TP53 Targeted Deep Sequencing of Cell-Free DNA in Esophageal Squamous Cell Carcinoma Using Low-Quality Serum: Concordance with Tumor Mutation.

Circulating cell-free DNA (cfDNA) is emerging as a potential tumor biomarker. CfDNA-based biomarkers may be applicable in tumors without an available non-invasive screening method among at-risk populations. Esophageal squamous cell carcinoma (ESCC) and residents of the Asian cancer belt are examples of those malignancies and populations. Previous epidemiological studies using cfDNA have pointed to the need for high volumes of good quality plasma (i.e., >1 mL plasma with 0 or 1 cycles of freeze-thaw) rather than archival serum, which is often the main available source of cfDNA in retrospective studies. Here, we have investigated the concordance of TP53 mutations in tumor tissue and cfDNA extracted from archival serum left-over from 42 cases and 39 matched controls (age, gender, residence) in a high-risk area of Northern Iran (Golestan). Deep sequencing of TP53 coding regions was complemented with a specialized variant caller (Needlestack). Overall, 23% to 31% of mutations were concordantly detected in tumor and serum cfDNA (based on two false discovery rate thresholds). Concordance was positively correlated with high cfDNA concentration, smoking history (p-value = 0.02) and mutations with a high potential of neoantigen formation (OR; 95%CI = 1.9 (1.11-3.29)), suggesting that tumor DNA release in the bloodstream might reflect the effects of immune and inflammatory context on tumor cell turnover. We identified TP53 mutations in five controls, one of whom was subsequently diagnosed with ESCC. Overall, the results showed that cfDNA mutations can be reliably identified by deep sequencing of archival serum, with a rate of success comparable to plasma. Nonetheless, 70% non-identifiable mutations among cancer patients and 12% mutation detection in controls are the main challenges in applying cfDNA to detect tumor-related variants when blindly targeting whole coding regions of the TP53 gene in ESCC.

Predicting the outcome of asymptomatic univesicular liver hydatids: diagnostic accuracy of unenhanced CT.

OBJECTIVES: To establish the role of unenhanced CT in predicting the outcome of liver hydatid cysts. We sought to determine whether the presence of laminated membrane detachment (LMD) or pericyst degenerative changes (PDCs) detected on CT were reliable signs for predicting a favorable outcome of liver hydatids. METHODS: In a 20-year-long study, we prospectively followed changes occurred in CT of 106 cysts of 98 patients with incidentally discovered asymptomatic univesicular liver hydatids who accepted to enter a watch-and-wait program. An "unfavorable" outcome was defined as the occurrence of a complication (most commonly, cyst fistula or infection) or increase in the cyst size during the follow-up; otherwise, the outcome was considered "favorable." The parameters derived from a binary logistic regression analysis (with the outcome taken as the dependent variable), after appropriate transformation of the independent variables (presence of LMD or PDCs on CT), were used to calculate the sensitivity, specificity, positive and negative likelihood ratios, and positive and negative predictive values of the presence of either the abovementioned CT findings for the prediction of a favorable outcome. RESULTS: The presence of LMD or PDCs had a high specificity (88%) and positive predictive value (96%) for a favorable outcome; they had high false-negative rates. CONCLUSIONS: The presence of either LMD or PDCs on unenhanced CT, in incidentally discovered asymptomatic univesicular liver hydatids, was associated with a high probability of a favorable outcome. Their absence does not rule out a favorable outcome. KEY POINTS: • Computed tomography can be used for predicting the outcome of those with incidentally discovered univesicular liver hydatids. • The presence of laminated membrane detachment and/or pericyst degenerative changes is associated with a favorable outcome. • Their absence does not necessarily indicate an unfavorable outcome.

The Non-operative Management of Asymptomatic Liver Hydatids: Ending Echinococcophobia.

BACKGROUND: Cystic echinococcosis is common in many countries and involves the liver in 75% of cases. The current belief is that all incidentally discovered asymptomatic liver hydatids must be subjected to some types of interventional treatments pre-emptively for fear of some presumably impending complications. We conducted this study to assess the prognosis of patients with liver hydatids without any surgical interventions. Furthermore, we tried to predict the outcome of the cyst using two radiological signs. METHODS: Of 434 asymptomatic patients with liver cysts who referred to our center, 297 were found eligible to be monitored at 1-3-year intervals by unenhanced computed tomography (CT) over a 20-year period. The patients were given the option of either having their cysts removed by open operation (the surgical group) or entering an open-ended "watch-and-wait" program (the surveillance group). Using unenhanced CT, cyst size, laminated membrane detachment (LMD), and pericyst degenerative changes (PDCs) were evaluated every 1-3 years. If a complication happened or the cyst size increased during the follow-up period, the cyst outcome was considered "unfavorable"; otherwise, it was considered "favorable." RESULTS: Forty-three patients with 56 liver cysts were operated upon without morbidity or mortality. One hundred and twenty-seven patients with 137 cysts were followed. After a median follow-up of 6 (IQR 3 to 10) years, 111 (81.0%) had favorable outcome and 26 (19.0%) cysts had an unfavorable outcome. The size of 69 (50.4%) cysts with favorable outcome was not changed. It was decreased by a mean of 35.7% (SD 17.5%) in 34 (24.8%) cysts; 8 (5.8%) cysts disappeared. Of 26 cysts with unfavorable outcome, 8 (5.8%) developed complications (6 fistulas and 2 secondary infections) after a median follow-up of 6.5 (IQR 2.3 to 12.8) years. The size of the remaining 18 cysts increased by a mean of 42.2% (SD 29.5%). Albendazole administered at a dose of 400 mg, po, bid for 3 years, had a significant effect on hydatids. After adjustment for cyst size at diagnosis and patient's age and sex, logistic regression analysis showed that being univesicular (OR 6.00, 95% CI 1.32 to 27.29), having LMD (OR 5.51, 95% CI 1.03 to 29.43), and the presence of PDCs (OR 4.25, 95% CI 1.36 to 13.30) were independent predictors of a favorable outcome for a cyst. CONCLUSIONS: More than 80% of asymptomatic liver hydatids have a favorable outcome without pre-emptive surgical interventions. The presence of LMD at any stage and/or PDCs of any grade justifiably exempts asymptomatic univesicular liver hydatids from any treatments. Those without LMD or PDCs can be effectively managed with long-term albendazole, artificially inducing larval involution.

