Occurrence of p53 Gene Abnormalities in Gastric Carcinoma Tumors and Cell Lines Joo-Hang Kim, Takashi Takahashi, Itsuo Chiba, Jae-Gahb Park, Michael J. Birrer, Jae Kyung Roh, Hy De Lee, Jin-Pok Kim, John D. Minna, Adi F. Gazdar*

Inactivation of tumor suppressor genes is believed to be important in the development of many human malignancies (7-7). Recently, several lines of evidence have indicated that the wildtype p53 gene, located at 17pl3.3, may function as a tumor suppressor gene and that a mutant p53 gene could promote transformation by inactivating normal p53 function in a dominant negative fashion (5,9). In addition to gross DNA abnormalities (5,6,10-12), many different point mutations in the highly conserved regions of the open reading frame have been observed in a variety of common cancers, such as lung, colon, and breast cancers (10,13-15). Allelic deletions of chromosome 17p have been detected in over 75% of colon cancers (14), and point mutations of the p53 gene were detected in nine of 10 tumors selected for 17p allele loss (13). Recently, intronic point 938

Materials and Methods Gastric specimens and cell lines. Primary gastric tumor tissues and samples of corresponding nonmalignant gastric mucosa were obtained during routine surgical diagnostic or curative procedures and were stored at -70 °C until processed. For restriction fragment length polymorphism (RFLP) studies, metastases to regional lymph nodes (if present) also were obtained. Gastric cancer cell lines were established and characterized as previously described (78). Five of the cell lines were established from metastatic lesions, and one was established from a primary tumor (Table 1). With the exception of one cell line (initiated from a patient from the United States), all samples were from Korean patients. Northern and Southern blotting. Methods for preparing DNA, RNA, and probe fragments along with analysis of Northern and Southern blots were performed as previously described (79). The genomic DNAs were digested with Msp I for Southern blot analysis. The probe used is a 1.8-kilobase Xba l-Xba I fragment prepared from a normal human p53 complementary DNA (cDNA) clone, php53cl (20), labeled with 32P by the random primer technique (27). Ribonuclease (RNase) protection assays. RNase protection assays were performed with 5 \ig of RNA as previously

ReceivedOctoberl5,199O, revised March 13, 1991; accepted April 2, 1991. National Cancer Institute-Navy Medical Oncology Branch, Division of Cancer Treatment, National Cancer Institute, and the Naval Hospital, Bethesda, Md. Present address: J.-H. Kim, J. K. Roh, Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Korea. Present Address: J.-G. Park, J.-P. Kim, Department of Surgery, College of Medicine, Seoul National University and Hospital, Seoul, Korea. Present address: H. D. Lee, Department of Surgery, Yonsei University College of Medicine, Seoul, Korea. We thank D. Givol for supplying the p53 probes and Marion Nau for many helpful suggestions and comments. 'Correspondence to: Adi F. Gazdar, M.D., National Cancer Institute-Navy Medical Oncology Branch, Bldg. 8, Rm. 5101, Naval Hospital Bethesda, Bethesda, MD 20814.

Journal of the National Cancer Institute

Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

We explored the state of the p53 gene in gastric cancer. Using one or more methods, we examined 15 specimens from primary carcinomas (14 tumors, one cell line), five cell lines derived from metastases, and seven paired samples of nonmalignant gastric mucosa. Sequence analyses of complementary DNA containing the entire p53 gene open reading frame demonstrated abnormalities in one of five samples from primary tumors and in all five samples from metastases. The single cell line derived from a primary carcinoma had no abnormality of the gene. The six abnormalities included four point mutations, one base-pair deletion resulting in a frame shift, and a 24 base-pair deletion caused by an intronic point mutation (as determined by sequence analysis of genomic DNA). Four of the six mutations mapped to regions highly conserved among species or involved in simian virus 40 T-antigen binding. Restriction fragment length polymorphism studies confirmed that chromosome 17p allelic deletions occur only in a minority of primary tumors, but that they may occur more frequently in metastases. Northern blotting and ribonuclease protection assays detected only a fraction of the p53 gene abnormalities detected by sequencing. Our findings indicate that mutations of the p53 gene are relatively rare in primary gastric tumors but appear to be relatively frequent in cell lines derived from metastatic lesions. Our results may help in understanding the molecular events associated with progression and metastasis in gastric carcinoma. [J NatI Cancer Inst 83:938-943,1991]

mutations have been identified as an alternative mechanism for p53 gene inactivation (75). Cancer of the stomach is still a leading malignant disease in many countries (16,17). Since the state of the p53 gene in gastric cancer has not been previously reported, we examined its structure and expression in gastric cancers and in cell lines derived from them.

