Cancer Letters

Cancer Letters

Volume 122, Issues 1–2, 9 January 1998, Pages 25-30
Cancer Letters

Cyclooxygenase-2-selective antagonists do not inhibit growth of colorectal carcinoma cell lines

https://doi.org/10.1016/S0304-3835(97)00361-3Get rights and content

Abstract

Non-steroidal anti-inflammatory drugs (NSAIDs) reduce the incidence of colorectal carcinoma. We now report that the potent cyclooxygenase-1 inhibitor indomethacin had no effect on the growth of human colorectal carcinoma cell lines in vitro at concentrations up to 30 μM. The selective cyclooxygenase-2 inhibitors L-745337 and NS-398 reduced cyclooxygenase activity, but had no effect on cell growth at concentrations as high as 100 μM. Our results provide direct evidence that inhibition of cyclooxygenase activity does not necessarily inhibit the growth of colorectal carcinoma cell lines and imply that the growth-inhibitory effects of NSAIDs in vitro are not mediated by inhibition of cyclooxygenases.

Introduction

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit colorectal tumour growth in several experimental models. Indomethacin and other NSAIDs inhibited the development of chemically-induced colon cancers in rats and mice 1, 2. Sulindac reduced the size and number of colorectal polyps in patients with familial adenomatous polyposis (FAP) [3]and inhibited tumour formation in a murine model of FAP [4]. In addition, several epidemiological studies have revealed that NSAIDs and in particular aspirin reduced the risk of colorectal carcinoma by approximately 50% 5, 6. Finally, several NSAIDs have been shown to reversibly inhibit the growth of colorectal carcinoma cell lines in vitro 7, 8.

The effect of NSAIDs on colon cancer has generally been accepted to be the result of inhibition of cyclooxygenases [9]. The two forms of cyclooxygenase, which are key enzymes in prostaglandin biosynthesis [9], differ in inducibility, tissue distribution and susceptibility to NSAIDs [10]. Levels of mRNA for the inducible cyclooxygenase-2 [11]and of cyclooxygenase-1 and -2 immunoreactivity [12]were significantly higher in human colorectal tumour samples than in the accompanying normal mucosa. In addition, treatment of rats with a selective cyclooxygenase-2 inhibitor significantly reduced the induction of aberrant crypt foci by the carcinogen azoxymethane [13].

Several indirect lines of evidence have recently suggested that cyclooxygenases may not be the only target of the anti-proliferative effects of NSAIDs. Firstly, treatment of colorectal carcinoma cell lines with prostaglandins did not reverse the inhibitory effects of NSAIDs on cell growth [14]. Secondly, sulindac sulphoxide, which does not inhibit prostaglandin synthesis, has been shown to inhibit the formation of aberrant crypt foci in carcinogen-treated rats [15]and the growth of human colorectal carcinoma cell lines in vitro [7]. Thirdly, sulindac sulphoxide and several other NSAIDs reduced the amounts and activities of the cyclin-dependent kinases, with a concomitant increase in the proportion of cells in the G0/G1 phase 8, 16. Fourthly, NSAIDs have been shown to induce apoptosis in the rectal epithelium of patients with FAP [17]and in the human colon carcinoma cell line HT 29 8, 16, 18. To investigate the role of cyclooxygenases in the anti-proliferative effects of NSAIDs on colon cancer, we studied the expression of cyclooxygenases and the concentration dependence of growth inhibition by NSAIDs and cyclooxygenase-2-selective antagonists in two colorectal cancer cell lines. Our results suggest that cyclooxygenases are not the target for the inhibitory effects of NSAIDs on colorectal cancer cell growth in vitro.

Section snippets

Materials

The human colorectal carcinoma cell lines LIM 1215 [19]and LIM 1899 [20]were obtained from Dr R.H. Whitehead, Ludwig Institute for Cancer Research, Melbourne. NSAIDs were from Sigma (St. Louis, MO). The cyclooxygenase-2-selective inhibitors NS-398 [21]and L-745 337 [22]were generously provided by Taisho (Tokyo, Japan) and Merck, Frosst (Pointe-Claire-Dorval, Canada), respectively, and sulindac sulphide and sulphone were from Merck, Sharp and Dohme (West Point, PA).

Cyclooxygenase assay

Total cyclooxygenase activity

Results

The colorectal carcinoma cell lines LIM 1215 and LIM 1899 were screened for the expression of cyclooxygenase-1 and -2 mRNA by RT-PCR. Levels of cyclooxygenase-2 mRNA in LIM 1215 cells were similar to levels in human fibroblasts (Fig. 1). Levels of cyclooxygenase-2 mRNA in LIM 1899 cells and of cyclooxygenase-1 mRNA in both LIM 1215 and LIM 1899 cells were considerably lower than levels in human fibroblasts (Fig. 1). Levels of cyclooxygenase activity in either LIM 1215 or LIM 1899 cells measured

Discussion

Herein we present evidence that inhibition of cyclooxygenases does not necessarily inhibit the growth of the colorectal carcinoma cell lines LIM 1215 and LIM 1899. Firstly, the potent cyclooxygenase-1 inhibitor indomethacin (IC50 30 nM) was only a moderately effective inhibitor of colorectal carcinoma cell growth (IC50 300 μM). Secondly, the potent and selective cyclooxygenase-2 inhibitors NS-398 and L-745337 (IC50 values of 3.8 μM and 23 nM for cyclooxygenase-2, respectively) had no effect on

Acknowledgements

We thank Dr R.H. Whitehead for the colorectal carcinoma cell lines and for helpful discussions. This work was supported in part by grants no. 920527 and 960182 from the Australian NHMRC.

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