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BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers, with around 9% of patients surviving >5 years. Asymptomatic in its initial stages, PDAC is mostly diagnosed late, when already a locally advanced or metastatic disease, as there are no useful biomarkers for detection in its early stages, when surgery can be curative. We have previously described a promising biomarker panel (LYVE1, REG1A, and TFF1) for earlier detection of PDAC in urine. Here, we aimed to establish the accuracy of an improved panel, including REG1B instead of REG1A, and an algorithm for data interpretation, the PancRISK score, in additional retrospectively collected urine specimens. We also assessed the complementarity of this panel with CA19-9 and explored the daily variation and stability of the biomarkers and their performance in common urinary tract cancers. METHODS AND FINDINGS: Clinical specimens were obtained from multiple centres: Barts Pancreas Tissue Bank, University College London, University of Liverpool, Spanish National Cancer Research Center, Cambridge University Hospital, and University of Belgrade. The biomarker panel was assayed on 590 urine specimens: 183 control samples, 208 benign hepatobiliary disease samples (of which 119 were chronic pancreatitis), and 199 PDAC samples (102 stage I-II and 97 stage III-IV); 50.7% were from female individuals. PDAC samples were collected from patients before treatment. The samples were assayed using commercially available ELISAs. Statistical analyses were performed using non-parametric Kruskal-Wallis tests adjusted for multiple comparisons, and multiple logistic regression. Training and validation datasets for controls and PDAC samples were obtained after random division of the whole available dataset in a 1:1 ratio. The substitution of REG1A with REG1B enhanced the performance of the panel to detect resectable PDAC. In a comparison of controls and PDAC stage I-II samples, the areas under the receiver operating characteristic curve (AUCs) increased from 0.900 (95% CI 0.843-0.957) and 0.926 (95% CI 0.843-1.000) in the training (50% of the dataset) and validation sets, respectively, to 0.936 in both the training (95% CI 0.903-0.969) and the validation (95% CI 0.888-0.984) datasets for the new panel including REG1B. This improved panel showed both sensitivity (SN) and specificity (SP) to be >85%. Plasma CA19-9 enhanced the performance of this panel in discriminating PDAC I-II patients from controls, with AUC = 0.992 (95% CI 0.983-1.000), SN = 0.963 (95% CI 0.913-1.000), and SP = 0.967 (95% CI 0.924-1.000). We demonstrate that the biomarkers do not show significant daily variation, and that they are stable for up to 5 days at room temperature. The main limitation of our study is the low number of stage I-IIA PDAC samples (n = 27) and lack of samples from individuals with hereditary predisposition to PDAC, for which specimens collected from control individuals were used as a proxy. CONCLUSIONS: We have successfully validated our urinary biomarker panel, which was improved by substituting REG1A with REG1B. At a pre-selected cutoff of >80% SN and SP for the affiliated PancRISK score, we demonstrate a clinically applicable risk stratification tool with a binary output for risk of developing PDAC ('elevated' or 'normal'). PancRISK provides a step towards precision surveillance for PDAC patients, which we will test in a prospective clinical study, UroPanc.

