An attempt to define sets of cooperating genetic alterations in human breast cancer.

The extent and the variation of losses of genetic material were examined in a series of 191 human breast cancers by means of a set of 18 polymorphic DNA probes, specific of 7 chromosomal arms (1p, 1q, 3p, 11p, 13q, 17p and 18q) known to be frequently affected by allele losses. Frequencies of losses of heterozygosity ranged from a low of 3.5% (chromosome 13q) to a high of 27% (chromosome 3p). The number of sites involved in breast cancer added to the frequent occurrence of concomitant losses at several chromosomal arms within the same tumor suggest cooperative effects of these LOHs. We were therefore interested in assessing the existence of preferential associations between sets of LOHs in our panel of tumors. Statistically significant associations were found between LOHs at chromosomes 1p and 17p, and between LOHs at chromosomes 11p and 17p. Furthermore, since all the tumors presently studied had previously been analyzed for proto-oncogene amplification at 5 distinct chromosomal sites, we tested for associations between LOH and DNA amplification. Such associations were indeed observed as exemplified by the correlations observed between the LOH at 11p and amplification of the erbB2 gene and LOH at 17p and the amplification of the flg gene. The only correlation with clinico-pathological parameters that could be observed linked the occurrence of LOHs on 11p with recurrent breast cancer (p = 0.015). Sets of several LOHs or LOHs and gene amplifications could not be significantly related to any marker of tumor aggressiveness.     

非小细胞肺癌中3p^25杂合性丢失研究

目的为探讨非小细胞肺癌（ＮＳＣＬＣ）细胞中第三号染色体短臂上３ｐ２５位点上等位基因的杂合性丢失（ＬｏｓｓｏｆＨｅｔｅｒｏｚｙｇｏｓｉｔｙＬＯＨ）与肺癌发生发展的相关性。方法采用银染聚合酶链反应（ＰＣＲ）结合二核苷酸（ＣＡ）ｎ重复程序出现多态性评价杂合性丢失方法，分析５８例ＮＳＣＬＣ组织中３ｐ２５等位基因的杂合性丢失。结果５８例肺癌组织中有２９例出现３ｐ２５ＬＯＨ，总的杂合性丢失率为５０％。３ｐ２５的杂合性丢失率在１８例腺癌中为１２例（６６．７％），明显高于鳞癌（４２．９％，１５／３５），有显著性统计学意义（Ｐ＜０．０５），但３ｐ２５杂合性丢失与ＮＳＣＬＣ临床分期关系不明显，Ⅰ期为４２．１％，Ⅱ期为５５．６％和Ⅲ期为５２．２％，５例正常胎儿肺组织均不出现３ｐ２５ＬＯＨ。结论３ｐ２５杂合性丢失在ＮＳＣＬＣ中普遍存在，从而说明３ｐ２５位点处存在着某些与肺癌发生发展有关的抑癌基因。     

Loss of heterozygosity in malignant rat schwannomas chemically induced in hybrids of inbred rat strains with differential tumor susceptibility

Rats of the inbred BD strains strongly differ in their susceptibility to the induction of tumors of the central (CNS) and peripheral nervous system (PNS) by N-ethyl-N-nitrosourea (EtNU). Malignant schwannomas induced in (BDIX x BDIV) and (BDIX x BDVI) rat hybrids were analyzed to identify genetic alterations associated with EtNU-induced tumorigenesis in the PNS. EtNU-induced schwannomas exclusively exhibit an A:T T:A transversion mutation of the neu/Erbb-2 gene located on chromosome 10, with subsequent loss of the wild-type neu/Erbb-2 allele at a post-initiation stage. Targeted allelic deletion mapping previously revealed losses of heterozygosity (LOH) at the distal end of chromosome 10 in a large majority of (BDIX x BDIV) schwannomas. The aims of the present study were (i) to scan the whole genome for further LOHs; (ii) to narrow down the consensus regions of frequently occurring allelic deletions using tumors from different crosses of BD rats; and (iii) to determine the sequence of genetic alterations during schwannoma development. A limited number of (BDIX x BDIV) F(1) tumors were initially screened for LOH and microsatellite instability (MI) by amplifying 58 microsatellite markers spanning the whole genome. LOHs on chromosome 5 were detected in 9/17 tumors, with random loss of the parental alleles. Ninety-two schwannomas from different BD rat-crosses were then analyzed to solidify these data and to determine the consensus region of frequent LOHs. The results indicate that LOHs on chromosomes 10 and 5 are required for the development of EtNU-induced malignant schwannomas from immature neu/Erbb-2 mutant glial cells, and that putative tumor suppressor genes are localized on chromosome 10q32.3, corresponding to human chromosome 17q25.3, and the telomeric region of mouse chromosome 11, and on the telomeric quarter of chromosome 5. MI was detected in <0.2% of cases.     

