REVIEW Cyclooxygenase-2 Inhibitors in Tumorigenesis (Part I) Makoto M. Taketo In Part II of this review, I will summarize earlier data of The rate-limiting enzyme in arachidonate metabolism is me- NSAIDs on colorectal tumors and then present an overview of diated by enzymes known as cyclooxygenases (COXs). These research on COX-2 and its inhibitors relating to cancer. I will enzymes catalyze the biosynthesis of prostaglandin H2, the focus on their roles in colorectal cancer and its animal models, Downloaded from https://academic.oup.com/jnci/article/90/20/1529/2519830 by guest on 27 September 2021 precursor of molecules, such as prostaglandins, prostacyclin, with some extension to other types of cancer, and discuss their and thromboxanes. The COX enzyme family consists of the clinical relevance. classical COX-1 enzyme, which is constitutively expressed in To understand the effects of COX-2 on cancer, however, it many tissues, and a second enzyme, i.e., COX-2, which is would be essential for us to have a comprehensive knowledge of induced by various stimuli, such as mitogens and cytokines, the biochemistry and pharmacology of the COXs and their in- and is involved in many inflammatory reactions. Because hibitors. I am going to review these studies first in Part I. Be- nonsteroidal anti-inflammatory drugs inhibit both COX-1 cause I am unable to adequately review all of this literature in the and COX-2, these drugs also cause unwanted side effects, space available, I would urge the readers to refer to some ex- exemplified by gastrointestinal bleeding. Accumulating evi- cellent reviews that cover the related topics: Marnett (2), Smith dence indicates that nonsteroidal anti-inflammatory drugs and DeWitt (3,4), DuBois et al. (5), Smith et al. (6), and can reduce the incidence of colorectal cancers in human and Levy (7). This review covers literature available before August experimental animals and can reduce the polyp number and 1997. size in patients with familial adenomatous polyposis. This Part I (of a two-part review) focuses on the discovery of the ARACHIDONATE METABOLISM AND CYCLOOXYGENASES COXs; their biochemical, molecular, and structural proper- ties; and on the discovery of isozyme-specific inhibitors of The enzyme activity that catalyzes formation of prostaglandin COX activity. [J Natl Cancer Inst 1998;90:1529–36] G2 (PGG2) from arachidonic acid—followed by its conversion into PGH2 (Fig. 1)—was designated as COX, prostaglandin– endoperoxide synthase, or prostaglandin H synthase. There are CYCLOOXYGENASES AND CANCER two mammalian isozymes encoded by different genes: the con- stitutive COX-1 (the first one to be described, EC 1.14.99.1) and More than 20 years ago, high concentrations of prostagland- the inducible COX-2. Both enzymes are the major pharmaco- ins were found in human and animal tumor tissues. This rela- logic targets of NSAIDs. The enzyme we now call COX-1 was tionship of neoplastic tumors to increased levels of prostaglan- first purified and characterized from bull vesicular glands by dins stimulated various experimental and clinical research Miyamoto et al. in 1976 (8), followed by Hemler and Lands (9) thereafter. Accumulating evidence indicates that nonsteroidal from sheep, Van der Ouderaa et al. (10) from sheep, and Ogino anti-inflammatory drugs (NSAIDs) that inhibit prostaglandin (or et al. (11) from bull. In fact, two catalytic reactions take place on prostanoid1) synthesis can reduce the incidence of colorectal the same enzyme: a di-oxygenase activity that cyclizes and oxy- cancers. There are three lines of evidence, from chemical car- genates arachidonic acid to form a15-hydroperoxy-9, 11- cinogen-induced rodent models, from clinical trials of familial endoperoxide with a substituted cyclopentene ring (PGG ), and adenomatous polyposis patients, and from epidemiologic stud- 2 a peroxidase activity that reduces PGG to its 15-hydroxy me- ies. 2 tabolite (PGH ) (8,10,12–15). The approximately 3-kilobase The enzyme involved in the first step of prostanoid synthesis 2 (kb) sheep complementary DNA (cDNA) encoding COX-1 was from arachidonic acid is designated as cyclooxygenase (COX), cloned and analyzed from the seminal vesicles in 1988 (16–18), prostaglandin H synthase, or prostaglandin–endoperoxide syn- followed by cloning from human, mouse, and rat sources (19– thase2. Two isoenzymes exist in the mammalian body, consti- tutive COX-1 and inducible COX-2. While COX-1 is involved in the homeostasis of various physiologic functions, COX-2 is responsible for many inflammatory processes. After the discov- Affiliation of author: Laboratory of Biomedical Genetics, Graduate School of ery of COX-2 in the early 1990s, many effects of NSAIDs on Pharmaceutical Sciences, University of Tokyo, Japan. Correspondence to: Makoto M. Taketo, M.D., Ph.D., Laboratory of Bio- human colon cancer and in animal models of this disease were medical Genetics, Graduate School of Pharmaceutical Sciences, University of ascribed to these drugs’ effects on COX-2. However, direct ex- Tokyo, 7–3–1 Hongo, Bunkyo, Tokyo 113, Japan (e-mail: [email protected] perimental evidence of this relationship was missing. We re- tokyo.ac.jp). cently presented genetic and pharmacologic data to support the See ‘‘Notes’’ following ‘‘References.’’ hypothesis (1). © Oxford University Press Journal of the National Cancer Institute, Vol. 90, No. 20, October 21, 1998 REVIEW 1529 repeats (Shaw/Kamen sequences) that are known to mediate rapid and selective mRNA degradation (30,31,33,34). The TIS10 gene was expressed in COS-1 cells and was demonstrated to encode a func- tional COX containing both hydroperoxidase and COX catalytic activities (35). These data, taken to- gether, established the existence of a novel COX isozyme that was designated as mitogen-inducible prostaglandin G/H synthase (36), prostaglandin H synthase 2, or COX-2. Although the overall exon- intron organization of the mouse and human COX-2 genes is similar to that of human COX-1, the COX-2 genes contain 10 exons, respectively, in about 8 kb, instead of 11 exons in 22 kb for COX-1 (19,24). The relatively small genomic size for the COX-2 gene Downloaded from https://academic.oup.com/jnci/article/90/20/1529/2519830 by guest on 27 September 2021 fits one of the characteristics of the immediate-early genes (37). Such a difference in the gene structure is reflected in the structural and functional differences Fig. 1. Arachidonate metabolism pathways for prostanoid synthesis by cyclooxygenase 1 between the COX-1 and COX-2 proteins as well (COX-1) and cyclooxygenase 2 (COX-2). (see below). The human COX-2 cDNA was soon cloned from endothelial cells (38,39). Through hu- man–hamster somatic cell hybrid studies, the geno- 22). The amino acid sequences predicted from the nucleic acid mic genes encoding COX-1 and COX-2 were assigned to dif- sequences of the cDNAs are very similar among the species ferent human chromosomes, with the COX-2 gene on (about 90% identity). Sequence comparisons of mouse and chromosome 1 (21,40,41). More precise fluorescence in situ sheep COX-1 suggested that His309 of the sheep enzyme is the hybridization analysis demonstrated that the human genes for axial heme ligand (20), which was confirmed by site-directed COX-1 (PTGS1) and COX-2 (PTGS2) map to 9q32–q33.3 and mutagenesis experiments (23). The genomic sequences for the 1q25.2–q25.3, respectively (42), whereas linkage analyses in human and mouse COX-1 genes span about 22 kb and are com- interspecific backcross mice showed that their mouse homologs posed of 11 exons, respectively (19,24). Ptgs1 and Ptgs2 map to distinct loci on chromosome 2 and Although prostaglandin production and COX-1 messenger chromosome 1, respectively (43). RNA (mRNA) levels were studied extensively with the cDNA probes, most attempts failed to demonstrate a direct association REGULATION OF PROSTAGLANDIN SYNTHESIS (25,26). Rather, analyses using antibodies against COX-1 led to the detection of another cross-reacting band in rat ovary (27) and Before the discovery of COX-2, it was known that prosta- sheep tracheal epithelial cells (28). In 1989, Rosen et al. (29) glandin synthesis is stimulated by a variety of substances, in- found by northern analysis an mRNA band of 4.0 kb in sheep cluding growth factors and tumor promoters. These effects were tracheal mucosa cells that cross-hybridized with the sheep COX thought to be a result of the activation of phospholipases, which cDNA probe under a low stringency condition. The intensity of supply more arachidonic acid to COX (2). The fact that two this band showed a more direct association with prostaglandin isozymes exist that are regulated independently helped clear levels than did the 2.8-kb COX-1 band, and the researchers much of the earlier confusion about the control of prostaglandin suggested that this mRNA encoded a mitogen-inducible COX. production and revealed an elaborate regulatory mechanism for In 1991, cDNAs for COX-2, a novel isozyme of COX, were COX-2. Essentially, COX-1 is expressed constitutively and isolated and sequenced by two independent groups. In chick ubiquitously, whereas COX-2 is expressed only in response to embryo fibroblasts transformed with a temperature-sensitive certain stimuli (3). Rous sarcoma virus (RSV) mutant, Xie et al. (30) found a 4.1-kb mRNA among the immediate early genes that encoded a protein NSAIDS AND THE INHIBITION OF COXS containing about 60% amino acid identity with sheep COX-1. Independently, TIS10, one of seven primary-response genes rap- The early history of aspirin and its mechanism of action has idly and transiently induced by the tumor promoter TPA (12-O- been reviewed in some excellent reviews, especially one by tetradecanoylphobol-13-acetate) in Swiss mouse 3T3 cells, was Vane et al.