Novel Autoantibody Signatures in Sera of Patients with Pancreatic Cancer, Chronic Pancreatitis and Autoimmune Pancreatitis: A Protein Microarray Profiling Approach

Abstract

Identification of disease-associated autoantibodies is of high importance. Their assessment could complement current diagnostic modalities and assist the clinical management of patients. We aimed at developing and validating high-throughput protein microarrays able to screen patients’ sera to determine disease-specific autoantibody-signatures for pancreatic cancer (PDAC), chronic pancreatitis (CP), autoimmune pancreatitis and their subtypes (AIP-1 and AIP-2). In-house manufactured microarrays were used for autoantibody-profiling of IgG-enriched preoperative sera from PDAC-, CP-, AIP-1-, AIP-2-, other gastrointestinal disease (GID) patients and healthy controls. As a top-down strategy, three different fluorescence detection-based protein-microarrays were used: large with 6400, intermediate with 345, and small with 36 full-length human recombinant proteins. Large-scale analysis revealed 89 PDAC, 98 CP and 104 AIP immunogenic antigens. Narrowing the selection to 29 autoantigens using pooled sera first and individual sera afterwards allowed a discrimination of CP and AIP from PDAC. For validation, predictive models based on the identified antigens were generated which enabled discrimination between PDAC and AIP-1 or AIP-2 yielded high AUC values of 0.940 and 0.925, respectively. A new repertoire of autoantigens was identified and their assembly as a multiplex test will provide a fast and cost-effective tool for differential diagnosis of pancreatic diseases with high clinical relevance.

Use of Autoreactive Antibodies in Blood of Patients with Pancreatic Intraductal Papillary Mucinous Neoplasms (IPMN) for Grade Distinction and Detection of Malignancy

“…To select protein antigens of high interest, a PubMed literature search was performed. The proteins included in the study matched one or more of the following criteria: reported in previous IPMN-related antibody reports [ 22 ], pancreatic cancer-related antibody reports [ 18 , 20 , 23 , 24 , 25 , 26 , 27 ], or cancer autoantibody publications [ 14 , 28 , 29 , 30 ]. Additionally, some proteins of importance to pancreatic function, such as CCKBR, PNLIP, LDH, VEGF, LOX, G6PD, and GPX-4, were included.…”

“…Protein microarrays were produced and incubated as described in full detail elsewhere [ 20 , 21 , 32 , 33 ]. Briefly, the cDNAs were used for a PCR-based generation of protein expression templates.…”

“…The possible utilization of autoantibodies as biomarkers for cancer has already been studied for several tumor entities [ 16 , 17 , 18 ]. In this study, autoantibodies (AAbs) were detected and analyzed by protein microarray analyses [ 19 , 20 , 21 ], with the following objectives: (1) The identification of distinct antibody signatures for different pathological stages of IPMN; and (2) the assessment of the performance of AAbs signatures for the discrimination of low- and high-risk IPMN.…”

Liquid biopsy approach to pancreatic cancer

“…The low prevalence of PC means that biomarkers should be highly specific[ 164 ]. It is difficult to properly correlate AAbs with PC for diagnostic purposes because of the low immune response to this cancer, attributable to the protection offered by the stromal network and immunosuppressive mechanisms[ 159 - 161 ].…”

“…A further concern is the low capacity of AAbs to differentiate between PC and other inflammatory diseases, especially CP or autoimmune pancreatitis, which are responsible for a large production of AAbs. In this regard, some authors have proposed a panel of PP1R15A, CYP3A5, and WDR45 antigens to discriminate between PC and benign inflammatory disease[ 164 ].…”