Carcinogen-specific targeting of chromosome 12 for loss of heterozygosity in mouse lung adenocarcinomas: implications for chromosome instability and tumor progression

Genotoxic carcinogens exert their tumorigenic effects in part by inducing genomic instability. We recently showed that loss of heterozygosity (LOH) on chromosome 12 associates significantly with the induction of chromosome instability (CIN) by the likely human lung carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and vinyl carbamate (VC) during mouse lung carcinogenesis. Here, we demonstrate the carcinogen specificity of this event and its effect on lung tumor evolution. LOH on chromosome 12 was observed in 45% of NNK-induced, 59% of VC-induced, 58% of aflatoxin B1 (AFB1)-induced, 14% of N-ethyl-N-nitrosourea (ENU)-induced and 12% of spontaneous lung adenocarcinomas. The frequency of LOH in each of the carcinogen-induced groups, except ENU, was significantly higher than in the spontaneous group (P<0.001). Deletion mapping revealed four potential candidate regions of 1-4 centiMorgans suspected to contain targeted tumor suppressor genes, with at least one expected to have a role in CIN. The relationship between LOH on chromosome 12 and additional chromosomal alterations occurring during lung tumor progression was also examined. LOH on chromosomes 1 and 14 were moderately frequent during malignant progression in tumors from all treatment groups, occurring in 21-35 and 18-33% of tumors. However, these alterations showed significant concurrence with LOH on chromosome 12 in VC-, NNK- and AFB1-induced tumors (P<0.05). The results suggest that a carcinogen-selective mechanism of lung cancer induction involves the frequent inactivation of genes on chromosome 12, including a stability gene that evidently promotes the evolutionary selection of additional chromosomal alterations during malignant progression.     

Specific association between alcohol intake, high grade of differentiation and 4q34-q35 deletions in hepatocellular carcinomas identified by high resolution allelotyping

One of the most frequent deletions in hepatocellular carcinoma (HCC) is that involving the long arm of chromosome 4 (30 to 70% of the cases). These chromosomal deletions are closely related to hepatitis B virus (HBV) infection. A tumor suppressor gene (TSG) located on 4q has been proposed in liver carcinogenesis, but has not been identified as yet. Despite previous LOH studies focused on 4q in HCC, a clear minimal common region of deletion (MCRD) could not be delimited. To further investigate the role of chromosome 4q LOH in the pathogenesis of HCC, 85 microsatellite markers spanning chromosome 4q were systematically analysed in a series of 154 well-characterized primary liver tumors. In 59 tumors (38%), LOHs were observed for at least two adjacent markers. Analysis of 31 tumors demonstrating a partial or interstitial 4q deletion allowed to define three MCRDs of 15, 9 and 8 Mb at the 4q22, 4q34 and 4q35 regions, respectively. Seven putative candidate genes located in 4q22, DAPP1, BMPR1B, PKD2, HERC3, SMARCAD1, CEB1 and ENH were screened for mutations but no somatic alterations were identified. Search for relationships between the specific regions of deletion and clinical parameters showed a significant association between loss of the 4q34-35 region with alcohol intake (P=0.005) and with high grade of differentiation (P=0.02). These results are in contrast with the close association between HBV infection and the whole 4q LOH and reveal heterogeneity of 4q LOH in relation to different risk factors. In the light of these new findings, which link different 4q LOH regions to different etiologic factors, the molecular mechanisms underlying 4q deletions in HCC and the targeted gene(s) remain to be identified.     