Table 1. Summary of assay data Specimen type* Specimen source

Assay

Gastric mucosa

Northern bkx RNase protection

Primary tumors

Northern blot RNase protection Sequencing

Metastases

Fresht

Cell line

0/7 0/7

on 1/14 1/4

Northern blot RNase protection Sequencing

Total (%) 0/7(0) 0/7(0)

0/1 0/1 0/1

0/8(0) 1/15 (7) 1/5(20)

1/3 3/5 5/5

1/3 (33) 3/5(60) 5/5(100)

•Values = No. of abnormal specimens/total No. of specimens examined. tSamples obtained directly from surgical procedures.

Results Northern and Southern Blot Analyses RNAs from eight primary carcinomas (seven tumors and one cell line), four cell lines from metastases, and seven paired nonVol. 83, No. 13, July 3,1991

RNase Protection Assay RNAs from none of the seven nonmalignant mucosa samples, one of 15 primary carcinomas (14 tumors and one cell line), and three of five metastases (all cell lines) showed definite abnormal RNase cleavage patterns (Table 1, Fig. 2). Of these four total abnormalities, two were detected both by the p53M probe (midregion) and by the p53PA probe (3' end). Single abnormalities unique to each of these probes were detected. No abnormalities were detected by the p53XP probe (5' end). In samples having abnormal cleavage patterns, the sum of the protected fragments equaled the length of the fully protected fragments, indicating that the abnormalities were either point mutations or small deletions in the messenger RNA (mRNA). Results from the use of overlapping probes permitted mapping of the mutations, which was confirmed by sequence analysis (see below).

Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

described (22) using three overlapping probes spanning the p53 open reading frame. Amplification and sequencing of cDNA. First-strand cDNA synthesis using 5-10 (Xg of total cellular RNA with p53-specific oligonucleotide primers and subsequent polymerase chain reaction (PCR) amplification were performed as previously described (10,23). Using the nucleotide (nt) numbers of the sequence published by Lamb and Crawford (24) as reference points, the sense and antisense primers used for samples of cell lines SNU-16 and T52 were S3 (nt 966-989) and AS2 (nt 13471370), respectively; S2 (nt 906-929) and AS3 (nt 1170-1193) for samples of the SNU-55 cell line; and SI (nt 540-563) and AS1 (nt 1740-1763) for other samples. All the sequencing data were reconfirmed with clones pooled from the products of two independent p53 gene cDNA/PCR reactions. Amplification and sequencing of genomic DNA. PCR amplification was performed using 2 ng of genomic DNA and the same cycles as those for cDNA/PCR following denaturation at 94 CC for 5 minutes. A pair of sense and antisense primers was used for SNU-5 cell line genomic DNA/PCR, including nt 1299-1325 and nt 1506-1529. All primers had extraneous nucleotides comprising an ZscoRI site at their 5' ends. The PCR products were cloned into pGEM4. The sequencing result was reconfirmed with an independently generated genomic DNA/PCR product. RFLP analysis. We compared nonmalignant mucosal tissues and corresponding primary gastric tumors and lymph node metastases for retention or loss of chromosome 17p heterozygosity. For this comparison, we used the highly polymorphic (86% heterozygosity) probe pYNZ22 (locus D17S5) located in chromosome region 17pl3.3 (25) as well as the less informative 17p probe pMCT35.1 (D17S31) (26). Msp I digestion identifies a two-allele polymorphism. RFLP analyses were performed as previously described (7), using Msp I digestion.

malignant gastric mucosa samples were analyzed by RNA blotting. In all the nonmalignant and primary carcinoma samples and in two of three cell lines from metastases (Table 1, Fig. 1), p53 transcripts of the expected 2.8-kilobase size were readily detected. No transcript was detected in cell line SNU-5 established from a metastasis. Southern blot analysis revealed no evidence of gross structural DNA rearrangements or deletions (data not shown).

