The acquisition of genomic instability is a crucial feature in tumor development and there are at least 3 distinct pathways in colorectal cancer pathogenesis: the chromosomal instability (CIN), microsatellite instability (MSI), and CpG island methylator phenotype (CIMP) pathways. Most cases of colorectal cancer arise through the CIN pathway, which is characterized by widespread imbalances in chromosome number (aneuploidy) and loss of heterozygosity (LOH). It can result from defects in chromosomal segregation, telomere stability, and the DNA damage response, although the full complement of genes underlying CIN remains incompletely described. Coupled with the karyotypic abnormalities observed in CIN tumors are the accumulation of a characteristic set of mutations in specific tumor suppressor genes and oncogenes that activate pathways critical for colorectal cancer initiation and progression. Whether CIN creates the appropriate milieu for the accumulation of these mutations or vice versa remains a provocative and unanswered question. The goal of this review is to provide an updated perspective on the mechanisms that lead to CIN and the key mutations that are acquired in this pathway.

Bortezomib (PS-341, Velcade) is a potent and selective inhibitor of the proteasome that is currently under investigation for the treatment of solid malignancies. We have shown previously that bortezomib has activity in pancreatic cancer models and that the drug induces endoplasmic reticulum (ER) stress but also suppresses the unfolded protein response (UPR). Because the UPR is an important cytoprotective mechanism, we hypothesized that bortezomib would sensitize pancreatic cancer cells to ER stress-mediated apoptosis. Here, we show that bortezomib promotes apoptosis triggered by classic ER stress inducers (tunicamycin and thapsigargin) via a c-Jun NH 2 -terminal kinase (JNK)-dependent mechanism. We also show that cisplatin stimulates ER stress and interacts with bortezomib to increase ER dilation, intracellular Ca 2+ levels, and cell death. Importantly, combined therapy with bortezomib plus cisplatin induced JNK activation and apoptosis in orthotopic pancreatic tumors resulting in a reduction in tumor burden. Taken together, our data establish that bortezomib sensitizes pancreatic cancer cells to ER stress-induced apoptosis and show that bortezomib strongly enhances the anticancer activity of cisplatin. (Cancer Res 2005; 65(24): 11658-66)

The proteasome inhibitor bortezomib ( formerly known as PS-341) recently received Food and Drug Administration approval for the treatment of multiple myeloma, and its activity is currently being evaluated in solid tumors. Bortezomib triggers apoptosis in pancreatic cancer cells, but the mechanisms involved have not been fully elucidated. Here, we show that pancreatic cancer cells exposed to bortezomib formed aggregates of ubiquitin-conjugated proteins (''aggresomes'') in vitro and in vivo. Bortezomib-induced aggresome formation was determined to be cytoprotective and could be disrupted using histone deacetylase (HDAC) 6 small interfering RNA or chemical HDAC inhibitors, which resulted in endoplasmic reticulum stress and synergistic levels of apoptosis in vitro and in an orthotopic pancreatic cancer xenograft model in vivo. Interestingly, bortezomib did not induce aggresome formation in immortalized normal human pancreatic epithelial cells in vitro or in murine pancreatic epithelial cells in vivo. In addition, these cells did not undergo apoptosis following treatment with bortezomib, suberoylanilide hydroxamic acid, or the combination, showing tumor selectivity. Taken together, our study shows that inhibition of aggresome formation can strongly potentiate the efficacy of bortezomib and provides the foundation for clinical trials of bortezomib in combination with HDAC inhibitors for the treatment of pancreatic cancer.

Systematic EPC in advanced pancreatic cancer patients significantly improved QoL with respect to on-demand EPC.