Prognostic impact of multiple allelic losses on metastatic recurrence in hepatocellular carcinoma after curative resection

Loss of heterozygosity (LOH) on chromosomes 13q, 16q and 17p has been associated with the progression of hepatocellular carcinoma (HCC). To investigate the prognostic impact of such LOH, we examined the metastasis-free survival of curatively resected HCC cases, in whom these LOHs were analyzed. Among the 49 HCC patients examined, the frequency of LOHs was 28% on 13q, 33% on 16q and 40% on 17p. The patients were followed up for metastatic recurrence after surgery and for analysis of the relationship between chromosomal changes and patients' metastasis-free survival. Univariate survival analysis showed the presence of LOH on 16q, 17p and the number of chromosomes with LOH were significantly and negatively associated with metastasis-free survival, indicating that patients with LOH on multiple chromosomes had a poorer prognosis after surgery than those with LOH on a single chromosome or no LOH. Multivariate Cox survival analysis identified the presence of LOH on 16q and the number of chromosomes with LOH as the most significant independent negatively predictive factors for metastasis-free survival. These findings indicate that accumulation of chromosomal changes is associated with metastatic behavior, and that LOH on 16q was the most useful prognostic indicator for metastasis after curative resection of HCC.     

乳腺癌组织BRCA1、BRCA2基因区域杂合性缺失的研究

目的　研究乳腺癌组织BRCA1、BRCA2基因区域的杂合性缺失 (LOH)情况。方法　采用聚合酶链式反应(PCR)和聚丙烯酰胺凝胶电泳 ,对 30例散发性乳腺癌和 2例姐妹乳腺癌进行LOH的检测 ,同时还采用G显带法对姐妹乳腺癌患者的外周血淋巴细胞染色体畸变情况进行分析。结果　在散发性乳腺癌中 ,BRCA1、BRCA2基因区域的杂合性缺失率分别为 6 12 %和 5 77% ;姐妹二人乳腺癌组织在BRCA2基因区域的D13S173位点处存在LOH ,且她们的外周血淋巴细胞发生了大量的稳定性和非稳定性畸变 ,畸变率远远高于自发畸变率。结论　乳腺癌易感机理在东西方种族有可能存在差异 ,应深入进行研究     

Interstitial chromosomal deletion of the tuberous sclerosis complex 2 locus is a signature for radiation‐associated renal tumors in Eker rats

Ionizing radiation can damage DNA and, therefore, is a risk factor for cancer. Eker rats, which carry a heterozygous germline mutation in the tumor‐suppressor gene tuberous sclerosis complex 2 (Tsc2), are susceptible to radiation‐induced renal carcinogenesis. However, the molecular mechanisms involved in Tsc2 inactivation are unclear. We subjected Fischer 344 × Eker (Long Evans Tsc2^+/?) F1 hybrid rats to gamma‐irradiation (2 Gy) at gestational day 19 (GD19) or postnatal day 5 (PND5) and investigated the patterns of genomic alterations in the Tsc2 allele of renal tumors that developed at 1 year after irradiation (N = 24 tumors for GD19, N = 10 for PND5), in comparison with spontaneously developed tumors (N = 8 tumors). Gamma‐irradiation significantly increased the multiplicity of renal tumors. The frequency of LOH at the chromosome 10q12 region, including the Tsc2 locus, was 38%, 29% and 60% in renal carcinomas developed from the nonirradiated, GD19 and PND5 groups, respectively. Array comparative genomic hybridization analysis revealed that the LOH patterns on chromosome 10 in renal carcinomas were classified into chromosomal missegregation, mitotic recombination and chromosomal deletion types. LOH of the interstitial chromosomal deletion type was observed only in radiation‐associated carcinomas. Sequence analysis for the wild‐type Tsc2 allele in the LOH‐negative carcinomas identified deletions (nonirradiated: 26%; GD19: 21%) and base‐substitution mutations (GD19: 4%). Reduced expression of Tsc2 was also observed in the majority of the LOH‐negative carcinomas. Our results suggest that interstitial chromosomal deletion is a characteristic mutagenic event caused by ionizing radiation, and it may contribute to the assessment of radiation‐induced cancer risk.     