Sequence Analysis of p53 mRNA by cDNA/PCR We performed nucleotide sequencing of p53 cDNA containing the entire open reading frame on all 10 samples (four tumors and six cell lines) from which RNA was available, including all four that showed abnormal cleavage patterns by the RNase protection assays. Sequencing demonstrated abnormalities in one of five primary gastric carcinomas (four tumors and one cell line) and in all five cell lines established from metastases (Tables 1, 2). Five of these abnormalities were single missense mutations (T52, SNU-16, SNU-55, YCC-3, and NCI-N87) (Table 2; Fig. 3, A); one sample from a metastasis (cell line SNU-55) contained a single base-pair (bp) deletion causing a frame shift at codon 143 (Fig. 3, B), which resulted in a TGA termination Tissue Samples

Cell Lines 1

2/3-Actin

M

l i

— 2.8 kb

^^

• *

• • • 1

• •-

— 2.0 kb

Fig. 1. Northern blot analysis of p53 gene expression. p53 RNA of the expected 2.8-kb size is expressed by all gastric tumor (prefix T) and normal gastric mucosa (prefix N) samples tested and by three or four gastric cancer cell lines. A transcript was not detected in cell line SNU-5. The P-actin probe was used as a control for RNA loading.

ARTICLES

939

G

B

A

T

C

o. "i to to en to

i. 9" IS.

Point mutation G- T

P53PA .

5'C Amino Acid

200

100 YCC3

300 NCI-N87

393

Fig. 2. Demonstration of p53 mutations by the RNase protection assay. Three overlapping probes spanning the entire open reading frame of the p53 gene were used (panel D). Untranslated regions of the gene are indicated by gray areas at the 5' and 3' ends. Abnormal cleavage patterns are seen with probe p53M (panel B) in cell lines SNU-16 and SNU-55 and in tumor sample T52 and with probe p53PA (panel C) in cell lines SNU-5 and SNU-16 and in tumor sample T52. Abnormal patterns were not detected with probe p53XP (panel A). The locations of p53 mutations detected by the RNase protection assay are plotted above the schematic diagram of the gene (panel D), while p53 mutations detected only by sequencing are plotted below the diagram.

G

A

T

C

B

Table 2. p53 gene mutations in gastric carcinoma* Gastric cancer

Specimen type

Primary T37 T38 T52 T56 SNU-1

Tumor Tumor Tumor Tumor Cell line

Metastases SNU-5 SNU-16 SNU-55 YCC-3 NCI-N87

Cell line Cell line Cell line Cell line Cell line

Mutation nucleotide

Amino acid change

216

Not detected Not detected GTGtoTTG Not detected Not detected

Val to Leu

262-269 205 143 175 248

24-bp deletion TAT to TTT Cl deleted CGC to CAC CGG to CAG

8 amino acids Tyr to Phe Frame shift Arg to His Arg to Gin

Codon

•Cell line NCI-N87 was established from a Caucasian patient in the United States; all other tumor and cell line samples were from Korean patients. Cell line NCI-N87 is well differentiated; other lines are poorly differentiated. Nucleotide abbreviations: A = adenosine; G = guanosine; C = cytidine; T = thymidine.

940

•Deletion!

Fig. 3. Abnormalities of the p53 gene demonstrated by sequencing of cDNA/PCR products. Panel A: Gastric tumor sample T52 contains a point mutation at codon 216 (GTG -> TTG). Panel B: Gastric cancer cell line SNU-55 shows a 1 base deletion (nucleotide C) at codon 143, which results in a frame shift mutation.

