No abstract

Nonsteroidal antiinflammatory drugs (NSAIDs) appear to reduce the risk of developing cancer. One mechanism through which NSAIDs act to reduce carcinogenesis is to inhibit the activity of cyclooxygenase-2 (COX-2), an enzyme that is overexpressed in various cancer tissues. Overexpression of COX-2 increases cell proliferation and inhibits apoptosis. However, selective COX-2 inhibitors can also act through COX-independent mechanisms. In this review, we describe the COX-2-independent molecular targets of these COX-2 inhibitors and discuss how these targets may be involved in the anticarcinogenic activities of these selective COX-2 inhibitors. We also compare the concentrations of these inhibitors used in in vitro and in vivo experiments and discuss the implications of the in vitro studies for clinical management of cancer with these drugs.

The regular use of various nonsteroidal anti-inflammatory drugs (NSAIDs) was shown to decrease the incidence of colorectal cancer. This effect is thought to be caused predominantly by inhibition of cyclooxygenase-2 (COX-2) and, subsequently, prostaglandin synthesis. However, recent studies have suggested that COX-independent pathways may contribute considerably to these antiproliferative effects. To evaluate the involvement of COX-dependent and COX-independent mechanisms further, we assessed the effects of celecoxib (selective COX-2 inhibitor) and SC560 (selective COX-1 inhibitor) on cell survival, cell cycle distribution, and apoptosis in three colon cancer cell lines, which differ in their expression of COX-2. Both drugs induced a G0/G1 phase block and reduced cell survival independent of whether or not the cells expressed COX-2. Celecoxib was more potent than SC560. The G0/G1 block caused by celecoxib could be attributed to a decreased expression of cyclin A, cyclin B1, and cyclin-dependent kinase-1 and an increased expression of the cell cycle inhibitory proteins p21Waf1 and p27Kip1. In addition, celecoxib, but not SC560, induced apoptosis, which was also independent of the COX-2 expression of the cells. In vivo, celecoxib as well as SC560 reduced the proliferation of HCT-15 (COX-2 deficient) colon cancer xenografts in nude mice, but both substances had no significant effect on HT-29 tumors, which express COX-2 constitutively. Thus, our in vitro and in vivo data indicate that the antitumor effects of celecoxib probably are mediated through COX-2 independent mechanisms and are not restricted to COX-2 over-expressing tumors.

Beside their structural role for the cell membrane the family of sphingolipids act as effector molecules in signal transduction with links to various aspects of cancer initiation, progression and treatment response. The "sphingolipid rheostat" balances between apoptosis inducing ceramid and growth promoting sphingosine-1-phosphate. We analyzed gene expression of 43 proteins from this pathway in different subtypes of breast cancer using microarray data of 1,269 tumor samples (test set n=171; validation sets n=1098) and observed significant differences for several genes. Sphingosine kinase 1 (SPHK1), ceramide galactosyltransferase (UGT8), and Ganglioside GD3-Synthase (ST8SIA1) displayed higher expression among ER negative tumors. In contrast, glucosylceramidsynthase (GCS), dihydroceramidsynthases (LASS4, LASS 6) and acid ceramidase (ASAH1) were higher expressed in ER positive samples. Survival analysis revealed a worse outcome of patients with high SPHK1 expression. To avoid a confounding effect of the ER status we also restricted the analysis to 750 patients with ER positive tumors. Again a worse outcome was observed for tumors displaying high SPHK1 expression. While 75.8+/-1.9% of the patients with tumors low in SPHK1 expression were free of metastasis at 5 years, this was the case for only 64.9+/-3.6% of patients with tumors displaying high SPHK1 expression (P=0.008). Immunohistochemistry identified the carcinoma cells as the major source of SPHK1 expression in the tumor. The correlation of SPHK1 with a poor prognosis as well as its high expression among ER negative tumors are in line with the antiapoptotic and proliferative properties of its product sphingosine-1-phosphate. Targeting of the sphingolipid rheostat may thus open new treatment options.

Several in vitro studies have correlated dysfunction of the sphingolipid-signaling pathway with promotion of tumor cell growth as well as progression and resistance of tumors to chemotherapeutic agents. As ceramides (Cer) constitute the structural backbones of all sphingolipids, we investigated the endogenous ceramide levels in 43 malignant breast tumors and 21 benign breast biopsies and compared them with those of normal tissues using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The total ceramide levels in malignant tumor tissue samples were statistically significantly elevated when compared with normal tissue samples. Upregulation of the total ceramide level averaged 12-fold and 4-fold higher than normal tissue samples, for malignant tumors and benign tissues, respectively. Specifically, the levels of C(16:0)-Cer, C(24:1)-Cer and C(24:0)-Cer were significantly raised in malignant tumors as compared with benign and normal tissue. The augmentation of the various ceramides could be assigned to an increase of the messenger RNA levels of ceramide synthases (CerS) LASS2 (longevity assurance), LASS4 and LASS6. Notably, elevated levels of C(16:0)-Cer were associated with a positive lymph node status, indicating a metastatic potential for this ceramide. Moreover, the levels of C(18:0)-Cer and C(20:0)-Cer were significantly higher in estrogen receptor (ER) positive tumor tissues as compared with ER negative tumor tissues. In conclusion, progression in breast cancer is associated with increased ceramide levels due to an upregulation of specific LASS genes.