Carcinogenic induction directs the selection of allelic losses in mouse lung tumorigenesis

Recent evidence suggests that genome-wide allelic imbalances are inducible by carcinogens and may occur as cells adapt to carcinogenic exposure during tumorigenesis. We investigated the role of carcinogenic exposure on global and selected loss of heterozygosity (LOH) during mouse lung adenocarcinogenesis. Tumor induction by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) or vinyl carbamate (VC) resulted in a significant overall increase in the number of chromosomes affected by LOH per tumor when compared with spontaneous lung tumors. Allelic loss on chromosome 12 occurred at a frequency of 35% and 40% in NNK- and VC-induced tumors, respectively, compared with 8.3% in spontaneous tumors (P < 0.01). In contrast, spontaneous lung adenocarcinomas displayed LOH on chromosome 4 at a frequency of 77%, whereas a frequency of only 36% (P < 0.001) was observed in tumors induced by NNK. Sixty-four percent of the VC-induced tumors displayed LOH on chromosome 4. In addition, allelic losses on chromosomes 12 and 14 were significantly associated with an increase in chromosomal instability, suggesting that genes inactivated on these chromosomes may contribute to this effect. The results from this study demonstrate that genotoxic carcinogens increase chromosome instability, as evidenced by a significant increase in global LOH frequency, and significantly alter the selection of chromosomal alterations during lung tumor development.     

Identification of candidate genes on chromosome band 20q12 by physical mapping of translocation breakpoints found in myeloid leukemia cell lines.

Deletions of the long arm of chromosome 20 have been reported in a wide range of myeloid disorders and may reflect loss of critical tumor suppressor gene(s). To identify such candidate genes, 65 human myeloid cell line DNAs were screened by polymerase chain reaction (PCR) for evidence of allelic loss at 39 highly polymorphic loci on the long arm of chromosome 20. A mono-allelic pattern was present in eight cell lines at multiple adjacent loci spanning the common deleted regions (CDRs) previously defined in primary hematological samples, suggesting loss of heterozygosity (LOH) at 20q. Fluorescence in situ hybridization (FISH) was then performed using a series of yeast artificial chromosomes (YACs) ordered in the CDR, and in five of eight cell lines, the deletions resulted from cytogenetically detectable whole chromosomal loss or large interstitial deletion, whereas in another cell line deletion was associated with an unbalanced translocation. LOH in the CMK megakaryocytic cell line, which has a hypotetraploid karyotype, was associated with a der(20)t(1;20)(q32;q12)x2 leading to complete deletion of the CDR. Three additional unbalanced translocations were found within the CDR and all three breakpoints mapped to a single YAC. We then used a series of P1 artificial chromosomes (PACs) spanning this YAC clone, and two PACs produced 'split' signals suggesting that they each span one of these breakpoints. Exon trapping using PACs that overlap the breakpoint regions yielded portions of six genes and evaluation of these genes as candidate tumor suppressor genes is underway. The limited information available about these genes suggests that the h-l(3)mbt gene is the most attractive candidate.     

Frequent homozygous deletion of the LKB1/STK11 gene in non-small cell lung cancer