Journal of the National Cancer Institute

Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

_p53M_ . p53XP.

codon 88 bp downstream; another sample (cell line SNU-5) contained a 24-bp deletion (nt 1343-1366) (Fig. 4, A, C); no mutation was detected in three primary tumor samples or in the cell line derived from a primary tumor (SNU-1). Sequence Analysis of Genomic DNA The finding of a 24-bp deletion at the 5' end of exon 8 in the cDNA of the SNU-5 cell line suggested an abnormal mRNA splicing event. We amplified and sequenced intron 7 and the surrounding region of the p53 gene in SNU-5. A point mutation in the splice acceptor site at the 3' end of intron 7 was found (Fig. 4, B). This intronic point mutation converts a splice acceptor sequence, 5'-GAGT4G-3', into the sequence 5'GAGTCG-3' (Fig. 4, C). The point mutation appears to abolish normal splicing at this site and results in complex abnormal

splicing that utilizes a cryptic splice acceptor site (5'-GAACAG3'), located near the 5' end of exon 8, resulting in a 24-bp deletion in the mRNA of this cell line. Of interest, two of the p53 gene abnormalities occurred in highly conserved regions of the gene (corresponding to domains II to V in Fig. 5), which are 92% homologous with the comparable region in mouse p53 {2027), while four (including the previously mentioned two) occurred in the simian virus 40 T-.antigen-binding areas (Fig. 5). RFLP Analyses We tested mucosal tissues and corresponding primary tumors and lymph node metastases for retention or loss of chromosome 17p heterozygosity. Of 14 informative cases examined with the polymorphic probe pYNZ22, four (29%) of 14 primary tumors Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

5' G A C T C C A G C T T T G A G 3'

Fig. 4. Structural analysis of cDNA and genomic DNA of the p53 gene in gastric cancer cell line SNU5. Panel A: cDNA sequencing demonstrates a 24-bp deletion in exon 8. Panel B: Sequencing of genomic DNA demonstrates a point mutation (nucleotide A —> C) at the splice acceptor site of intron 7. Panel C: Schematic analysis of the cDNA and genomic DNA of the wild-type p53 gene and of the abnormality in SNU-5. The site of the intronic point mutation in genomic DNA (lower panel) is displayed in reverse video (arrow). This mutation converts a splice acceptor sequence, 5'-GAGT/4G-3', into the sequence 5'-GAGTCG-3' (lower part of panel). The resulting abnormal cDNA is shown in the upper part of the panel.

