Cell cycle-regulated factors in esophageal cancer.

Alterations of cell cycle-regulated genes play an important role in the process of carcinogenesis, and some of them are thought to be prognostic factors in esophageal cancer. The expressions of p53, p16, pRB and Cyclin D1 proteins were evaluated immunohistochemically in 144 patients who underwent curative esophagectomy without any adjuvant therapy before surgery. p53 overexpression was observed in 99 (69%) out of the 144 patients. No significant correlation was noted between p53 and any other gene expression. p16 expression was observed in 12 (8.3%) out of all cases. A negative correlation was recognized between p16 and Cyclin D1 expression (P=0.0004). pRB expression was observed in 130 (90.3%) out of all cases, whereas pRB expression was not observed in 11 out of the 12 patients with p16-positive tumors. A negative correlation was also found between p16 and pRB (P < 0.0001). A positive correlation was noted between pRB and Cyclin D1 expression (P=0.0009). The cumulative survival rate of patients without pRB expression was significantly lower than that of patients with positive expression (P=0.003). In the multivariate survival analysis, pRB expression was an independent prognostic factor. In 98% of all patients with esophageal cancer, impairment of the G1 checkpoint is due to a loss of function by p16, pRB or Cyclin D1, which showed a negative correlation in each factor. In addition, aberrant expression of pRB is useful as a prognostic factor in esophageal cancer.

Different susceptibility of each L-myc genotype to esophageal cancer risk factors.

To understand the relationship between the L-myc genotypes and esophageal cancer risk, a polymerase chain reaction-based restriction fragment length polymorphism analysis was performed on 91 Japanese patients with esophageal cancer and 241 non-cancer outpatients. No significant difference in the distribution of genotypes was observed between patients and controls; 18.7% LL genotype, 56.0% LS and 25.3% SS among patients, and 24.5%, 55.6% and 19.9%, respectively, among controls. Frequency of the s-allele in patients (0.533) was slightly higher than in controls (0.477), but the difference was not statistically significant. However, the odds ratios (ORs) for smoking or heavy drinking were markedly higher in SS and LS genotypes than in LL genotype; age-sex-adjusted ORs for smoking was 7.57 in the SS genotype, 6.40 in the LS genotype and 1.77 in the LL genotype. Age-sex-adjusted ORs for heavy drinking were 19.78, 18.20 and 7.40, respectively. The age-sex-adjusted ORs for both factors combined were 12.77, 18.45 and 1.44, respectively. These results suggested that the L-myc polymorphism might modify the effects of lifestyle factors on esophageal cancer risk.

Possible risk reduction in esophageal cancer associated with MPO -463 A allele.

Myeloperoxidase (MPO), an enzyme found in lysosomes of phagocytes, causes hydroxy radicals linked to DNA damage and activation of smoking related carcinogens. A -463 G/A polymorphism in the promoter region of the MPO gene results in reduced gene expression, which would imply lower susceptibility of esophageal cancer in mutant carriers. We conducted case-control study to test this hypothesis. Cases were 91 patients with esophageal cancer and controls were 241 non-cancer outpatients. MPO genotypes were examined by PCR-RFLP. The allele frequency for MPO -463A was found to be 8.2% for cases and 10.5% for controls. The age, sex, smoking and drinking status adjusted odds ratio for all subjects for MPO -463 GG/GA as compared to the AA was 0.61 (95% CI: 0.28-1.32). The adjusted odds ratio for the GG/GA genotype was significantly low (0.15; 0.03-0.76, P=0.022) for those aged 61 years or older who had a significantly higher odds ratio for smoking than younger subjects. No difference was observed in disease risk when prevalent and incident cases were compared. Although there are limitations for interpretation of this study because of prevalent case-control study and partial statistical significance, these results suggest that MPO -463 A allele reduce the risk of esophageal cancer.

Gene-environment interaction between an aldehyde dehydrogenase-2 (ALDH2) polymorphism and alcohol consumption for the risk of esophageal cancer.

