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The people of Linxian County, China have one of the world's highest rates of esophageal cancer. Two intervention trials were conducted to determine whether supplementation with specific vitamins and minerals could lower mortality from or incidence of cancer in this population and whether supplementation with multiple vitamins and minerals would reduce esophageal and gastric cardia cancer in persons with esophageal dysplasia. About 30,000 general population (GP) subjects in the GP trial were randomly assigned to one of eight intervention groups according to a one-half replicate of a 2(4) factorial experimental design and were supplemented for 5.25 y with four combinations of micronutrients at doses from one to two times the US recommended dietary allowance (RDA). About 3000 subjects in whom dysplasia was diagnosed in the dysplasia trial were randomly assigned to groups receiving daily supplementation with 14 vitamins and 12 minerals at two to three times the US RDA or placebo for 6 y. Results of the dysplasia trial indicate that in individuals with esophageal dysplasia, micronutrient supplementation had little effect on T lymphocyte responses. In contrast, male participants in the GP trial who were supplemented with beta-carotene, vitamin E, and selenium showed significantly (P0.05) higher mitogenic responsiveness of T lymphocytes in vitro than those not receiving these micronutrients.

Linxian, China, has one of the highest esophageal-cancer mortality rates in the world. In 1983, esophageal balloon-cytology screening was performed to identify subjects eligible for 2 nutrition-intervention trials in Linxian; 12,877 subjects had cytology slides which were satisfactory for diagnosis. Of the 12,649 subjects with squamous-cell diagnoses, 31% were normal by Chinese cytologic criteria; 38% showed hyperplasia; 21% showed dysplasia 1; 6% showed dysplasia 2; 2% showed near-cancer; and 2% showed cancer. Of the 1,471 subjects with columnar-cell diagnoses, 31% were normal; 44% showed hyperplasia; 16% showed dysplasia 1; 4% showed dysplasia 2; 2% showed near-cancer; and 3% showed cancer. Squamous dysplasia and cancer were more common among females than males, while columnar dysplasia and cancer showed male predominance. The prevalence of dysplasia and cancer of both cell types increased with age. The prevalence of squamous dysplasia was significantly higher than in earlier balloon-cytology screenings in Linxian, probably reflecting changes in cytologic classification.

To determine the association between tooth loss and the risk of developing esophageal squamous cell carcinoma, gastric cardia adenocarcinoma, or gastric non-cardia adenocarcinoma in a prospective study.Cox proportional hazards regression was used to examine these associations in a 28,868-person cohort followed prospectively for 5.25 years. The baseline questionnaire included questions regarding tooth loss, and individuals reporting lost teeth had their teeth counted by study personnel. The analytic cohort included 620 esophagus, 431 gastric cardia, and 102 gastric non-cardia cancer cases.Tooth loss was associated with a significantly elevated risk of developing all three cancers. When examined as median splits, tooth loss was associated with a relative risk (RR) (95% confidence interval, CI) of 1.3 (1.1-1.6) in the esophagus, 1.3 (1.0-1.6) in the gastric cardia, and 1.8 (1.1-3.0) in the gastric non-cardia. Further analysis demonstrated that this increased risk was most strongly associated with the loss of the first few teeth and was primarily confined to the younger members of our cohort.In this cohort tooth loss increased the risk of developing upper gastrointestinal cancer. We hypothesize that this may be related to alterations in oral bacterial flora and subsequent increases in the in-vivo production of carcinogens such as nitrosamines.

Ecologic studies of esophageal squamous cell carcinoma (ESCC) have reported an association with consumption of maize contaminated with Fusarium verticillioides, which produce fungal toxins referred to as fumonisins. Fumonisins disrupt sphingolipid metabolism and serum sphingolipids have been proposed as biomarkers of fumonisin exposure. We conducted a prospective nested case-control study to examine the relationship between serum sphingolipids and ESCC incidence.Cases and controls were selected from a large prospective trial conducted in Linxian, People's Republic of China. Ninety-eight ESCC cases were randomly selected from the 639 incident ESCC ascertained during the initial 5.25 years of follow-up: 185 controls were also randomly selected based on the distribution of cases among six age and sex strata. Concentrations of sphinganine and sphingosine were determined by high-performance liquid chromatography in serum collected at the study baseline.No significant associations were found between serum sphingosine, sphinganine, or the sphinganine/ sphingosine ratio and ESCC incidence in conditional and unconditional logistic regression models with adjustment for age, sex, tobacco use. and alcohol use.Our study is the first prospective study to assess the relationship between sphingolipid levels, as biomarkers of fumonisin exposure, and cancer incidence. We found no significant association between sphingolipid levels and risk of ESCC.