cONA EXON 7

Normal

T C C S e r

SNU-5

A G

EXON 8

T

G G T G l y

S e r

A A T A t n

C T A L e u

C T G L e u

G G A G l y

C G G A r g

A A C A s n

T C C AG S e r S e r

A G C S e r

C

T T T P h i

G A G G l u

G T G V ) I

T T T G A G P he G l u

G T G V•I

GENOMIC DNA EXON 7

EXON 8

Normal T C C AG

g tca...333bp...t g a g

SNU-5

g t c a . . . 3 3 3 b p . . . t g a g t Q g

T C C AG

Vol. 83, No. 13, July 3, 1991

t ag T

G G T

A A T

C T A

C T G

G G A

C G G

A A C

A G C

t g g t

a a t

e t a

c t g

g g a

e g g

a a c a g C

T T T

G A G

G T G

TTT

GAG

GTG

ARTICLES

941

carcinomas also frequently map to these regions (7,13), indicating that mutations in the p53 gene often occur in important func216 tional regions. RFLP analysis confirmed and extended these 143 175 205 findings. A minority (29%) of primary gastric tumors demonstrated loss of heterozygosity for chromosome 17p. In addition, loss of heterozygosity was present in the regional node Evolutionary conserved regions (Domains I,II,III,IV,V) metastases of two cases, while the corresponding primary tumor The predicted binding sites of p53 to large T antigen was heterozygous. Neumann et al. (25) have reported a similar incidence (21%) of 17p allelic deletions in gastric cancers. Allelic deletions of chromosome 17p are common in colon cancers Fig. 5. Location of the six p53 gene mutations detected in gastric carcinoma samples (arrows). Large, open box represents a schematic diagram of the p53 (14), and most allelic deletions in lung and colon cancers have open reading frame (5' to 30, with amino acid positions noted. Thefiveconmutated p53 genes (729). Northern blotting and RNase protecserved regions (black bars, roman numerals) and the two simian virus 40 T-antion assays detected only a fraction of the p53 gene abnortigen-binding regions (gray boxes) are indicated. malities in gastric carcinomas detected by sequencing. RNase protection assays detect only approximately 50% of single bp demonstrated loss of heterozygosity. Four (44%) of nine substitutions (30 Jl). regional lymph node metastases demonstrated loss of Allelic loss of chromosome 17p is common in primary heterozygosity, including two cases in which both tumor and colorectal tumors and often is present at the time of malignant node metastasis showed loss of heterozygosity and two cases in transformation of adenomas (32). Both primary and metastatic which the node metastasis showed loss of heterozygosity while colorectal tumors are relatively easy to establish as continuous the corresponding primary tumor was heterozygous. Of four incultures with an overall success rate of about 40% (33), possibly formative cases examined with probe pMCT35.1, one primary because p53 mutations occur at a relatively early stage in tumor showed loss of heterozygosity. One of the tumors (T52) tumorigenesis. In contrast, our findings suggest that p53 mutaexamined by RFLP analysis was sequenced and found to have tions in gastric carcinomas occur after development of the p53 gene mutation at codon 216 (Table 2). This tumor specimen malignant phenotype and are more frequent in regional or disdemonstrated loss of heterozygosity for chromosome 17p by tant metastases. Gastric cell lines are difficult to establish, and both of the probes. most of the existing lines originated from metastatic lesions (18). In our experience, only 3% of primary tumors and 25% of Discussion metastatic tumors can be successfully cultured (18). Of interest, all five cell lines established from metastatic lesions had p53 Gastric carcinoma is the most common form of gastrointesmutations, while the single cell line established from a primary tinal malignancy in certain parts of the world, including most of tumor lacked an abnormality. While the possibility that in vitro the Far East, yet relatively little is known about the molecular culture results in the induction of p53 gene mutations or selects events leading to its development Recent data have shown that for tumor cells having mutated genes cannot be excluded, Baker allelic loss of chromosome 17p and mutations of the p53 gene et al. (29) found a similar incidence of p53 mutations in primary are common in certain human tumors, including lung and colon colon tumors and cell lines. Since wild-type, but not mutant, p53 tumors {10,13,14). Therefore, we explored the state of the p53 protein suppresses cell growth in vitro (9J4J5), ourresultssuggene in samples from primary and metastatic gastric cargest that these differences in cell line establishment rates may be cinomas. Cell lines constituted all of the five metastatic samples due to the relative frequency of p53 gene mutations in metasand one of 15 of the primary tumor samples. In addition, seven tatic tumors. paired samples of nonmalignant gastric mucosa from patients In summary, mutations of the p53 gene are common in gastric with primary tumors were examined. cancer cell lines established from metastatic lesions, but they are Abnormalities of the p53 gene were detected by cDNA seless common in primary gastric tumors. These findings may be quencing in one (25%) of four primary gastric tumors and in important in understanding the molecular events associated with five (83%) of six cell lines; all five of these cell lines were inprogression and metastasis in gastric carcinoma. itiated from metastatic lesions. (The single negative cell line was initiated from a primary tumor.) The abnormalities included four point mutations in the open reading frame, resulting in four missense mutations, a single base deletion resulting in a frame References shift, and a 24-bp deletion. The latter abnormality was caused by (/) HANSEN MF, CAVENEE WK: Genetics of cancer predisposition. Cancer Res an intronic point mutation in the splice acceptor site of intron 7. 47:5518-5527,1987 Intronic point mutations of the p53 gene also have been (2) KNUDSON AG JR: Hereditary cancer, oncogenes, and antioncogenes. Cancer Res 45:1437-1443,1985 described in lung cancers (75). Two of the six mutations (i) MURPHREE AL, BENEDICT WF: Retinoblastoma: Clues to human onmapped to regions conserved in evolution (1020). Four of the cogenesis. Science 223:1028-1033,1984 mutations (including the two previously mentioned) map to two (4) MASHAL R, SHTALRID M, TALPAZ M, ET AL: Rearrangement and expression of p53 in the chronic phase and blast crisis of chronic myelogenous discontinuous regions of p53 gene, which include domains III leukemia. Blood 75:180-189,1990 and V of the conserved regions and which have been identified (5) MULLIGAN LM, MATLASHEWSKJ GJ, SCRABLE HJ, ET AL: Mechanisms of as coding for the simian virus 40 large T-antigen-binding site p53 loss in human sarcomas. Proc Natl Acad Sci USA 87:5863-5867, 1990 (27). Mutations of the p53 gene in colorectal and pulmonary 100

