Growth-suppressive effect of non-steroidal anti-inflammatory drugs on 11 colon-cancer cell lines and fluorescence differential display of genes whose expression is influenced by sulindac.

In addition to an anti-inflammatory effect, sulindac, one of the non-steroidal anti-inflammatory drugs (NSAIDs), has been shown to have a protective effect against the incidence and mortality of colorectal cancer. However, the molecular basis of its anti-proliferative function remains unclear. To investigate its molecular mechanism, we exposed 11 colon-cancer cell lines to NSAIDs such as aspirin, sulindac and the sulfide and sulfone metabolites of sulindac. Sensitivity to these drugs was dose- and time-dependent but varied from one cell line to another. Among the cell lines examined, sulindac showed a moderate anti-proliferative effect on HT-29 colon cancer cells and caused morphological changes, including an increase of cells with abnormal DNA content. We used the mRNA fluorescence differential display method with these cells to identify molecules that might contribute, through altered expression, to cellular changes in response to NSAIDs. Sixty-eight cDNA fragments were confirmed by RT-PCR to have significantly different expression levels following sulindac treatment. Thirty of these fragments proved to be novel cDNA sequences or identical to expressed sequence tags; the other 38 fragments were identical, or showed significant homology, to genes whose function was already known. Among the known genes differentially expressed in HT-29 cells after sulindac treatment were those encoding acetylglucosaminyltransferase, ferritin heavy chain, zinc finger protein 165, aldose reductase, carcinoembryonic antigen, aldoketoreductase, NF-kappaB-activating kinase, lysosome-associated protein, RhoE = 26 kDa GTPase homologue, NADH oxidoreductase, G/T mismatch bindingprotein, TM7SF3, ADP/ATP carrier-like protein and chromosome segregation protein. This variety among classes of proteins affected by sulindac in our experiments underscores the complexity of anti-proliferative mechanisms that may operate in colon-cancer cells treated with NSAIDs. Furthermore, identification of genes regulated by NSAIDs in colon-cancer cells should provide useful information to identify novel therapeutic targets for treatment and/or prevention of colon cancer.

Isolation and characterization of a human cDNA encoding a protein homologous to the 7.2-kDa protein (subunit X) of bovine ubiquinol-cytochrome C reductase.

Through large-scale sequencing of clones randomly selected from a library of human cDNAs, we have isolated a novel human gene termed hUQCR10. Its open reading frame encodes 63 amino acids that share 88.5% identity with the sequence of bovine ubiquinol-cytochrome C reductase 7.2-kDa protein (subunit X). A single 0.6-kb transcript was expressed in all human tissues examined, but was particularly abundant in heart and skeletal muscle, tissues that consume a large amount of oxygen. The gene product therefore may play a significant role in the cellular respiratory system. In support of this hypothesis, our immunohistochemical analysis revealed that the hUQCR10 protein is located in mitochondria. A homology search using computer programs determined the chromosomal localization of the gene at 22q12.