LKB1/STK11 is a tumor suppressor and a negative regulator of mammalian target of rapamycin signaling. It is inactivated in 30% of lung cancer cell lines but only 5-15% of primary lung adenocarcinomas. There is evidence that homozygous deletion (HD) of chromosome 19p at the LKB locus contributes to the inactivation of the gene in primary human lung cancers. Here, we used several complementary genetic approaches to assess the LKB1 locus in primary non-small cell lung cancers (NSCLCs). We first analyzed 124 NSCLC cases for allelic imbalance using eight microsatellite markers on chromosome 19p, which revealed an overall rate of 65% (80 of 124) loss of heterozygosity (LOH). We next used chromogenic in situ hybridization (CISH) to directly examine the chromosomal status of the LKB1 locus. In all, 65 of 124 LOH tested samples were available for CISH and 58 of those (89%) showed either loss of one copy of chromosome 19p (LOH, 40 of 65 cases, 62%) or both copies (HD 18 of 65 cases, 28%). The occurrence of HD was significantly more frequent in Caucasian (35%) than in African-American patients (6%) (P=0.04). A total of 62 of 124 samples with LOH at one or both markers immediately flanking the LKB1 gene were further analyzed by directly sequencing the complete coding region, which identified 7 of 62 (11%) tumors with somatic mutations in the gene. Jointly, our data identified total inactivation of the LKB1 gene by either HD or LOH with somatic mutation in 39% of tested samples, whereas loss of chromosome 19p region by HD or LOH at the LKB1 region occured in 90% of NSCLC.     

Allelic loss in ovarian cancer

Loss of heterozygosity (LOH) was examined at 86 loci distributed on every chromosomal arm in 50 human ovarian tumors. Frequent allele losses were observed on chromosomes 13q (42%), 17p (42%), 17q (45%), and Xp (41%). Deletion mapping on chromosome 17 revealed a candidate gene on the long arm distal to D17S41/S74 for ovarian cancer which is distant from the locus for early onset breast cancer. LOH on chromosome 17q was found to be concordant with LOH on chromosomes 3p, 13q, 17p and Xp suggesting that it may be an early event in neoplastic development. These findings indicate that multiple tumor-suppressor genes for ovarian cancer possibly exist on chromosomes 13q, 17, and/or Xp and provide the basis for the identification of candidate gene(s) associated with ovarian cancer. The chromosomal mechanisms resulting in allele losses in ovarian cancer include deletion, deletion/duplication, mitotic recombination and monosomy, in concordance with the developed genetic model.     

Tobacco smoke-induced DNA damage and an early age of smoking initiation induce chromosome loss at 3p21 in lung cancer

The short arm of chromosome 3 is thought to harbor a novel oncogenic locus that is important in the genesis of lung cancer. The region at 3p21 is believed to contain a distinct locus that is sensitive to loss from the action of tobacco smoke carcinogens and has been reported to be specifically targeted for deletion in lung cancer. To investigate whether 3p21 alteration in lung cancer is associated with carcinogen exposure, PCR-based analysis was performed to detect loss of heterozygosity (LOH) on chromosome 3 at 3p21 in non-small cell lung carcinoma (NSCLC). We also measured instability at the BAT-26 locus, because the mismatch DNA repair gene, hMLH1, is found at 3p21. LOH at 3p21 was analyzed for association with the clinical features of NSCLC, p53 mutation status, polynuclear aromatic hydrocarbon-DNA adduct levels (measured using 32P-postlabeling) and carcinogen exposure information including cigarette smoking and asbestos exposure. Of 219 lung cancers, 150 cases (68.5%) were informative at the D3S1478 locus, and 44.2% of squamous cell carcinoma cases and 30.2% of adenocarcinoma cases showed 3p21 LOH. None of the cancers showed BAT-26 instability. The prevalence of 3p21 LOH was higher in both current and former smokers compared with never smokers and was higher in p53 mutated cases. Among squamous cell carcinoma cases, there was a strong association of increased 3p21 LOH with increasing polynuclear aromatic hydrocarbon-DNA adducts levels (P = 0.03), as well as an increased prevalence LOH with earlier age of smoking initiation (P = 0.02). Our results confirm that 3p21 LOH is strongly associated with measures of biologically effective dose of exposure to tobacco carcinogens. Our results also suggest that alterations of hMLH1 are not related to any of the reported associations, because there was no evidence of microsatellite instability. Finally, LOH in 3p21 may be an early molecular event in NSCLC, because it is significantly associated with a tendency to start smoking at a young age.     

