Quantitative RT-PCR analysis of MMR genes on EUS-guided FNA samples from focal pancreatic lesions.
OBJECTIVE
Pancreatic cancer is characterized by a variety of molecular alterations. Mismatch excision repair (MMR) is a DNA repair system that eliminates mismatched base pairs and it plays an important role in the maintaining of genomic integrity. The aim of the study was to assess the role of several MMR genes in the diagnosis of pancreatic cancer in samples collected by EUS-FNA procedure.


METHODS
The prospective study included 44 consecutive patients with pancreatic cancer (n=24) and chronic pseudotumoral pancreatitis (n=20). EUS-FNA was performed in all the patients. Gene analysis was performed by extracting the mRNA and by determining the expression of DNA repair genes (MLH1, MLH3, MSH6) using a standard algorithm.


RESULTS
Total RNA was successfully isolated from all the EUS-FNA pancreatic samples. We analyzed ROC curves to assess the significance of determining the expression of analyzed genes in EUS-FNA samples, obtaining a cutoff value of 476621mRNA copies/mL for MSH6, and sensitivity and specificity of 75.0% and 100%, respectively. For MLH1 and MLH3, sensitivity and specificity were only satisfactory (64.65% and 76.11%, and 75.0% and 63.64%, respectively).


CONCLUSIONS
The quality and amount of cellular sampling using pancreatic EUS-FNA allow the extraction of sufficient quantities of RNA to perform qRT-PCR analysis. The use of MMR genes for the differentiation between pseudotumoral chronic pancreatitis and pancreatic cancer using a minimally invasive sampling technique could be a promising technique.
University of Medicine and Pharmacy, Bucharest, Romania.
Dan Ionuţ
DI
Gheonea
Marius Eugen
ME
Ciurea
Adrian
A
Săftoiu
Mihai
M
Ioana
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D000465:Algorithms; D044963:Biopsy, Fine-Needle; D002352:Carrier Proteins / Q000235:genetics; D053843:DNA Mismatch Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D003937:Diagnosis, Differential; D019160:Endosonography; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D009687:Nuclear Proteins / Q000235:genetics; D010190:Pancreatic Neoplasms / Q000000981:diagnostic imaging / Q000235:genetics* / Q000473:pathology; D010195:Pancreatitis / Q000000981:diagnostic imaging / Q000235:genetics* / Q000473:pathology; D011237:Predictive Value of Tests; D011379:Prognosis; D011446:Prospective Studies; D012333:RNA, Messenger / Q000032:analysis; D012372:ROC Curve; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012383:Romania; D012680:Sensitivity and Specificity; D018084:Ultrasonography, Interventional / Q000379:methods*
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C094443:G-T mismatch-binding protein; C501486:MLH1 protein, human; C403623:MLH3 protein, human; D009687:Nuclear Proteins; D012333:RNA, Messenger; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins