[CANCER RESEARCH 59, 5927–5931, December 1, 1999]

Interferon-inducible Gene Family 1-8U Expression in Colitis-associated ColonCancer and Severely Inflamed Mucosa in Ulcerative Colitis1

Tadakazu Hisamatsu, Mamoru Watanabe, Haruhiko Ogata, Toshihiko Ezaki, Shigenari Hozawa, Hiromasa Ishii,Takanori Kanai, and Toshifumi Hibi 2

Keio Cancer Center [M. W., T. K., T. Hib.], and Department of Internal Medicine, School of Medicine, Keio University [T. His., H. O., T. E., S. H., H. I., T. Hib.], Tokyo 160-8582, Japan

ABSTRACT

Persistent severe inflammation in colonic mucosa is thought to causethe development of colon cancer in patients with ulcerative colitis (UC).However, predisposing genetic abnormalities have not been identified inthis sequence. Using differential display PCR, we isolated cDNA frag-ments corresponding to mRNAs that were differentially expressed incolitis-associated cancer tissues and mucosa with mild inflammation in thecolon of five UC patients. This molecular screening approach identified 60cDNA fragments, and we sequenced 34 fragments. One cDNA fragment,which is identical to IFN-inducible gene family 1-8U, was strongly ex-pressed in all five UC-associated cancers. 1-8U was also expressed insporadic colon cancer tissues and colon cancer cell lines, but not in normalmucosa. This gene was strongly expressed in severely inflamed colonicmucosa of UC without colitis-associated colon cancer, although 1-8Uexpression was not related to the extent and duration of the disease.However, 1-8U was expressed in the colonic mucosa of all patients withchronic, continuously severe inflammation. These results indicated thatIFN-inducible gene family 1-8U expression in inflamed colonic mucosamight be used as a preferential marker of colitis-associated colon cancerin UC.

INTRODUCTION

The risk for colorectal carcinoma increases considerably in patientswith chronic UC3 after 10 years. Dysplasia in UC mucosa is thoughtto be a precursor of carcinoma and an indication for colectomy (1–3).Molecular mechanisms underlying UC-associated neoplasms havebeen studied for years, but understanding of these mechanisms re-mains incomplete. It is likely that an accumulation of multiple genemutations leads to dysplasia and cancer in UC in a manner similar tothat of sporadic colon malignancy. However, biological features differbetween UC-associated cancer and sporadic colorectal cancer. Muta-tions in the APC gene and ras proto-oncogenes are not consistentlypresent in UC-associated cancer, in contrast to sporadic colorectalcancer resulting from the adenoma-carcinoma sequence pathway (4–8). Conversely, p53 mutation is a frequent occurrence in early dys-plastic UC lesions, whereas p53 mutation is rare in early dysplasticadenomas (9, 10). Moreover, the time interval between UC-associateddysplasia and cancer is short compared to that between sporadicadenoma and carcinoma (8). Patients with dysplasia or cancer in UCare younger than those with sporadic colorectal cancer. These resultsindicate that destroyed molecular mechanisms might underlie UC-associated cancer, although conclusive evidence is limited.

Findings in patients with chronic UC are paralleled in animalmodels, the cotton top tamarin, a new world monkey that spontane-ously develops an idiopathic colitis with histopathological similarityin human UC. In this model, adenocarcinoma develops frequently(50%) in association with chronic colitis (11–14). HLA-B27 trans-genic rats (15), Gai2-deficient mice (16), and dominant negativeN-cadherin mice (17) that express chronic colitis also develop asso-ciated colonic adenoma or adenocarcinoma. In both UC patients andanimal models, persistent severe inflammation in the colonic mucosais thought to cause the development of colorectal cancer. However,molecular biological mechanisms of carcinogenesis due to colonicinflammation have not yet been identified.

The DD-PCR using arbitrary primers may be used to identifychanges in gene expression (18). Candidate oncogenes and tumorsuppresser genes have been isolated using this method (19, 20). UsingDD-PCR, we isolated cDNA fragments corresponding to mRNAs thatare differentially expressed in mucosa with UC-associated cancer andmild inflammation, and we sequenced those fragments. In the presentstudy, we show evidence that the IFN-inducible gene family 1-8U isexpressed not only in UC-associated cancer tissues, but also in se-verely inflamed colonic mucosa of UC. Our results indicated thatIFN-inducible gene family 1-8U expression in inflamed colonic mu-cosa may be used as a preferential marker of colitis-associated coloncancer in UC.

MATERIALS AND METHODS

Tissue Samples.Tissue samples were obtained at surgery from five pa-tients with UC who developed colonic adenocarcinoma. Clinical features ofUC in patients with UC-associated colon cancer are shown in Table 1.Inflamed colonic mucosa was obtained from 14 UC patients without coloncancer at surgery and endoscopically. We defined the quantification of inflam-mation in UC mucosa according to the widely accepted histopathologicalclassification by Matts (21). Based on Matts’s classification, we defined mildinflammation as inflammation, grade 3, and severe inflammation was de-fined as inflammation5 grade 4–5. The clinical type of UC patients included4 chronic continuous and 10 relapsing-remitting type. Eight patients were inactive stage, and six were in remission. Duration of the disease was less than5 years in seven patients and more than 5 years in seven patients. Sporadiccolon cancer mucosa was obtained from 10 patients with colon cancer atsurgery. Histopathological examination showed that no severe inflammationwas observed in the mucosa surrounding the cancer. Normal colonic mucosaobtained at colonoscopy from 10 healthy volunteers served as a control.Tissues were frozen in liquid nitrogen and stored at280°C until use.

mRNA DD-PCR. Total cellular RNA was isolated by RNazol B (Tel-Test,Inc., Friendswood, TX), and 0.2mg of the respective DNA-free RNAs wasreverse transcribed with Moloney murine leukemia virus reverse transcriptaseusing three different pools of oligodeoxythymidylic acid-anchored 39 primers(RNA image; GenHunter Co., Brookline, MA). The RT reaction was amplifiedin the presence of [32P]dATP in a PCR protocol. PCR reactions were carriedout with 2 mM HT11M primer, 2mM arbitrary primer (see below), 25mM

dNTP, 103reaction buffer, 0.3 unit of Taq DNA polymerase (Perkin-ElmerCorp., Foster City, CA). Reactions were performed at 94°C for 60 s and thenat 94°C for 30 s, 40°C for 120 s, and 72°C for 30 s for a total of 40 cycles. PCRproducts were displayed on standard sequence gels. Bands representingpotentially differentially expressed mRNAs were excised from the gel and

Received 6/18/99; accepted 10/6/99.The costs of publication of this article were defrayed in part by the payment of page

charges. This article must therefore be hereby markedadvertisementin accordance with18 U.S.C. Section 1734 solely to indicate this fact.

1 Supported in part by grants in aid from the Japanese Ministry of Education, Cultureand Science; the Japanese Ministry of Health and Welfare; Chiyoda Mutual Life Foun-dation; Keio University; and the Keio University Medical Fund.

2 To whom requests for reprints should be addressed, at Keio Cancer Center, 35Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. Phone/Fax: 011-81-3-3357-6156.E-mail: thibi@med.keio.ac.jp.

3 The abbreviations used are: UC, ulcerative colitis; DD, differential display; LPL,lamina propria lymphocyte; RT, reverse transcription; dNTP, deoxynucleotide triphos-phate; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; CMF-HBSS, calcium- andmagnesium-free HBSS; mAb, monoclonal antibody.

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eluted in distilled water. cDNA PCR products were then amplified usingthe same primers. The following 39 and 59 primers were used for RT andDD: (a) HT11A, 59-AAGCTTTTTTTTTTTA-39; (b) HT11C, 59-AAGCTTTTTTTTTTTC-39; and (c) HT11G, 59-AAGCTTTTTTTTTTTG-39.Randomly selected 59 primers for DD were as follows: (a) AP-9, 59-AAGCT-TCATTCCG-39; (b) AP-10, 59-AAGCTTCCACGTA-39; (c) AP-11, 59-AAGCTTCGGGTAA-3; (d) AP-12, 59-AAGCTTGAGTGCT-39; (e) AP-13,59-AAGCTTCGGCATA-39; (f) AP-14, 59-AAGCTTGGAGCTT-39; (g) AP-15, 59-AAGCTTACGCAAC-39; and (h) AP-16, 59-AAGCTTTAGAGCG-39.

Northern Blot Analysis. Tenmg of the indicated total RNA extracted fromtissue samples were electrophoresed on a 1.2% formaldehyde agarose gel andtransferred to a positively charged nylon membrane (Hybond N1; Amersham,Buckinghamshire, United Kingdom). Equivalent sample loading was con-firmed by ethidium bromide staining and visualization of 28S rRNA bands andby the mRNA levels of GAPDH. Labeling of DNA probes by [a-32P]dCTPwas carried out using DNA Labeling Beads (Ready To Go; Amersham), andpositively charged nylon membranes were hybridized with radiolabeled probein Rapid-Hyb Buffer (Amersham).

Subcloning, DNA Sequencing, and Homology Search of cDNA Frag-ments. The cDNA fragments were reamplified and subcloned into the TAcloning vector (Invitrogen, Carlsbad, CA). These fragments were sequencedwith the ABI PRISM Dye Terminator Cycle Sequencing Ready Reaction Kit(Perkin-Elmer Corp.) using221M13 primers. All sequences of cDNA frag-ments were compared to the National Center for Biotechnology InformationGenBank database using the BLAST algorithm.

Cell Lines. The human colon carcinoma cell lines HT29, Caco-2, andWiDr were obtained from the American Type Culture Collection (Rockville,MD), and ACM and CW2 were kindly provided by Dr. M. Mitsuhashi (TokyoWomen’s Medical College, Tokyo, Japan). Human cervical cancer cell lineHeLa was kindly provided by Dr. K. Toda (Kitasato Institute Hospital, Tokyo,Japan).

RT-PCR of IFN-inducible Gene Family 1-8U. Total RNA was isolatedfrom endscopically obtained specimens and by surgical resections usingRNazol (Cinna/Biotech Laboratory, Houston, TX). Twomg of total RNA wereincubated at 70°C for 10 min and reverse transcribed in the presence ofoligodeoxythymidylic acid primer, 103 reaction buffer, 10 mM dNTP, 0.1M

DTT, and Superscript reverse transcriptase (Life Technologies, Inc., Rockville,MD). The RT reaction was performed at 42°C for 50 min, 90°C for 5 min, and37°C for 20 min after the addition of RNase H (Life Technologies, Inc.). PCRreactions were carried out with an equal amount of cDNA, 2mM DD40 39and59primers (see below), 25mM dNTP, 103reaction buffer, and 0.3 unit of TaqDNA polymerase (Perkin-Elmer Corp.). Reactions were performed at 94°C for60 s and then at 94°C for 30 s, 40°C for 120 s, and 72°C for 30 s for a totalof 30 cycles. PCR products were displayed on 1.6% agarose gels and stainedwith ethidium bromide. GAPDH was used as an internal control. The follow-ing 39 and 59primers of DD40 (predicted size of the product was 97 bp) andGAPDH (predicted size of the product was 440 bp) were used for PCRs: (a)39primer of DD40, 59-GTGCACTTTATTGAATGCC-39; (b) 59 primer ofDD40, 59-CTCCAACTTCCATTCCTCG-39; (c) 39 primer of GAPDH, 59-CGACGCCTGCTTCACCACCT-39; and (d) 59 primer of GAPDH, 59-TCATCTCTGCCCCCTCTGCT-39. All specific primers synthesized by thephosphoramine method using a DNA synthesizer (model 392 PCR-MATA;Applied Biosystems, Inc., Foster City, CA) were purchased from Sawady(Tokyo, Japan).

Isolation of Colonic Epithelial Cell and Mucosal Lymphocytes. Humancolonic epithelial cells were isolated from resected specimens as describedpreviously (22). Briefly, surgical samples of colonic mucosa were dissectedfree from underlying musculature and washed in CMF-HBSS. Tissues werethen treated with 1 mM DTT (Sigma Chemical Co., St. Louis, MO) in

CMF-HBSS for 15 min at 22°C. After washing three times with CMF-HBSS,the tissue pieces were incubated in CMF-HBSS containing 1 mg/ml dispase(Boehringer Mannheim Biochemicals, Indianapolis, IN) for 30 min at 37°C.During this treatment, epithelial cells and intraepithelial lymphocytes werereleased from the tissues. Dispase treatment was repeated once. The cellsuspensions resulting from dispase treatments were washed twice, pelleted,and resuspended in 3 ml of 100% Percoll (Pharmacia Biotech, Piscataway, NJ).Three-ml layers of 60%, 40%, and 30% Percoll were layered successively ontop before centrifugation at 1500 rpm for 30 min at 4°C. Cells at the top0%–30% layer interface contained epithelial cells. The purity of the epithelialcells was confirmed by flow cytometric analysis using antihuman intestinalepithelial cell mAb B9 and antilymphocyte mAbs against CD3 and CD20. B9(IgG1) was kindly provided by Prof. L. Mayer (Mount Sinai Medical Center,New York, NY; Ref. 23) and was biotinylated in our laboratory. Cells in thispreparation method contained.95% pure epithelial cells, as reported previ-ously (24). Cells were washed three times in RPMI 1640, and viability wasdetermined by trypan blue exclusion.

Colonic LPLs were also isolated from colonic mucosa. After treatment with0.5 mM DTT, the tissue was washed and incubated twice with stirring for 60min at 37°C in CMF-HBSS containing 0.75 mM EDTA to remove epithelialcells. The supernatant was decanted, and the remaining tissue was incubatedwith stirring at 37°C in 5% CO2 in RPMI 1640 containing antibiotics, 10%FCS, 0.02% collagenase (CLSPA; Worthington Biochemical Co., Freehold,NJ), and 0.01% DNase I (DP; Worthington Biochemical Co.) for 18 h. Thecrude cell suspension was filtered through 50-mm stainless mesh and thenwashed. After resuspension in 100% FCS, the cell suspension was separatedover Ficoll-Hypaque. The lymphoid cell suspension was washed and resus-pended in a complete medium.

RESULTS

DD of mRNA Expression between Colitis-associated Cancerand Mucosa with Mild Inflammation in the Same UC Patients.Using mRNA DD-PCR, we assessed the patterns of gene expressionin colitis-associated cancer tissues and mucosa with mild inflamma-tion in the same UC patients. We used eight different 59 primers incombination with three sets of 39 primer pools for the reaction.Combinations of the forward primers AP9–16 and the reverseHT11M primers HT11A, HT11G, and HT11C produced strikinglydifferent patterns when used in our DD-PCRs.

Subcloning and Sequencing of cDNA Fragments Correspond-ing to Differentially Expressed mRNAs. We identified 60 cDNAfragments corresponding to mRNAs differentially expressed in mu-cosa with colitis-associated cancer and with mild inflammation fromall five UC patients using all of the primer combinations. Thesefragments were then excised and subjected to reamplification usingthe same sets of primers. We subcloned 34 fragments and sequencedthem. Approximately 100–500 bp of nucleotide sequences were ob-tained from these fragments and compared to the National Center forBiotechnology Information GenBank database using the BLAST al-gorithm. Among them, 19 cDNA fragments were more stronglyexpressed in UC-associated cancer tissues than in mucosa with mildinflammation. These fragments were considered as candidate cancer-related genes. Only two fragments, DD2 and DD40, were highly(.95%) homologous to known cDNA sequences. Northern blot anal-ysis demonstrated that expression of DD2, which was completelyhomologous to human tyrosine phosphatase, was slightly differentbetween UC-associated cancer and mildly inflamed mucosa. In con-trast, DD40 was expressed quite strongly in UC-associated cancertissues. We identified the nucleotide sequence of DD40, and this114-bp cDNA fragment, including the polyadenylic acid site, wascompletely homologous to the partial sequence of IFN-inducible genefamily 1-8U.

Expression of IFN-inducible Gene Family 1-8U in Colon Can-cer Tissues.In three samples from five UC-associated colon cancerpatients, IFN-inducible gene family 1-8U mRNA was strongly ex-

Table 1 Clinical features of UC in patients with UC-associated colon cancer

Patientno. Age (yr) Clinical type

Duration(yr) Extension

1 67 Chronic continuous 8 Total2 42 Relapsing-remitting 13 Total3 35 Chronic continuous 23 Total4 36 Chronic continuous 14 Total5 25 Relapsing-remitting 12 Left-sided

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pressed in UC-associated cancer tissues rather than in mildly inflamedtissues by Northern blot analysis (Fig. 1A). In two other samples, aNorthern blot could not be done. However, PCR revealed that 1-8Uwas also more strongly expressed in UC-associated cancer tissue thanin inflamed mucosa in those patients (Fig. 1B).

To assess whether high expression of this gene was specific forUC-associated colon cancers, we examined IFN-inducible gene fam-ily 1-8U expression in 10 sporadic colorectal cancers. As shown inFig. 2, 1-8U mRNA was also expressed in all 10 sporadic colorectalcancer tissues. To clarify the expression of this gene in colonicepithelial cells, we examined its expression in some colon cancer celllines. 1-8U was expressed in cancer cell lines, including colon cancercell lines HT29, CW2, and ACM (Fig. 3A), and its expression wasincreased by IFN-a stimulation (data not shown).

In contrast, 1-8U was not expressed in normal colonic mucosa from10 healthy volunteers (Fig. 3B).

Expression of IFN-inducible Gene Family 1-8U in InflamedColonic Mucosa of UC. We then assessed the expression of IFN-inducible gene family 1-8U in inflamed colonic mucosa of UC pa-tients without colon cancer. To determine the cellular source of 1-8U,we first isolated colonic epithelial cells and LPLs from the colonic

mucosa of the patients. As shown in Fig. 4, this gene was stronglyexpressed in freshly isolated colonic epithelial cells. In contrast, thisgene was not strongly expressed in freshly isolated LPLs, even fromseverely inflamed mucosa. This result indicated that we could assessthe mRNA expression of 1-8U in colonic epithelial cells by usingmRNA from colonic mucosa.

Interestingly, this gene was strongly expressed in severely inflamedcolonic mucosa of UC patients without colitis-associated colon cancer(Fig. 5). Expression of 1-8U was quite strong in all eight patients withsevere colonic inflammation (Table 2). In contrast, expression of 1-8Uwas weak or negligible in four of six patients in the remission stagewith very mild colonic inflammation. This gene expression was notrelated to the extent and duration of the disease. However, 1-8U wasexpressed in the colonic mucosa of all four patients with chronic,continuously severe inflammation.

Fig. 1. A, expression of 1-8U mRNA in macroscopically normal mucosa (N), mildlyinflamed colon mucosa (I), and UC-associated cancer tissue (T) of patients 3–5. Thepredicted size of 1-8U mRNA was 0.8 kb. For the internal standard, GAPDH was used asa housekeeping gene.B, RT-PCR demonstrating 1-8U mRNA expression in UC-associ-ated cancer tissues rather than in mildly inflamed colonic mucosa in patients 1 and 2. Thepredicted size of RT-PCR products of 1-8U was 97 bp.

Fig. 2. Expression of 1-8U mRNA in sporadic colon cancer tissues. Ten representativesamples of 1-8U mRNA expression in normal colon mucosa (N) and colon cancer tissues(T) from the same patients are shown.

Fig. 3.A, expression of 1-8U mRNA in cancer cell lines. The human colon carcinomacell lines HT29, Caco2, ACM, WiDr, and CW2 were used. A human cervical cancer cellline, HeLa, was also used.B, expression of 1-8U mRNA in normal colon mucosa fromhealthy volunteers. Six representative samples of 1-8U mRNA expression in normal colonmucosa are shown. HeLa was used as a positive control.

Fig. 4. Expression of 1-8U mRNA in freshly isolated colonic epithelial cells (Epithelialcell) and LPLs from severely inflamed colonic mucosa of a UC patient. Ethidium bromidestaining of the PCR products by DD40 primers is shown.

Fig. 5. Expression of 1-8U mRNA in severely inflamed colonic mucosa and inremission stage mucosa from UC patients without colitis-associated colon cancer. Threerepresentative samples of severely inflamed colonic mucosa and four samples of 1-8UmRNA expression in remission stage mucosa from UC patients were shown. HeLa wasused as a positive control.

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DISCUSSION

Previous molecular and epidemiological studies suggest that tumor-igenesis in UC differs from sporadic colon cancer and that chronicinflammation likely plays a role in the former but not the latter (9, 10,25, 26). To clarify the mechanism of carcinogenesis from chronicinflammation in colonic mucosa, we tried to isolate cDNA fragmentscorresponding to mRNAs that are differentially expressed betweenUC-associated cancer tissue and inflamed mucosa. We isolated 60cDNA fragments from all five samples. Nineteen of 34 sequencedDNA fragments were strongly expressed in UC-associated coloncancer. Two of these fragments were highly homologous to knowncDNA sequences. Only DD40 was definitively overexpressed in UC-associated cancer. Therefore, we focused on the nucleotide sequenceof this DD40 fragment. The DD40 fragment was identical to thepartial sequence of human IFN-inducible gene family 1-8U that arehighly induced by both type I (a andb) and II (g) IFN. IFN-induciblegene family 1-8U was initially cloned from a human lymphoid cellcDNA library (27). 1-8U was expressed in HeLa cell line, and thisexpression was enhanced by IFN-a treatment. Human 1-8U gene-transfected Chinese hamster ovary cells are highly responsive toIFN-a. This gene has the IFN stimulation regulatory element in itspromoter region that is the binding site of IFN stimulated gene factor3 and the IFN regulatory factor family, and transcription of this geneis thought to be regulated by INF signal transduction (27). However,the function of IFN-inducible gene 1-8U in IFN systems is stillunknown.

Interestingly, 1-8U was strongly expressed in both UC-associatedcancer and sporadic colon cancer tissues. In contrast, 1-8U was notexpressed in normal colon mucosa. Consistent with these findings, arecent study reported that IFN-inducible gene family 1-8U and 1-8Dwere strongly expressed in colon primary tumors and in some coloncancer cell lines, but not in normal colon mucosa by serial analysis ofgene expression (28), suggesting that 1-8U may be a colon cancer-associated gene.

Importantly, this colon cancer-associated gene, 1-8U, was stronglyexpressed in severely inflamed mucosa of UC. Expression of 1-8Uwas weak or negligible in UC patients with very mild colonic inflam-mation. In contrast, expression of 1-8U was quite strong in thepatients with severe colonic inflammation. To confirm the purity ofcolonic epithelial cells isolated from mucosa of biopsied specimens,we have performed flow cytometric analysis by using an antihumanintestinal epithelial cell-specific mAb, not just by using lymphocyte-specific mAbs. Cells in our preparation method contained.95% pureepithelial cells (24). Moreover, our recent experiments using epithelialcells with high B9 expression isolated from our epithelial preparationsdemonstrated that those cells strongly expressed I-8U in mucosal

tissues with active UC.4 Therefore, we are quite sure of the purity ofepithelial cells. Our results indicated that 1-8U expression in LPLswas quite weak and was not comparable to that in purified epithelialcells. Although we cannot completely negate contamination by LPLsin our epithelial cell preparations, we are sure that the main cellularsource of 1-8U expression in the UC mucosa was epithelial cells.These issues are supported by the fact that 1-8U expression in LPLsfrom some active UC patients was not demonstrated. In contrast to thesituation regarding epithelial cells, LPLs isolated from mucosal biop-sied specimens contained up to 10% B9-positive colonic epithelialcells in our preparation method.