200

II

III

300 V del 248 262-269

393

IV

Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

942

Journal of the National Cancer Institute

(6) BRESSAC B, GALVIN KM, LIANG TJ, ET AL: Abnormal structure and expres-

(22) GREEN WR, UNNOILA RI, TRICHE TJ: Neuroendocrine carcinoma of skin

sion of p53 gene in human hepatocellular carcinoma. Proc Natl Acad Sci USA 87:1973-1977, 1990

with simultaneous cytokeratin expression. Ultrastruct Pathol 6:141-152, 1984

(7) CHIBA I, TAKAHASHI T, NAU MM, ET AL: Mutations in the p53 gene are fre-

(23) DOHERTY PJ, HUESCA-CONTRERAS M, DOSCH HM, ETAL: Rapid amplifica-

quent in primary, resected non-small cell lung cancer. Lung Cancer Study Group. Oncogene 5:1603-1610, 1990

tion of complementary DNA from small amounts of unfractionated RNA. Anal Biochem 177:7-10, 1989 (24) LAMB P, CRAWFORD L: Characterization of the human p53 gene. Mol Cell Biol 6:1379-1385, 1986

(8) EUYAHU D, MtCHALOvrrz D, ELIYAHU S, ET AL: Wild-type p53 can inhibit

oncogene-mediated focus formation. Proc Natl Acad Sci USA 86:87638767,1989 (9) FINLAY CA, HINDS PW, LEVINE AJ: The p53 proto-oncogene can a a as a

suppressor of transformation. Cell 57:1083-1093, 1989 (70) TAKAHASHI T, NAU MM, CHIBA I, ET AL: p53: A frequent target for genetic

abnormalities in lung cancer. Science 246:491-494, 1989 (11) MASUDA H, MILLER C, KOEFFLER HP, ET ALJ Rearrangement of the p53

(25) NAKAMURA Y, BALLARD L, LEPPERT M, ET AL: Isolation and mapping of a

polymorphic DNA sequence (pYNZ22) on chromosome 17p P17S30]. Nucleic Acids Res 16:5707,1988 (26) CARLSON M, NAKAMURA Y, PAYSON R, ET AL: Isolation and mapping of a

polymorphic DNA sequence pMCT35.1 on chromosome 17p [D17S31]. Nucleic Acids Res 16:783, 1988

gene in human osteogenic sarcomas. Proc Natl Acad Sci USA 84:77167719,1987

(27) Soussi T, CARON DE FROMENTEL C, MAY P: Structural aspects of the p53

(12) AHUM H, BAR-EU M, ADVANI SH, ET AL: Alterations in the p53 gene and

(28) NEUMANN WL, JACOBY RF, WASYLYSHYN M, ET AL: A comparison of

the clonal evolution of the blast crisis of chronic myelocytic leukemia. Proc Natl Acad Sci USA 86:6783-6787, 1989

genetic alterations occurring in colon, gastric and pancreatic adenocarcinomas. In The Molecular Basis of Human Cancer. Abstracts of the first meeting, 1990, p 32

(13) NKJRO JM, BAKER SJ, PRHSINGER AC, ET AL: Mutations in the p53 gene

occur in diverse human tumour types. Nature 342:705-708, 1989 (14) BAKER SJ, FEARON ER, NIGRO JM, ET AU Chromosome 17 deletions and

(29) BAKER SJ, PREISINGER AC, JESSUP JM, ET AL: p53 gene mutations occur in

combination with 17p allelic deletions as late events in colorectal tumorigenesis. Cancer Res 50:7717-7722, 1990

