Identification of genetic alterations in pancreatic cancer by the combined use of tissue microdissection and array-based comparative genomic hybridisation

Abstract

Pancreatic ductal adenocarcinoma (PDAC) is characterised pathologically by a marked desmoplastic stromal reaction that significantly reduces the sensitivity and specificity of cytogenetic analysis. To identify genetic alterations that reflect the characteristics of the tumour in vivo, we screened a total of 23 microdissected PDAC tissue samples using array-based comparative genomic hybridisation (array CGH) with 1 Mb resolution. Highly stringent statistical analysis enabled us to define the regions of nonrandom genomic changes. We detected a total of 41 contiguous regions (43.0 Mb) of copy number changes, such as a genetic gain at 7p22.2 -p15.1 (26.0 Mb) and losses at 17p13.3 -p11.2 (13.6 Mb), 18q21.2 -q22.1 (12.0 Mb), 18q22.3 -q23 (7.1 Mb) and 18q12.3 -q21.2 (6.9 Mb). To validate our array CGH results, fluorescence in situ hybridisation was performed using four probes from those regions, showing that these genetic alterations were observed in 37 -68% of a separate sample set of 19 PDAC cases. In particular, deletion of the SEC11L3 gene (18q21.32) was detected at a very high frequency (13 out of 19 cases; 68%) and in situ RNA hybridisation for this gene demonstrated a significant correlation between deletion and expression levels. It was further confirmed by reverse transcription -PCR that SEC11L3 mRNA was downregulated in 16 out of 16 PDAC tissues (100%). In conclusion, the combination of tissue microdissection and array CGH provided a valid data set that represents in vivo genetic changes in PDAC. Our results raise the possibility that the SEC11L3 gene may play a role as a tumour suppressor in this disease.

Yes-associated protein (YAP) functions as a tumor suppressor in breast

“…The staining was done on the Ventana Discovery system (USA) as previously described. 33 ERa (Novocastra, 1 : 100) and Her-2 (CB11; Dako, 1 : 100) were demonstrated using pressure-cooking antigen retrieval followed by standard Avidin-Biotin Complex IHC. Images were acquired using a Zeiss Axiophot microscope ( Â 40) and Nikon DXM1200 digital camera.…”

Differential Diagnosis of Pancreatic Tumors by Molecular Analysis of Clinical Specimens

“…Among the hallmark features of PDAC are mutations in the K-ras and HER2/neu oncogenes as well as the p53 , p16INK4a and SMAD4/DPC4 tumor suppressor genes [for an overview, see 6,25 ]. More recently, high-throughput screening analyses have identified a multitude of differences between malignant and benign processes in the pancreas, as well as between different malignant tumor entities, on the genome [26][27][28][29] , transcriptome [30][31][32][33][34][35][36][37][38][39][40][41][42][43] , proteome [44][45][46][47][48] and epigenome [49][50][51][52] levels. Based on these results, a growing number of studies report on the use of one or several of these molecular markers in diagnostic analysis of serum, pancreatic juice, brush cytologies or FNABs.…”

Genome-Wide Analysis of Pancreatic Cancer Using Microarray-Based Techniques

“…The extremely high mortality rate of patients with PDAC is clinically considered to be due to late diagnosis and lack of effective therapeutic strategies, but biologically its aggressiveness could be explained by the high level of genomic instability in PDAC cells [2][3][4][5][6] . In general, genomic aberrations are selected for the induction of tumour growth advantage and accumulated according to tumour progression [7][8][9][10] .…”