Esophageal squamous cell carcinoma (ESCC) is one of the most common fatal cancers worldwide, and north central China has some of the highest rates in the world. Previous studies from tumors in this area of China have shown high frequencies of allelic loss on chromosome 17p13-11, which includes the region where the TP53 gene is found. We examined 56 ESCC patients using single-strand conformation polymorphism and DNA sequencing to assess the frequency and spectrum of TP53 mutation and the association between allelic loss at microsatellite marker TP53 and TP53 mutations. Ninety-six % of cases were found to have at least one genetic alteration, including TP53 mutation (77%), allelic loss within the TP53 gene (73%), and/or loss of heterozygosity at the TP53 microsatellite marker (80%); 75% had two or more such alterations, including 59% with both a point mutation and an intragenic allelic loss ("two hits"). The majority of mutations observed were in exon 5, where the most common type of nucleotide substitution was a G:C--A:T or C:G--T:A transition, including half that occurred at CpG sites. Allelic loss was most commonly found in exon 4 but was very common in exon 5 as well. Taken together, the multiple genetic alterations of TP53 in this population at high risk for ESCC indicate that there is a very high degree of genetic instability in these tumors, that TP53 is a primary target for inactivation, and that this tumor suppressor gene plays a critical role in the carcinogenesis process for ESCC.

Esophageal cancer is one of the most common fatal cancers worldwide. Deletions of genomic regions are thought to be important in esophageal carcinogenesis. We conducted a genomewide scan for regions of allelic loss using microdissected DNA from 11 esophageal squamous-cell carcinoma patients with a family history of upper gastrointestinal tract cancer from a high-risk region in north central China. Allelic patterns of 366 fluorescently labeled microsatellite markers distributed at 10-cM intervals over the 22 autosomal chromosomes were examined. We identified 14 regions with very high frequency (/= 75%) loss of heterozygosity (LOH), including broad regions encompassing whole chromosome arms (on 3p, 5q, 9p, 9q, and 13q), regions of intermediate size (on 2q, 4p, 11p, and 15q), and more discrete regions identified by very high frequency LOH for a single marker (on 4q, 6q, 8p, 14q, and 17p). Among these 14 regions were 7 not previously described in esophageal squamous-cell carcinoma as having very high frequency LOH (on 2q, 4p, 4q, 6q, 8p, 14q, and 15q). The very high frequency LOH regions identified here may point to major susceptibility genes, including potential tumor suppressor genes and inherited gene loci, which will assist in understanding the molecular events involved in esophageal carcinogenesis and may help in the development of markers for genetic susceptibility testing and screening for the early detection of this cancer. Genes Chromosomes Cancer 27:217-228, 2000. Published 2000 Wiley-Liss, Inc.

Chromosomal regions with frequent allelic loss may point to major susceptibility genes that will assist in understanding molecular events involved in esophageal carcinogenesis. Esophageal squamous cell carcinoma samples and blood from 46 patients, including 23 patients with and 23 patients without a family history of upper gastrointestinal cancer, were screened using laser microdissected DNA and tested for loss of heterozygosity (LOH) at 18 marker loci representing 14 chromosomal regions (on 2q, 3p, 4p, 4p, 5q, 6q, 8p, 9p, 9q, 11p, 13q, 14q, 15q, and 17p) identified in an earlier genome-wide scan to have frequent LOH. Clinical/pathological and lifestyle risk factor data were also collected. For all 46 tumors combined, the lowest frequency LOH for any of the 18 markers was 37%, and 8 markers showed LOH inor =75% of informative tumors. One marker (D13S894 on 13q) showed greater LOH in patients with a positive family history (93% versus 50%; P = 0.04), whereas two markers (D6S1027 on 6q and D9S910 on 9q) had significantly more LOH in patients with metastasis, and one marker (D4S2361 on 4p) showed significantly higher LOH in patients with a lower pathological tumor grade. No relation was seen between LOH and lifestyle risk factors. This study confirms the previously observed high frequency LOH for these 14 chromosomal regions, including a locus on 13q where LOH is more common in patients with a family history of upper gastrointestinal cancer than in those without such history, suggesting that a gene in this area may be involved in genetic susceptibility to esophageal cancer.

