognostication.