br Keywords brush cytology ERCP microRNA pancreatobiliary br
Keywords: brush, cytology, ERCP, microRNA, pancreatobiliary
Projected cancer death of bile duct and pancreatic cancers, surpassing breast, prostate, and colorectal cancers, are expected to become second and third leading causes of cancer-related death by 2030 . Pancreatobiliary malignant strictures are hard to diagnose, manage, and often manifest in advanced unresectable stages. Early diagnosis and precise staging are hindered by the lack of specific symptoms, biomarkers and the shortcomings of cross-sectional imaging [2, 3].
ERCP-based brush 530141-72-1 is widely available, because it is a cheap, fast, relatively easy to perform and a safe procedure with a specificity approaching 100%. In a recent meta-analysis the pooled sensitivity and specificity of brushings for the diagnosis of malignant biliary strictures was 45% and 99% respectively, even in a combination with intraductal biopsies the sensitivity only reached 59.4% . The low sensitivities measured in several studies , combined with the availability of more recent diagnostic procedures such endoscopic ultrasound (EUS), have led to questions over the utility of ERCP brush cytology in diagnosing pancreatobiliary disease. ERCP is still the only methodology that can treat the jaundiced and yield a cytology diagnosis in a single procedure, however, some might even consider neglecting brush cytology during a therapeutic ERCP procedure due to the sensitivity problem. ERCP/brush cytology in jaundiced patients is a valuable technique in pancreatobiliary cancer diagnosis principally in the neoadjuvant and palliative setting, and as an adjunct procedure in indeterminate biliary strictures, but efforts should be made to increase the sensitivity of the procedure.
Some ancillary cytological methods have already been tested in clinical studies which identified additional patients with malignancy over routine cytology without additional false positives. These include mutational analysis (ie, KRAS, p16, p53) , immunohistochemistry (ie, mesothelin) , DNA ploidity tests . All of these tests have shown promise in accurately identifying malignant pancreatobiliary strictures, but with the exception of FISH, none of them are used in clinical practice at the present. The detection of polysomy by fluorescence in situ hybridization (FISH), has been shown to double sensitivity in the most comprehensive study to date , more recently the same group developed a set of FISH probes that separate pancreatobiliary malignancy with a sensitivity of 64.7% . The evaluation of brushings by both routine cytology and FISH is now common day practice in some cytopathology laboratories .
In recent years there has been a dramatic increase in the discovery of microRNAs (miRs) playing important roles in a variety of fundamental cellular processes and helping the early diagnosis of various diseases, mainly
cancers . MicroRNAs are disease specific, stable markers that can be detected quickly and reproducibly by PCR based methodology available more widely in regular cytopathology laboratories . Aims of the project were: (1) to prove that microRNAs can be detected and isolated from brush cytology samples, (2) to determine the expression of four tumor-associated microRNAs (miR-16, miR-21, miR-221 and miR-196a) on obtained cytology samples, (3) to give way to novel single or combined molecular markers in order to increase the sensitivity of brush cytology enough to impact clinical decision making.
PATIENTS/MATERIALS AND METHODS
Subjects and study design. Patients were prospectively enrolled at the Department of Interventional Gastroenterology, National Institute of Oncology, Budapest, Hungary. A total of 73 samples from Caucasian patients were included into this study. Samples were collected during n=57 ERCP procedures from pancreatobiliary strictures. Patients were categorized in the malignant group with (1) histological proof of malignancy (endoscopic or percutaneous biopsy, surgical exploration and sampling, autopsy), (2) clearly malignant clinical course over at least 12 months after sampling (evidence of progression such as large vessel involvement, appearance of malignant lymph nodes or new metastases on imaging), (3) pancreatobiliary tumor-related death during follow-up. Patients grouped under benign stricture had none of the mentioned features and were followed-up for 20 months to exclude progression or malignancy.
After exclusion of duplicates (repetitive sampling for suspected false negatives), parapapillary lesions, metastatic biliary strictures from non-pancreatobiliary primaries, and samples where metastatic origin of a pancreatic mass could not be ruled out, we ended up with n=35 samples (biliary malignant n=14, pancreatic malignant n=12, pancreatobiliary benign n=9). For a summary of patient characteristics we refer to Table 1.