In previous studies in the high risk population of Linxian, China, the majority of foci of high grade (moderate and severe) squamous dysplasia (HGD) and invasive squamous carcinoma (CA) of the esophagus were associated with endoscopically visible lesions that could be targeted for biopsy, but some foci of HGD were missed by routine endoscopic examination. This study examined whether spraying the mucosa with Lugol's iodine solution, which stains normal epithelium brown but leaves dysplasia and carcinoma unstained, could improve endoscopic detection and delineation of these lesions.Two hundred twenty-five Linxian adults with balloon cytologic evidence of dysplasia or carcinoma underwent endoscopy. All visible lesions were described and photographed before and after staining with 1.2% Lugol's iodine solution. Biopsies were taken from all lesions visible before staining, from all unstained lesions (USLs) after applying the stain, and from representative control areas of stained mucosa.Two hundred fifty-three USLs and 255 control sites were biopsied. No complications occurred. Ninety-four biopsy sites contained HGD and 20 contained CA. Before staining, the sensitivity of visible lesions for identifying HGD or CA was 62%, and the specificity was 79%. After staining, the sensitivity of USLs for identifying HGD or CA was 96%, and the specificity was 63%. Eighty-eight percent of the HGD and CA lesions were larger or more clearly defined after staining. The diagnostic lesions in 17 of 31 patients with moderate dysplasia (55%), 8 of 35 patients with severe dysplasia (23%), and none of the 19 patients with invasive carcinoma (0%) were identified only after staining.Mucosal iodine staining improved endoscopic detection and delineation of HGD and CA in these patients. This simple technique is highly sensitive for identifying these precursor and invasive squamous lesions, and it should be used whenever optimal visualization of squamous mucosal abnormalities is required.

The highest incidences of esophageal and gastric cardia cancer in the world occur in northern China. Chinese scientists have developed esophageal balloon cytology screening to detect these cancers, but traditional cytology is sometimes inadequate to find some early, curable lesions. Several studies suggest that quantitative fluorescence image analysis (QFIA) of DNA ploidy and nuclear morphology may be able to improve upon traditional cytology results. In October 1987, esophageal balloon cytology was performed on 1331 adults in Linxian, China, and all samples were evaluated both by traditional cytology and QFIA. From 1987 to May 1991, 62 new squamous esophageal cancers and 44 new adenocarcinomas of the cardia were identified in this cohort. Proportional hazards models were used to evaluate the relationship of cytological diagnoses and six QFIA variables to subsequent cancer risk. These models showed significant trends for increasing esophageal cancer risk, with increasing values in five of the QFIA variables and with increasing severity of the traditional cytological diagnoses. A comparison of models with only cytology variables versus models with both cytology and QFIA variables indicated that the QFIA provided an important additional predictive value. Persons with both cytological dysplasia and high cellular DNA were 8 times more likely to develop esophageal cancer than were individuals with neither of these conditions. For cardia cancer, associations between QFIA variables or cytological diagnoses and later cancer were more limited. This study suggests that the QFIA variables evaluated here are independent predictors of squamous esophageal cancer and that combining QFIA with traditional cytology can improve prediction of esophageal cancer risk.

Between 1986 and 1991, 29,584 persons took part in a randomized nutritional intervention trial in Linxian, China, an area whose residents had chronically low intakes of several nutrients and high rates of esophageal and gastric cardia cancer as well as stroke. Using a one-half replicate of a 2(4) factorial design, we randomized individuals to one of eight groups which received combinations of four supplements: retinol and zinc (factor A); riboflavin and niacin (factor B); vitamin C and molybdenum (factor C); and beta-carotene, alpha-tocopherol (vitamin E), and selenium (factor D). Deaths that occurred during 5 years of supplementation were ascertained and classified according to cause. At the end of the supplementation period, we measured blood pressure readings and determined the prevalence of hypertension. Participants who received factor D had reductions in total mortality (9%) and total cancer mortality (13%). These individuals also had the largest reduction in stroke mortality (relative risk = 0.91; 95% confidence interval = 0.76-1.07). End-of-trial hypertension, however, was not less prevalent among those receiving factor D. Our findings contrast with the larger reductions in stroke death and hypertension found in a parallel trial of Linxian subjects with esophageal dysplasia who received a multivitamin/mineral supplement, suggesting an effect largely derived from nutrients other than those received in the present study.

Esophageal cancer is the second leading cause of cancer death in China. Esophageal cancer has a very poor prognosis, principally because most tumors are asymptomatic until they are unresectable. Esophageal balloon cytology is an early detection method developed by Chinese scientists to identify resectable early cancers and precursor lesions. Previous studies have reported high sensitivities for detecting esophageal cancer in symptomatic patients. The current report describes several studies evaluating this diagnostic technique in asymptomatic individuals. A comparison of Chinese and U. S. cytological diagnoses of the same esophageal samples showed that the Chinese categories of precancerous neoplasia were more inclusive than the corresponding U. S. categories. Comparisons of both Chinese and U. S. cytological diagnoses with concurrent histological findings showed low (14-36%) sensitivities for the cytological detection of biopsy-proven cancers. Prospective follow-up studies of several screened cohorts showed a consistent progression of risk for developing esophageal cancer with increasing severity of initial cytological diagnosis. These preliminary studies suggest that esophageal balloon cytology is a useful technique that can benefit from additional research to improve its optimal performance.

Researchers have proposed a broad range of molecular, cellular and histological markers as surrogate end-points for cancer (SECs). The effect of an intervention on a 'valid' SEC is concordant with its effect on cancer incidence. The validity of a potential SEC is determined primarily by the extent to which the marker is a necessary event on the causal pathway to cancer. Colorectal adenomatous polyp formation is an example of a reasonably valid SEC because these lesions are obligate precursors of most large bowel malignancies. However, the existence of a plausible major alternative causal pathway--one bypassing the potential SEC--weakens inferences from that marker to cancer. Moreover, unless the pathway to cancer operates nearly exclusively through the SEC, an SEC that is valid for one intervention or exposure may not be valid for another. Metabolic, ecological, observational epidemiological and intervention studies may yield data that are useful in revealing these causal interrelations of intervention (exposure), SEC and cancer. Empirical studies of three questions are pertinent: (1) What is the relation of the SEC to cancer? (2) What is the relation of the intervention (exposure) to the SEC? (3) To what extent does the SEC mediate the relation between the intervention (exposure) and cancer? Data on SEC measurement error are important in ascertaining the extent to which marker results have been attenuated by such error. It is essential to carry out these studies to evaluate potential SECs (such as epithelial cell hyperproliferation) with plausible major alternative pathways to cancer. At the present time, definitive evidence on etiology and prevention will emerge only from studies with cancer end-points or SECs that are, by and large, necessary steps on the causal pathway to malignant disease.