Background Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers, with around 9% of patients surviving >5 years. Asymptomatic in its initial stages, PDAC is mostly diagnosed late, when already a locally advanced or metastatic disease, as there are no useful biomarkers for detection in its early stages, when surgery can be curative. We have previously described a promising biomarker panel (LYVE1, REG1A, and TFF1) for earlier detection of PDAC in urine. Here, we aimed to establish the accuracy of an improved panel, including REG1B instead of REG1A, and an algorithm for data interpretation, the PancRISK score, in additional retrospectively collected urine specimens. We also assessed the complementarity of this panel with CA19-9 and explored the daily variation and stability of the biomarkers and their performance in common urinary tract cancers. Methods and findings Clinical specimens were obtained from multiple centres: Barts Pancreas Tissue Bank, University College London, University of Liverpool, Spanish National Cancer Research Center, Cambridge University Hospital, and University of Belgrade. The biomarker panel was assayed on 590 urine specimens: 183 control samples, 208 benign hepatobiliary disease samples (of which 119 were chronic pancreatitis), and 199 PDAC samples (102 stage I–II and 97 stage III–IV); 50.7% were from female individuals. PDAC samples were collected from patients before treatment. The samples were assayed using commercially available ELISAs. Statistical analyses were performed using non-parametric Kruskal–Wallis tests adjusted for multiple comparisons, and multiple logistic regression. Training and validation datasets for controls and PDAC samples were obtained after random division of the whole available dataset in a 1:1 ratio. The substitution of REG1A with REG1B enhanced the performance of the panel to detect resectable PDAC. In a comparison of controls and PDAC stage I–II samples, the areas under the receiver operating characteristic curve (AUCs) increased from 0.900 (95% CI 0.843–0.957) and 0.926 (95% CI 0.843–1.000) in the training (50% of the dataset) and validation sets, respectively, to 0.936 in both the training (95% CI 0.903–0.969) and the validation (95% CI 0.888–0.984) datasets for the new panel including REG1B. This improved panel showed both sensitivity (SN) and specificity (SP) to be >85%. Plasma CA19-9 enhanced the performance of this panel in discriminating PDAC I–II patients from controls, with AUC = 0.992 (95% CI 0.983–1.000), SN = 0.963 (95% CI 0.913–1.000), and SP = 0.967 (95% CI 0.924–1.000). We demonstrate that the biomarkers do not show significant daily variation, and that they are stable for up to 5 days at room temperature. The main limitation of our study is the low number of stage I–IIA PDAC samples (n = 27) and lack of samples from individuals with hereditary predisposition to PDAC, for which specimens collected from control individuals were used as a proxy. Conclusions We have successfully validated our urinary biomarker panel, which was improved by substituting REG1A with REG1B. At a pre-selected cutoff of >80% SN and SP for the affiliated PancRISK score, we demonstrate a clinically applicable risk stratification tool with a binary output for risk of developing PDAC (‘elevated’ or ‘normal’). PancRISK provides a step towards precision surveillance for PDAC patients, which we will test in a prospective clinical study, UroPanc., Silvana Debernardi and colleagues establish a clinical risk score and a biomarker panel for early detection of pancreatic cancer., Author summary Why was this study done? Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers, with around 9% of individuals surviving more than 5 years. The poor outcome is largely due to late diagnosis; however, if the disease is detected at an early stage, when tumours are still small and resectable, 5-year survival can increase to up to 70%. The development of a non-invasive test for the early detection of PDAC is therefore a major clinical need, likely to improve the chances of patients’ survival. What did the researchers do and find? We have previously identified a panel of 3 protein biomarkers (LYVE1, REG1A, and TFF1) in urine that showed promise in detecting resectable PDAC. In this study, we have improved this panel by substituting REG1A with REG1B, validated this panel in an additional number of retrospectively collected samples, and shown the ability of our urinary panel to distinguish control individuals and patients with benign hepatobiliary diseases from early stage PDAC patients with specificity and sensitivity >85%. Different cutoffs were tested for the PancRISK score, an algorithm that enables the stratification of patients into those with ‘normal’ or ‘elevated’ risk for developing PDAC: PancRISK-Fam can be useful for surveillance of asymptomatic individuals with genetic predisposition for PDAC, and PancRISK-Sym for symptomatic patients (such as those with chronic pancreatitis) who are at risk of developing PDAC. What do these findings mean? The urine biomarker panel and the affiliated PancRISK score were successfully validated in the large number of retrospectively collected samples and present a promising approach for completely non-invasive early detection of PDAC. After testing in a prospective cohort, this patient stratification approach could be implemented in clinical practice, with the potential to improve the current diagnostic pathway for PDAC patients.