Sequential deletions in both arms of chromosome 9 are associated with the development of head and neck squamous cell carcinoma in Indian patients

In the deletion mapping of chromosome (chr) 9 in head and neck lesions of the Indian patient population by microsatellite markers, we have identified four discrete areas (D1-D4) with high loss of heterozygosities (LOHs) viz. 9p24-p23 (D1), 9p22-p21 (D2), 9q11-q13 (D3) and 9q22.3 (D4) regions. The deletions in D2 and D4 regions were suggested to be essential for the development of dysplastic lesions of head and neck, whereas the deletions in D1 and D3 regions were responsible for progression of the dysplastic lesions to early invasive head and neck squamous cell carcinoma (HNSCC). The microsatellite size alterations (MAs) were observed in the chromosomal 9pter-p23, 9p22-p21(D2), 9q13 and 9q21.1-q21.2 regions with gradual increase during progression of the tumor. Additional chromosomal alterations like loss of normal copy of chr.9 and biallelic alterations were also seen in our samples. There is a correlation between HPV infection with TNM stages, histopathological grades and LOHs at D1 and D4 regions. Whereas tobacco habit is associated with the occurrence of LOHs at D1 and LOHs / MAs at D2 region.     

人非小细胞肺癌中FHIT等位基因缺失和突变的研究

目的　探讨FHIT等位基因缺失、突变在肺癌发生、发展中的作用。方法　应用PCR SSCP和DNA序列分析方法对 3 5例人非小细胞肺癌和 4个肺癌细胞株中FHIT基因的 4个外显子 (外显子 3、4、5、8)和微卫星D3S13 0 0、D3S13 12、D3S13 13进行研究 ,并以远癌肺组织和 10例肺良性病变组织做对照。结果　在 3 5例肺癌中 ,2 2例肺癌发生了一个或两个以上的FHIT等位基因缺失 ,缺失率为 62 .86% ( 2 2 /3 5 )。在鳞癌中 ,FHIT等位基因缺失率 ( 88.2 4% ,15 /17)明显高于腺癌 ( 3 8.89% ,7/18) (P <0 .0 1) ;在吸烟患者中 ( 76.19% ,16/2 1)亦明显高于不吸烟患者 ( 4 2 .86% ,6/14 ) (P <0 .0 5 )。而FHIT等位基因缺失与肺癌的细胞分化程度、P TNM分期、原发肿瘤大小、部位、患者性别、年龄及有无转移均无明显关系 (P >0 .0 5 )。Lewis肺癌、A5 49细胞株亦有FHIT基因部分缺失。 4例肺癌组织具有微卫星灶D3S13 12点突变 ,经DNA序列分析显示均为D3S13 12微卫星灶基因的 87位点密码子发生了CT点突变。结论　FHIT基因异常主要以等位基因的缺失为主 ,而点突变发生率较低。FHIT等位基因缺失主要发生在肺鳞癌和吸烟患者中 ,且FHIT基因可能为烟草致肺癌的靶基因 ,其等位基因缺失可能是肺癌的早期分子事件。     

Relationship between the loss of heterozygosity and tobacco smoking in pulmonary adenocarcinoma

A loss of heterozygosity (LOH) is a major cause of lung carcinogenesis, and it is considered to be related to tobacco smoking in central type lung cancer. We investigated the relationship between LOH in lung adenocarcinoma and tobacco smoking. In a consecutive series of 50 patients with lung adenocarcinoma who underwent a surgical resection, cancer tissue specimens and corresponding normal peripheral lung and central bronchial tissue specimens were analyzed for LOH at the regions of D3S1234 (FHIT), D3S1300 (FHIT), D9S171 (CDKN2), and D17S796 (p53) by polymerase chain reaction using four fluorescence-labeled dinucleotide markers. To examine how cells are influenced by smoking, the A549 cell line was exposed to benzo[a]pyrene (B[a]P) for 24 weeks and then was subjected to the above analysis. The LOH in cancer tissue was thus detected in four (17%) patients at D3S1234, six (14%) at D3S1300, and seven (18%) at D17S796, but no LOH was detected in any normal tissue specimens. The incidence of LOHs in cancer tissue specimens from active smokers was 21% at D3S1234, 11% at D3S1300, and 19% at D17S796, whereas that of LOHs from nonactive smokers was 0% at D3S1234, 19% at D3S1300, and 14% at D17S796. Analyzing the relationship between the pack-year index and the presence of LOH, a significant difference was found among the active smokers. Besides, in the A549 cell line exposed to B[a]P, LOH was de novo detected in one (D2S123) of the nine regions examined. The incidence of LOH could be influenced by tobacco smoking in lung adenocarcinoma, thus suggesting the presence of an important event in the carcinogenesis of this disease.     