Aldehyde dehydrogenase-2 (ALDH2) degrades acetaldehyde metabolized from ethanol. Its encoding gene ALDH2 has a functional polymorphism: ALDH2 Glu487LYS: An association between this polymorphism and esophageal cancer among alcoholics has been reported. To further evaluate the gene-environment interaction, a hospital-based case-control study was conducted. Cases were 102 patients with histologically confirmed esophageal cancer and controls were 241 non-cancer outpatients of Aichi Cancer Center. ALDH2 genotypes were examined by a PCR-CTPP method developed in our laboratory, which does not require a digestion stage. Logistic regression analysis was employed for estimation of relative risk and gene-environment interaction. The allele frequency for ALDH2 Lys487 was 0.28, consistent with previous reports. The age, sex, smoking and drinking status adjusted odds ratio for the ALDH2 Glu/Lys and Lys/Lys genotypes as compared with the Glu/Glu genotype was 3.43 (95% CI 1.74-6.75). The odds ratio for heavy drinking was 49.6 (14.5-169.4) among Lys487 carriers and 7.84 (2.77-22.2) for the Glu/Glu genotype. The gene-environment interaction between alcohol drinking and the ALDH2 Lys487 allele was 6.84 (2.39-19.6), whereas no significant interaction was obtained with smoking status. Although limited because of its prevalent case-control design, our study revealed a strong gene-environment interaction between ALDH2 polymorphism and heavy alcohol consumption. Taking the observed high risk of esophageal cancer in association with the ALDH2 Lys487 allele into consideration, reducing alcohol intake may be most protective among Lys487 allele carriers of this polymorphism.

High frequency of clonally related tumors in cases of multiple synchronous lung cancers as revealed by molecular diagnosis.

In patients with multiple synchronous lung tumors, discrimination of multicentric lung cancers from intrapulmonary metastasis is important for treatment decision, but this is sometimes difficult. The aim of this study was to retrospectively distinguish multicentric lung cancers from intrapulmonary metastases in 14 such cases by loss of heterozygosity (LOH) and p53 mutational status. DNA was extracted from microdissected tumor cells in paraffin-embedded archival tissue, and 3p14.2, 3p21, 3p25, 9p21, and 18q21.1 were investigated for LOH. Exons 5-8 of the p53 gene were examined for mutations by the PCR, followed by single-strand conformation polymorphism analysis and DNA sequencing. For cases with the same LOH pattern, we calculated a clonality index, the probability of the given LOH pattern when these tumors were hypothesized to be independent in origin. Eleven of 14 cases (79%) were thus diagnosed as having pulmonary metastasis and only one case as having genuinely multicentric lung cancers. Two cases presented difficulty in diagnosis. In several cases, the LOH patterns conflicted with p53 mutation patterns, suggesting that clonal evolution is directly affected by certain genetic changes. The combination of p53 with LOH helped increase both the sensitivity and specificity of the assay.

Subsite-specific risk factors for hypopharyngeal and esophageal cancer (Japan).

OBJECTIVES: To clarify subsite-specific risk factors for hypopharyngeal and esophageal cancers (HC and EC), we concluded a hospital-based case-referent study in Nagoya, Japan. METHODS: Subjects comprised 346 male cases with cancer of the hypopharynx (n = 62) or esophagus (upper [U-EC] 53, middle [M-EC] 159, lower [L-EC] 72), and 11,936 male referents free from cancer among first-visit outpatients aged 40-79 years in 1988-1997. Of histological confirmed cases, 93% comprised squamous cell carcinoma. Odds ratios (ORs) were estimated by a logistic regression model with adjustment for potential confounding factors. RESULTS: Cigarette smoking increased the OR for M-EC, and alcohol drinking elevated the ORs for all subsites. The trend of ORs for combined cases of M- and L-EC tended to increase with number of cigarettes (p = 0.056), and a decreasing trend of the ORs was found with years after quitting smoking (p = 0.006). The ORs for smoking with drinking were multiplicatively greater than those for smoking or drinking in combined cases of HC and EC. In contrast, daily raw vegetable consumption lowered the ORs for all subsites. CONCLUSIONS: This study suggests that the magnitude of risk with smoking is stronger for M-EC within the esophagus, and drinking increases the risk at any subsite.

Cyclin D1 expression is useful as a prognostic indicator for advanced esophageal carcinomas, but not for superficial tumors.

The purpose of the present study was to define the overexpression of cyclin D1 in superficial and advanced esophageal carcinomas and to investigate whether the expression of this molecule indicates a poor prognosis. This study included 41 patients with superficial esophageal carcinomas (Tis and T1) and 48 patients with advanced esophageal carcinomas (T2, T3, and T4). The expression of cyclin D1 in surgically resected specimens was evaluated immunohistochemically with a monoclonal antibody. Positive immunoreactivity was found in 31 of 89 cases (35%). Overexpression of cyclin D1 did not correlate with TNM classification or histologic type. Of the 48 patients with advanced esophageal carcinomas, 32 patients with cyclin D1-negative tumors survived longer than did 16 patients with cyclin D1-positive tumors (P = 0.0017). In contrast, we observed no survival difference between patients with cyclin D1-positive and -negative superficial esophageal carcinoma. These results suggest that cyclin D1 indicates a poor prognosis in cases of advanced esophageal carcinoma but not in cases of superficial esophageal carcinoma.

Polymorphic variation of the ARP gene on 3p21 in Japanese esophageal cancer patients.

Allelic loss of the short arm of chromosome 3 is common in esophageal squamous cell carcinoma (ESC) and its premalignant lesions. The ARP gene (Arginine Rich Protein) was mapped to 3p21, and frequent variations of the triplet, AGG, repeat around codon 50 of the ARP gene were reported in a variety of human cancers. To examine the involvement of the ARP gene in esophageal cancer, we screened mutations around codon 50 in 35 ESC tumours and matched normal tissues. Sequence variants were observed in four ESC tumours; two (AGG)2 insertions, one ATG50-to-AGG substitution and one AGG deletion. However, they were also found in its corresponding normal tissues, suggesting that variation of the ARP gene found in ESC is polymorphic. We next analyzed sequence changes in 48 unrelated Japanese healthy individuals. They consist of 33 wild-type homozygotes, nine (AGG)2 insertion/wild-type heterozygotes, two (AGG)2 insertion homozygotes, one AGG deletion/wild-type heterozygote and three ATG50-to-AGG substitution/wild-type heterozygotes. Allele frequencies for wild-type, (AGG)2 insertion, ATG50-to-AGG substitution and AGG deletion are 0.82, 0.14, 0.03 and 0.01, respectively. Observed genotype frequencies fit well with the Hardy-Weinberg's law. Significant difference was not observed between allele distributions in normal and cancer patient populations.

Massive intrathoracic haemorrhage after CT-guided lung biopsy.

CT-guided lung biopsy is now widely performed for tumorous lesions in the lung, and both its usefulness in this context and the associated complications have been well described in the literature. Although severe complications are rare, we describe a case in which massive intrathoracic haemorrhage developed after lung biopsy and necessitated emergency operation for control. Intraoperative findings suggested that the source of the haemorrhage was a fibrous, cord-like substance present at the site of adhesion associated with old tuberculosis. We attributed this haemorrhage to a pneumothorax, which developed after lung biopsy and caused the new vessels penetrating the centre of the fibrous, cord-like substance to stretch and rupture. Numerous cases have been reported of spontaneous haemopneumothorax precipitated by spontaneous pneumothorax and resulting from the rupture of such vessels.

Clinical implications of p53 autoantibodies in the sera of patients with non-small-cell lung cancer.

BACKGROUND: The presence of autoantibodies to p53 protein has been associated with the presence of p53 (also known as TP53) gene mutations in primary tumors and with poor prognosis. This study was undertaken to determine the clinical significance of p53 autoantibodies in patients with non-small-cell lung cancer (NSCLC). METHODS: We studied 188 consecutive patients with NSCLC who underwent pulmonary resection and for whom preoperative serum was available. The presence of p53 autoantibodies, detected by use of two amino-terminal and two carboxy-terminal peptides (20-30 mers) as antigens and an enzyme-linked immunosorbent assay, was related to various clinicopathologic parameters and to overexpression of p53 protein in the primary tumor. For 22 patients who had p53 autoantibodies before surgery, we also examined sera taken during postoperative follow-up. Reported P values are two-sided. RESULTS: Autoantibodies to p53 protein were detected in 38 patients. Patients with squamous cell carcinoma, those with more advanced disease (stage III-IV), and those with tumors that overexpressed p53 had a significantly higher incidence of p53 autoantibodies (P = .05,.0079, and .02, respectively). In all but one of the patients with postoperative serum samples, the antibody titer declined after surgery; however, there was no relationship between clinical course and this change in antibody titer. In addition, there was no relationship between the presence of p53 autoantibodies and overall survival in 171 patients who underwent potentially curative resection (P = .28); however, 13 patients with autoantibodies to amino-terminal peptides had a worse overall survival (P = .02). CONCLUSIONS: In NSCLC, the incidence of p53 autoantibodies is associated with histologic type, stage, and p53 overexpression--but not with patient survival. Our data do not support the clinical utility of p53 autoantibodies as diagnostic or prognostic markers in patients with NSCLC.

Methylation of the 5' CpG island of the FHIT gene is closely associated with transcriptional inactivation in esophageal squamous cell carcinomas.

