Transcriptional profiles of peripheral blood leukocytes identify patients with cholangiocarcinoma and predict outcome.

Cholangiocarcinoma (CCA), a slow growing but highly metastatic tumor, is highly prevalent in Northeast Thailand. Specific tests that predict prognosis of CCA remain elusive. The present study was designed to investigate whether peripheral blood leukocyte (PBL) transcriptional profiles might be of use as a prognostic test in CCA patients. Gene expression profiles of PBLs from 9 CCA and 8 healthy subjects were conducted using the Affymetrix HG_U133 Plus 2.0 GeneChip. We indentified informative PBLs gene expression profiles that could reliably distinguish CCA patients from healthy subjects. Of these CCA specific genes, 117 genes were up regulated and 60 were down regulated. The molecular and cellular functions predicted for these CCA specific genes according to the Gene Ontology database indicated differential PBL expression of host immune response and tumor progression genes (EREG, TGF ß1, CXCL2, CXCL3, IL-8, and VEGFA). The expression levels of 9 differentially expressed genes were verified in 36 CCA vs 20 healthy subjects. A set of three tumor invasion related genes (PLAU, CTSL and SERPINB2) computed as "prognostic index" was found to be an independent and statistically significant predictor for CCA patient survival. The present study shows that CCA PBLs may serve as disease predictive clinically accessible surrogates for indentifying expressed genes reflective of CCA disease severity.

Exome sequencing of liver fluke-associated cholangiocarcinoma.

Opisthorchis viverrini-related cholangiocarcinoma (CCA), a fatal bile duct cancer, is a major public health concern in areas endemic for this parasite. We report here whole-exome sequencing of eight O. viverrini-related tumors and matched normal tissue. We identified and validated 206 somatic mutations in 187 genes using Sanger sequencing and selected 15 genes for mutation prevalence screening in an additional 46 individuals with CCA (cases). In addition to the known cancer-related genes TP53 (mutated in 44.4% of cases), KRAS (16.7%) and SMAD4 (16.7%), we identified somatic mutations in 10 newly implicated genes in 14.8-3.7% of cases. These included inactivating mutations in MLL3 (in 14.8% of cases), ROBO2 (9.3%), RNF43 (9.3%) and PEG3 (5.6%), and activating mutations in the GNAS oncogene (9.3%). These genes have functions that can be broadly grouped into three biological classes: (i) deactivation of histone modifiers, (ii) activation of G protein signaling and (iii) loss of genome stability. This study provides insight into the mutational landscape contributing to O. viverrini-related CCA.

Exome sequencing identifies frequent mutation of the SWI/SNF complex gene PBRM1 in renal carcinoma.

The genetics of renal cancer is dominated by inactivation of the VHL tumour suppressor gene in clear cell carcinoma (ccRCC), the commonest histological subtype. A recent large-scale screen of ∼3,500 genes by PCR-based exon re-sequencing identified several new cancer genes in ccRCC including UTX (also known as KDM6A), JARID1C (also known as KDM5C) and SETD2 (ref. 2). These genes encode enzymes that demethylate (UTX, JARID1C) or methylate (SETD2) key lysine residues of histone H3. Modification of the methylation state of these lysine residues of histone H3 regulates chromatin structure and is implicated in transcriptional control. However, together these mutations are present in fewer than 15% of ccRCC, suggesting the existence of additional, currently unidentified cancer genes. Here, we have sequenced the protein coding exome in a series of primary ccRCC and report the identification of the SWI/SNF chromatin remodelling complex gene PBRM1 (ref. 4) as a second major ccRCC cancer gene, with truncating mutations in 41% (92/227) of cases. These data further elucidate the somatic genetic architecture of ccRCC and emphasize the marked contribution of aberrant chromatin biology.

Tissue invasive macrophage density is correlated with prognosis in cholangiocarcinoma.

Cholangiocarcinoma (CCA) is a high metastatic cancer with no effective treatment. Here, the pro-metastatic action of tissue macrophages in CCA is demonstrated and suggested as a prognostic marker and novel target for the therapeutic intervention of CCA. Fifty CCA tissues were immunohistochemically stained with a marker for reactive/infiltrating monocytes/macrophages (MAC387) and matrix metalloproteinase (MMP)-9. The antigenic densities in positively-stained cells along the leading edge of tumors were scored. Correlations between the densities of MAC387, MMP-9-positive cells, clinicopathological features and patient survival were investigated. High densities of MAC387-positive cells were detected in more than 60% of the CCA tissues. This was significantly associated with poor prognosis parameters (non-papillary and mass-forming type CCA). Overall survival was worst in patients with high-density MAC387-positive cells. Double immunofluorescent staining indicated that MAC387-positive cells co-expressed MMP-9. Immunohistochemical staining of MMP-9 in serial sections of CCA tissues indicated that MMP-9 was rarely expressed in CCA tumor cells, but highly expressed in MAC387-positive cells and polymorphonucleated infiltrating cells. Patients with high tissue expression levels of MAC387 in combination with MMP-9-expressing cells had the worst survival. These factors were found to be independent predictors of the post-resectional survival of CCA patients. Since CCA tumor cells rarely expressed MMP-9, it is likely that tissue macrophages are critical for degrading the extracellular matrix and for facilitating tumor metastasis. They may therefore serve as a prognostic marker for poor clinical outcome, and represent novel targets for the therapeutic intervention of CCA.