(15) TAKAHASHI T, D'AMICO D, CHIBA I, ET AL: Identification of intronic point

(50) WINTER E, YAMAMOTO F, ALMOGUERA C, ET AL: A method to detect and

mutations as an alternative mechanism for p53 inactivation in lung cancer. J Clin Invest 86:363-369, 1990

characterize point mutations in transcribed genes: Amplification and overexpression of the mutant c-Ki-ras allele in human tumor cells. Proc Natl Acad Sci USA 82:7575-7579, 1985

(16) MINISTRY OF HEALTH AND SOCIAL AFFAIRS, REPUBLIC OF KOREA; One

year's report for cancer registry programme in the Republic of Korea. July 1, 1985-June3O, 1986. J Kor Cancer Assoc 19:131-258,1987 (17) METTUN C: Epidemiologic studies in gastric adenocarcinoma. In Gastric Cancer (Douglass HO, ed). New York: Churchill Livingstone, 1988, pp 1-25 (18) PARK JG, FRUCHT H, LAROCCA RV, ET AL: Characteristics of cell lines es-

tablished from human gastric carcinoma. Cancer Res 50:2773-2780, 1990 (19) DAVIS LG, DIBNER MD, BATTEY JF: Basic Methods in Molecular Biology.

Amsterdam: Elsevier/North Holland, 1986

(31) MYERS RM, LARIN Z, MANIATIS T: Detection of single base substitutions

by ribonuclease cleavage at mismatches in RNA:DNA duplexes. Science 230:1242-1246,1985 (32) VOGELSTEIN B, FEARON ER, HAMILTON SR, ET AL: Genetic alterations

during colorectal-tumor development. N Engl J Med 319:525-532, 1988 (33) PARK JG, OIE HK, SUGARBAKER PH, ET AL: Characteristics of cell lines es-

tablished from human colorectal carcinomas. Cancer Res 47:6710-6718, 1987

(20) ZAKUT-HOURI R, BIENZ-TADMOR B, GIVOL D, ET AL: Human p53 cellular

(34) BAKER SJ, MARKOWITZ S, FEARON ER, ET AL: Suppression of human

tumor antigen: cDNA sequence and expression in COS cells. EMBO J 4:1251-1255,1985

(35) MERCER WE, SHIELDS MT, AMIN M, ET AU Negative growth regulation in

(21) FEINBERG AP, VOGELSTEIN B: A technique for radiolabeling DNA restric-

tion endonuclease fragments to high specific activity. Anal Biochem 132:6-13,1983

12

colorectal carcinoma growth by wild-type p53. Science 249:912-915, 1990 a glioblastoma tumor cell line that conditionally expresses human wildtype p53. Proc Natl Acad Sci USA 87:6166-6170, 1990

Research Resources

Issues each year filled with current biomedical research findings Subscribe Today

Vol. 83, No. 13, July 3, 1991

Reporter Don't Miss an Issue of the Research Resources Reporter. For Direct Mail Delivery to Your Home or Office Every Month Only $9 Per Year (U.S.) or $11.25 (Foreign Address, Via Airmail)

Published by the National Center (or Research Resources, NH Fin out the order form below and mail to: The Superintendent of Documents, US. Government Priming Office, Washington D.C. 20402 Send the Research Resources Reporter for one year to:

• My check or money order for is enclosed. D Charge to my: D Visa D MasterCard number Interbank number Expires To order by telephone using Visa or MasterCard, cafl (202) 783-3238.

ARTICLES 943

Downloaded from http://jnci.oxfordjournals.org/ at Laurentian University on November 5, 2014

p53 gene mutations in colorectal carcinomas. Science 244:217-221, 1989

protein in relation to gene evolution. Oncogene 5:945-952, 1990

Occurrence of p53 gene abnormalities in gastric carcinoma tumors and cell lines.

We explored the state of the p53 gene in gastric cancer. Using one or more methods, we examined 15 specimens from primary carcinomas (14 tumors, one c...
4MB Sizes 0 Downloads 0 Views