Previous studies suggested that the FHIT (fragile histidine triad) gene on 3p14.2 might be involved in the development of esophageal squamous cell carcinomas, but the mechanisms for inactivating the gene have not been fully revealed. In the present study, we examined aberrations of the FHIT gene in 23 esophageal squamous cell carcinoma cell lines and 35 primary tumors. We detected aberrant expression in seven cell lines (30%), including a shorter transcript in two cell lines and loss of apparent transcript in five cell lines. Genomic PCR or cDNA sequencing analysis revealed a single exon deletion in two cell lines with a shorter transcript and one cell line without expression, but no structural alterations were found in the other 20 cell lines, including transcriptionally repressed four cell lines. Next we examined methylation of the 5' CpG island of the FHIT gene by bisulfite genomic sequencing. Hypermethylation of the 5' CpG island of the FHIT gene was observed in three of four structurally unaltered but transcriptionally repressed cell lines. The remaining cell line harbored a point mutation upstream of exon 1. All methylated cell lines exhibit re-expression of the FHIT gene and demethylation in the CpG island after treatment with demethylating agent 5-aza-2'-deoxycytidine. Hypermethylation was also found in 5 of 35 (14%) primary tumors, whereas corresponding normal tissue shows no methylation. These findings suggest that methylation of the 5' CpG island of the FHIT gene is closely associated with transcriptional inactivation and might be involved in tumor development of the esophagus.

Mutations of the P53 tumor suppressor gene as clonal marker for multiple primary lung cancers.

OBJECTIVES: Second primary lung cancers are prevalent after treatment for initial lung cancer, and the lung is also one of the most frequent sites for recurrence after removal of early-stage lung cancer. The objective of the present study is to clarify the clonal origin of the second tumor with the p53 gene mutation used as a clonal marker. METHODS: Of 794 consecutive patients who underwent pulmonary resection for primary lung cancer from 1980 to 1993, 22 required second pulmonary resection during the follow-up period, with a median interval of 38 months. We examined 16 of these patients for mutations of the p53 gene occurring in exons 5 through 8 by the polymerase chain reaction/single strand conformation polymorphism method. Differential diagnosis was also made on a morphologic basis, considering the degree of cellular differentiation and cytologic subtypes. RESULTS: Nine of the 16 patients analyzed had at least one p53 mutation in their tumors. We were thus able to make molecular diagnoses for these patients. The mutational status of the p53 gene was discordant in all nine patients, suggesting a different clonal origin despite the fact that six of them had almost identical histologic features. CONCLUSIONS: Analysis of p53 gene mutations was thus useful in distinguishing second primary lung cancers from recurrent tumors. The observed heterogeneity of p53 status was also in line with the "field cancerization" concept.

[A carcinoma arising from benign pleomorphic adenoma of the trachea].

A 69-year-old female complained of husky voice and dyspnea. Bronchofiberscopic examination revealed a tumor at the left side of the trachea, which obstructed approximately 80% of the tracheal lumen. Benign pleomorphic adenoma was diagnosed by biopsy. The tumor was removed by circumferenctial resection of the trachea with partial sternotomy adding to the 3 th intercostal transverse resection. Historical finding of the resected specimen revealed a carcinoma arising from benign pleomorphic adenoma of the trachea and residue of malignancy at the margin of the trachea and esophagus. Additional radiotherapy was performed (60 Gy). The postoperative course was uneventful for 4 months and one year.

Papillary carcinoma in a huge intrathoracic goiter with tracheal stenosis and invasion. Case report.

A rare case in tracheal invasion by a papillary carcinoma in an intrathoracic goiter is reported. Subtotal thyroidectomy and tracheoplasty were performed. Surgery should be considered as first-line treatment of intrathoracic goiter in order to avoid future compression of the respiratory tract and to remove possible concomitant malignant lesion.

[Clinical evaluation of myocardial protection by oxygenated crystalloid cardioplegic solutions with DBcAMP].

Possible enhancement of myocardial protection by adding DBcAMP and oxygenation of a crystalloid cardioplegic solution (CCS) was evaluated in a three group study. The patients having coronary bypass operation or valvular operation were divided into three groups, each consisting of 15 patients, and differing only in the type of CCS employed. Group I was protected by nonoxygenated CCS (PO2 190 mmHg, PCO2, 32 mmHg, pH 7.78, K 30 mmEq/L), Group II by adding DBcAMP to nonoxygenated CCS and Group III by adding DBcAMP to oxygenated CCS (PO2 790 mmHg, PCO2 26 mmHg, pH 7.87). Group III had significantly improved CI and double product (p less than 0.05) compared with Group II. However, CPK, CPK-MB, and myoglobin in the serum were similar in each group. Lactate and pyruvate ratio (L/P) in the coronary sinus bloods were improved to lower value after the pump than before the pump only in Group III. Base excess in the coronary sinus held on alkalosis after aortic declamp only in Group III. The refunction time was significantly shortest with Group III than with other groups (p less than 0.01, 0.05) and Group II was significantly shorter than Group I (p less than 0.05). It is concluded that oxygenation and adding DBcAMP to CCS are effectual for the myocardial metabolism and protect the myocardial damage during cardiac arrest.