Linxian, China, has some of the highest rates of esophageal cancer in the world. Previous authors have proposed that esophagitis, atrophy, and dysplasia may be precursor lesions of esophageal cancer in such high risk populations.To examine the relationship between squamous esophageal histology and subsequent esophageal cancer in Linxian, the authors prospectively followed 682 participants of a 1987 endoscopic survey for 3.5 years and compared their initial biopsy diagnoses with the occurrence of squamous cell carcinoma during this follow-up period.Squamous cell carcinoma of the esophagus was identified in 52 (7.6%) of the participants during the follow-up period. After adjusting for potential confounding factors, relative risks (95% confidence intervals) for squamous cell carcinoma incidence by initial histologic diagnoses were as follows: normal, 1.0 (reference); basal cell hyperplasia, 2.1 (0.4-9.8); mild dysplasia, 2.2 (0.7-7.5); moderate dysplasia, 15.8 (5.9-42.2); severe dysplasia, 72.6 (29.8-176.9); dysplasia not otherwise specified, 22.9 (6.7-78.0); and carcinoma in situ, 62.5 (24.1-161.9).In this study, moderate dysplasia, severe dysplasia, and carcinoma in situ were the only histologic lesions associated with a significantly increased risk of developing squamous cell carcinoma of the esophagus within 3.5 years after endoscopy. Increasing grades of dysplasia were associated with increasing risk, but severe dysplasia were associated with increasing risk, but severe dysplasia and carcinoma in situ had similar degrees of risk, findings that suggest a continuous spectrum of esophageal intraepithelial neoplasia, without morphologically distinguishable dysplasia and in situ carcinoma. A longer follow-up will be necessary to fully evaluate the less severe diagnostic categories, which may take more than 3.5 years to affect the occurrence of squamous cell carcinoma in this high risk population.

Abnormalities of epithelial proliferation have been proposed as an early step in gastrointestinal carcinogenesis. To determine whether micronutrient supplementation may reduce squamous epithelial proliferation in the esophagus, we evaluated proliferation in subjects participating in a randomized nutrition intervention trial in Linxian, China, where esophageal cancer rates are among the highest in the world. After 30 months of intervention involving daily supplementation with multiple vitamins and minerals, an endoscopic survey was performed and squamous biopsies from 512 subjects were labeled with tritiated thymidine and autoradiographed. Analysis showed no treatment effect on the overall amount of squamous epithelial proliferation measured by the total labeling index. However, a measure of the vertical distribution of labeled cells showed lower values with supplementation: a 14% reduction in all subjects (P = 0.29), and a 29% reduction in nonsmokers (P = 0.03). These results suggest a potential modest benefit for short-term intervention with multiple vitamins and minerals on squamous epithelial cell proliferation of the esophagus in this high-risk population.

In Linxian China, the esophageal/gastric cardia cancer mortality rates are among the highest in the world. There is suspicion that the population's chronic deficiencies of multiple micronutrients are etiologically involved. We conducted two randomized, placebo-controlled nutrition intervention trials to test the effects of vitamin and mineral supplements in lowering the rates of esophageal/gastric cancer. In the first trial, the dysplasia trial, 3318 adults with a cytological diagnosis of esophageal dysplasia received daily supplementation with 26 vitamins and minerals in doses typically 2-3 times the United States Recommended Daily Allowances, or placebos, for 6 years. The second trial, the general population trial, involved 29,584 adults and used a one-half replicate of a 2(4) factorial experimental design which tested the effects of four combinations of nutrients: A, retinol and zinc; B, riboflavin and niacin; C, vitamin C and molybdenum; and D, beta-carotene, vitamin E, and selenium. Doses for these daily supplements ranged from 1 to 2 times the United States Recommended Daily Allowances, and the different vitamin/mineral combinations or placebos were taken for a period of 5.25 years. As part of the general population trial, and end-of-intervention endoscopy survey was carried out in a small (1.3%) sample of subjects to see if supplementation affected the prevalence of dysplasia and early cancer. Herein we review the methods of these trials and the results of the endoscopic survey. Fifteen esophageal and 16 gastric cancers were identified in endoscopic biopsies from the 391 subjects evaluated from two villages, and nearly all were asymptomatic. No significant reductions in the prevalence of esophageal or gastric dysplasia or cancer were seen with any of the four supplement groups. However, the prevalence of gastric cancer among participants receiving retinol and zinc was 62% lower than those not receiving those supplements (P = 0.09), while participants receiving beta-carotene, vitamin E, and selenium had a 42% reduction in esophageal cancer prevalence (0.34). We have reported separately that cancer mortality over the entire 5.25-year period was significantly reduced among those receiving beta-carotene, vitamin E, and selenium. The findings from the overall trial and the endoscopic sample offer a hopeful sign and should encourage additional studies with these agents in larger numbers of subjects.