No abstract

Although it displays promising activity in other tumor models, the effects of tumor necrosis factor -related apoptosis-inducing ligand (TRAIL) on human pancreatic cancer cells have not been comprehensively explored. We report that a majority of human pancreatic cancer cell lines (seven of nine) underwent apoptosis when they were exposed to recombinant human TRAIL in vitro. Characterization of surface TRAIL receptors by fluorescenceactivated cell sorting showed that TRAIL-resistant cells (Panc-1 and HS766T) expressed lower levels of DR4 and DR5 than did TRAIL-sensitive cells. The proteasome inhibitor bortezomib (PS-341, Velcade) further increased TRAIL responsiveness in the TRAIL-sensitive cells and synergized with TRAIL to reverse resistance in Panc-1 and HS776T cells. The effects of bortezomib were mimicked by transfection with a small interfering RNA construct specific for the p65 subunit of nuclear factor-KB (NF-KB) or exposure to a selective chemical inhibitor of IKK (PS-1145). Silencing IKBA prevented TRAIL sensitization by PS-1145, confirming that IKBA mediated the effects of PS-1145. NF-KB inhibition resulted in down-regulation of BCL-X L and XIAP, and silencing either restored TRAIL sensitivity in TRAIL-resistant cells. Finally, therapy with TRAIL plus PS-1145 reversed TRAIL resistance in vivo to produce synergistic growth inhibition in orthotopic Panc-1 tumors. Together, our results show that NF-KB inhibits TRAIL-induced apoptosis in human pancreatic cancer cells and suggest that combination therapy with TRAIL and NF-KB inhibitors, such as bortezomib, PS-1145, or curcumin, should be considered as a possible treatment strategy in patients with pancreatic cancer. [Mol Cancer Ther 2006;5(9):2251 -60]

Background & Aims Colorectal cancers (CRCs) displaying DNA microsatellite instability (MSI) are associated with a favorable natural history, but the molecular basis for this observation has not been defined. We sought to determine whether the epithelial to mesenchymal transition (EMT) is impaired in MSI-positive CRCs that characteristically have a mutant transforming growth factor-β receptor type II (TGFBR2) gene. Methods The induction of EMT by TGF-β1 was analyzed by phase contrast microscopy, immunofluorescence, qRT-PCR, immunoblotting, and cellular migration and invasion assays. Expression of EMT markers was evaluated by immunohistochemistry and qRT-PCR in a series of human colorectal tumors. Results TGF-β1 induced changes in cellular morphology, gene expression, motility, and invasion consistent with EMT in microsatellite stable (MSS) colon cancer cells whereas cells with MSI and mutant TGFBR2 were unresponsive to TGF-β1. These effects did not require Smad4 but depended upon the recruitment of ERK. Tumor cells with MSI but wildtype TGFBR2 underwent EMT in response to TGF-β1, indicating that TGFBR2 genotype is a key determinant of the EMT response in tumors with MSI. In human colorectal tumors, expression of EMT markers was significantly associated with adverse clinicopathologic features and the absence of MSI. Conclusions These findings define a unique genotype-phenotype relationship between TGFBR2 and EMT that may contribute to the improved prognosis consistently observed in colon cancers with MSI.

KRAS and BRAF mutations are frequently observed in human colon cancers. These mutations occur in a mutually exclusive manner, and each is associated with distinctive biological features. We showed previously that K-ras can interact with hypoxia to activate multiple signaling pathways. Many hypoxic responses are mediated by hypoxia-inducible factor (HIF)-1α and HIF-2α, and we sought to define the roles of mutant KRAS and BRAF in the induction of HIF-1α and HIF-2α in colon cancer cells. Ectopic expression of mutant K-ras in Caco2 cells enhanced the hypoxic induction of only HIF-1α, whereas mutant BRAF enhanced both HIF-1α and HIF-2α. Knockout or knockdown of mutant KRAS in DLD-1 and HCT116 cells impaired the hypoxic induction of only HIF-1α. HIF-1α mRNA levels were comparable in cells with and without a KRAS mutation. However, the rate of HIF-1α protein synthesis was higher in cells with a KRAS mutation, and this was suppressed by the phosphoinositide 3-kinase inhibitor LY294002. In contrast, knockdown of mutant BRAF in HT29 cells suppressed both HIF-1α and HIF-2α. Although BRAF regulated mRNA levels of both HIF-1α and HIF-2α, knockdown of BRAF or treatment with the MEK inhibitor PD98059 impaired the translation of only HIF-2α. Our data reveal that oncogenic KRAS and BRAF mutations differentially regulate the hypoxic induction of HIF-1α and HIF-2α in colon cancer, and this may potentially contribute to the phenotypic differences of KRAS and BRAF mutations in colon tumors. [Cancer Res 2009;69(21):8499-506]