Expression of the DMBT1 Gene Is Frequently Suppressed in Human Lung Cancer

DMBT1 (deleted in malignant brain tumors) is a candidate tumor suppressor gene that has been mapped to chromosome 10q25.3-q26.1, a region in which frequent loss of heterozygosity (LOH) has been observed in several human tumors. Since DMBT1 is highly expressed in the lung, we analyzed LOH at the DMBT1 locus and expression of this gene in lung cancer. Thirty-five (53%) of 66 primary lung cancers showed LOH, and diminished expression of DMBT1 was observed in 20 (91%) of 22 lung cancer cell lines: three (14%) of them showed loss of expression. We further determined the primary structure of DMBT1 and analyzed genetic alterations in this gene using 23 lung cancer cell lines. Two (9%) of them had homozygous deletion within the gene, and two cell lines had genetic aberrations: one was a rearrangement involving exons 5 and 6, and the other was a missense mutation at codon 52. These results suggest that inactivation of the DMBT1 gene plays an important role in human lung carcinogenesis.     

Expression of the DMBT1 gene is frequently suppressed in human lung cancer.

DMBT1 (deleted in malignant brain tumors) is a candidate tumor suppressor gene that has been mapped to chromosome 10q25.3-q26.1, a region in which frequent loss of heterozygosity (LOH) has been observed in several human tumors. Since DMBT1 is highly expressed in the lung, we analyzed LOH at the DMBT1 locus and expression of this gene in lung cancer. Thirty-five (53%) of 66 primary lung cancers showed LOH, and diminished expression of DMBT1 was observed in 20 (91%) of 22 lung cancer cell lines: three (14%) of them showed loss of expression. We further determined the primary structure of DMBT1 and analyzed genetic alterations in this gene using 23 lung cancer cell lines. Two (9%) of them had homozygous deletion within the gene, and two cell lines had genetic aberrations: one was a rearrangement involving exons 5 and 6, and the other was a missense mutation at codon 52. These results suggest that inactivation of the DMBT1 gene plays an important role in human lung carcinogenesis.     

Detailed characterization of a homozygously deleted region corresponding to a candidate tumor suppressor locus at distal 17p13.3 in human lung cancer

17p13.3 is one of the chromosomal regions most frequently affected by allelic loss in a variety of human neoplasms including lung cancer. A number of loss of heterozygosity (LOH) analyses have suggested the existence of a tumor suppressor gene at 17p13.3, distal to the p53 locus at 17p13.1. In the present study, we characterized a homozygous deletion at 17p13.3 in a small cell lung cancer cell line by constructing a bacterial artificial chromosome (BAC) contig and a restriction map surrounding the region, as well as by utilizing publicly available draft sequences. We defined the breakpoint, assigned and analysed two known genes, 14-3-3 epsilon and CRK, and a novel gene LOST1 within or at the end of the homozygous deletion of about 170 kb in size. Marked reduction of LOST1 expression was detected in 69% (11/16) of lung cancer specimens by quantitative real-time RT-PCR, while significant DNA hypermethylation was observed at the 5' end of the LOST1 gene in four of six lung cancer cell lines with negligible LOST1 expression. We also show here that a polymorphic marker D17S1174, which resides within the homozygous deletion, was apparently located in the middle of the minimum LOH region, providing further supportive evidence for the presence of a tumor suppressor gene(s) in this region.