This paper reports a 15-year follow-up study of 12,693 persons in Linxian, China, who were originally screened by esophageal balloon cytology in 1974. The purpose of the study was to evaluate the ability of esophageal balloon cytology to identify individuals at increased risk for developing esophageal cancer. Age, sex and cytologic diagnoses were obtained from the original 1974 records, and information on vital status, cancer experience and potential confounding risk factors was collected from interviews and medical abstracts in 1989. A total of 1,162 incident cases of esophageal cancer and 993 deaths due to esophageal cancer were identified and used in this analysis. The follow-up study showed that the risk of esophageal cancer incidence and mortality increased in parallel with the presumed severity of the 1974 Chinese cytologic diagnoses. After adjusting for potential confounding factors, the relative risks (and 95% confidence intervals) for esophageal cancer incidence, by cytologic diagnosis, were: normal, 1.00; esophagitis, 1.52 (1.07-2.14); hyperplasia, 1.17 (1.02-1.33); dysplasia 1, 1.53 (1.10-2.14); dysplasia 2, 1.89 (1.47-2.41); and suspicious for cancer, 5.77 (3.79-8.80). These results suggest that esophageal balloon cytology, as performed and interpreted in Linxian in 1974, successfully identified individuals at increased risk for esophageal cancer.

A randomized nutrition intervention trial was conducted among 29,584 adult residents of Linxian, China, to examine the effects of vitamin/mineral supplementation on the occurrence of esophageal/gastric cardia cancer in this high-risk population. A fractional factorial study design allowed evaluations of four different combinations of nutrients: (A) retinol and zinc; (B) riboflavin and niacin; (C) vitamin C and molybdenum; and (D) beta-carotene, vitamin E, and selenium. During the 5.25-year intervention, significant reductions in total mortality, total cancer mortality, and stomach cancer mortality occurred among those receiving beta-carotene, vitamin E, and selenium. At the end of intervention, an endoscopic survey was carried out in a sample of subjects to see if the nutritional supplements had affected the prevalence of clinically silent precancerous lesions and early invasive cancers of the esophagus or stomach. Endoscopy was performed on 391 individuals from two study villages. The prevalences of esophageal and gastric dysplasia and cancer were compared by nutrient factor. Cancer or dysplasia was diagnosed in 15% of the participants. No statistically significant reductions in the prevalence of esophageal or gastric dysplasia or cancer were seen for any of the four vitamin/mineral combinations. The greatest reduction in risk (odds ratio, 0.38; P = 0.09) was seen for the effect of retinol and zinc on the prevalence of gastric cancer. Although no significant protective effects were seen in this endoscopic survey, there was a suggestion that supplementation with retinol and zinc may protect against the development of gastric neoplasia in this high-risk population. Additional studies with larger numbers of endpoints will be needed to further evaluate this possibility.

Linxian, China has some of the highest rates of esophageal/gastric cardia cancer in the world, and epidemiological evidence suggests that chronically low intake of micronutrients may contribute to these high cancer rates. To examine whether supplementation with multiple vitamins and minerals can affect the occurrence of esophageal/gastric cardia cancer in this population, a two-arm randomized nutrition intervention trial was conducted among 3318 Linxian residents with cytological evidence of esophageal dysplasia. During the 6-year intervention, esophageal/gastric cardia cancer mortality was 8% lower among those receiving the active supplements. After 30 and 72 months of intervention, endoscopic surveys were carried out to see if the nutritional supplements had affected the prevalence of clinically silent precancerous lesions and early invasive cancers of the esophagus and stomach. In the first survey, in 1987, 833 subjects were endoscoped; in the second survey, in 1991, 396 subjects were examined. The histological diagnoses from each survey were compared by treatment group. Cancer or dysplasia was diagnosed in 28% of the subjects endoscoped in 1987 and 24% of those examined in 1991. The odds ratio for subjects in the treatment group (versus those in the placebo group) having esophageal or gastric dysplasia or cancer was 0.84 (95% confidence interval, 0.61-1.15) in 1987 and 0.86 (0.54-1.38) in 1991. Although modest protective effects on worst overall diagnosis were seen in the supplemented group in both surveys, none of the results was statistically significant, and the findings must be considered inconclusive.(ABSTRACT TRUNCATED AT 250 WORDS)