Interestingly, this gene was expressed in colonic mucosa fromall patients with chronic, continuously severe inflammation. There-fore, this gene expression may be related to the severity of thedisease. This gene expression was not correlated with duration ofthe disease. Long-term persistent inflammation in colonic mucosahas been thought to cause the development of colon cancer inpatients with UC (1–3). A recent study demonstrated that UCpatients with chronic continuously severe inflammation developedcolitis-associated colon cancer in shorter periods of time thanpreviously reported.5 In these patients, colitis-associated coloncancer developed in severely inflamed colonic mucosa. It has beenconsidered that the risk of UC-associated colon cancer dependsonly on the duration of the disease. Our results suggest that theseverity of inflammation may also be important in the tumorigen-esis of UC. Therefore, long-term persistent expression of 1-8U inthe colonic mucosa might be used as a marker that is preferentiallyexpressed in UC-associated colon cancer.

Production and expression of a variety of inflammatory cyto-kines are increased in severely inflamed colonic mucosa of UC,and colonic epithelial cells in severely inflamed colonic mucosa ofUC are exposed to several kinds of IFNs. Therefore, it is quitereasonable that the IFN-inducible gene family is strongly ex-pressed in severely inflamed mucosa, but not in normal colonicmucosa. In contrast, the molecular mechanism for induction of1-8U gene expression and function of the expressed gene incolitis-associated cancer tissues in UC remains unclear. How doesincreased IFN expression as a result of severe inflammation affectcarcinogenesis in UC? Although the function of IFN-induciblegene 1-8U remains unclear, recent studies demonstrated that someIFN-inducible genes and proteins are overexpressed in cancertissues. p27 was overexpressed in breast carcinoma (29), and a newglycoprotein that cross-reacts with carcinoembryonic antigen wasreported to be up-regulated by IFN-g (30). Therefore, IFN-induc-ible genes such as p27, the new glycoprotein, and 1-8U that areoverexpressed in cancer tissues may be cancer-related genes. IFN-inducible genes mediate several kinds of functional roles. IFN-inducible genes such as double-stranded RNA-dependent proteinkinase exert an antiviral and antiproliferative effect (31). IFN-inducible protein p202 binds retinoblastoma protein and inhibitscell growth (32). IFN-inducible protein IP-10 is an antitumor agentthat promotes damage in established tumor vasculature and inducestissue necrosis in human Burkitt lymphoma (33). Therefore, 1-8Uexpression in severely inflamed mucosa and colitis-associated can-cer tissues in UC may also be important in protection against theproliferation of inflammation-mediated cells and tumor cells. Al-though it is not defined whether 1-8U act as antitumor gene orcarcinogenic gene, our study indicates that expression of this genein colonic mucosa may be a useful marker in the identification ofthe high-risk group of UC-associated colon cancer.

4 Unpublished observation.5 T. Hibi and T. Hisamatsu, unpublished observation.

Table 2 Relationship between 1-8U expression in the colonic mucosa and clinicalfeatures in UC patients without cancer

Category

1-8U Expression

1 2

Clinical typeChronic continuous 4 0Relapsing-remitting 6 4

ActivityActive 8 0Remission 2 4

Duration,5 yrs 6 1$5 yrs 4 3

ExtensionTotal 5 3Left sided 5 1

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ACKNOWLEDGMENTS

We thank Motomi Yamazaki for technical assistance; Drs. Akira Sugita,Masahiko Watanabe, Tatsuo Teramoto, and Masaki Kitajima for providing thespecimens; and Reiko Fujisaki for manuscript preparation.

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1-8U EXPRESSION IN UC-ASSOCIATED COLON CANCER

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