The gastro-esophageal malignancies in Northern Iran research project: impact on the health research and health care systems in Iran.

BACKGROUND: The Gastro-Esophageal Malignancies in Northern Iran (GEMINI) research project is an example of recent progress in health research in Iran. The original aim of this project was to identify etiologic factors and prevention measures for upper gastrointestinal cancers in Northern provinces of Iran, but its achievements have gone much beyond this initial goal. METHODS: GEMINI consists of several projects including cancer registries, pilot studies, case-control studies, and the Golestan Cohort Study. GEMINI has been conducted through extensive collaborations between the Digestive Disease Research Center of Tehran University of Medical Sciences with other domestic and international health organizations. The achievements of GEMINI include producing new knowledge, introducing new research methods, developing and expanding health research and health care infrastructures, investing in human resources, and increasing the awareness and knowledge of policy makers and officials at all levels about the importance of chronic diseases in Iran's health priorities. CONCLUSION: The success of GEMINI reveals the feasibility of large-scale health research studies in developing countries and serves as a successful model not only for health research in Iran, but also for similar research studies in other developing nations.

Epidermal growth factor receptor (EGFR) mutations and expression in squamous cell carcinoma of the esophagus in central Asia.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) shows geographic variations in incidence, with high incidences (>50/105 person-years) in central Asia, including North Eastern Iran (Golestan) and Northern India (Kashmir). In contrast to Western countries, smoking does not appear to be a significant risk factor for ESCC in central Asia. In lung adenocarcinoma, activating mutations in the gene encoding epidermal growth factor receptor (EGFR) are frequent in tumors of never smokers of Asian origin, predicting therapeutic sensitivity to Egfr-targeting drugs. METHODS: In this study 152 cases of histologically confirmed ESCC from Iran (Tehran and Golestan Province) and North India (Kashmir Valley) have been analyzed for EGFR mutation by direct sequencing of exons 18-21. Egfr protein expression was evaluated by immunohistochemistry in 34 samples from Tehran and HER2 mutations were analyzed in 54 cases from Kashmir. RESULTS: A total of 14 (9.2%) EGFR variations were detected, including seven variations in exons. Among those, four (2.6%) were already documented in lung cancers, two were reported as polymorphisms and one was a potentially new activating mutation. All but one variation in introns were previously identified as polymorphisms. Over-expression of Egfr was detected in 22/34 (65%) of tested cases whereas no HER2 mutation was found in 54 cases from Kashmir. CONCLUSION: Overall, EGFR mutations appear to be a rare event in ESCC in high incidence areas of central Asia, although a very small proportion of cases may harbor mutations predicting sensitivity to anti-Egfr drugs.

A new technical approach to cancers of the cervical esophagus.

BACKGROUND: The aim of this study was to assess the possibility of a primary end-to-end pharyngoesophageal anastomosis after standard tumor resection of the cervical esophagus by acute flexion of the neck. METHODS: A total of 34 consecutive patients with primary cervical esophageal cancer, none having received prior radio- or chemotherapy, were treated by two methods based on intraoperative findings. In 18 patients, reconstruction after esophageal resection was carried out by the standard gastric pull-through technique (control group). In 16 patients, acute flexion of the neck after tumor resection allowed for reconstruction by primary end-to-end pharyngoesophagostomy (experimental group). RESULTS: There was no operative mortality in either group. The mean operative time for the experimental group was about 50 minutes less compared to the control group. Self-limited postoperative anastomotic leakage in the neck was twice as common in the experimental group. Postoperative dysphagia was about three times as common in the experimental group [5 patients (31%)] compared to the control group [2 patients (11%)]. CONCLUSION: In selected cases, segmental resection of primary cervical esophageal cancers reconstructed by end-to-end pharyngoesophagostomy is technically feasible by bending the neck acutely forward during anastomosis and maintaining it in the flexed position during a postoperative period of about 7 days. The advantages are reduced scope and duration of the operation. The downside is doubling of the frequency of postoperative cervical leakage.

Extremely high Tp53 mutation load in esophageal squamous cell carcinoma in Golestan Province, Iran.

BACKGROUND: Golestan Province in northeastern Iran has one of the highest incidences of esophageal squamous cell carcinoma (ESCC) in the world with rates over 50 per 100,000 person-years in both sexes. We have analyzed TP53 mutation patterns in tumors from this high-risk geographic area in search of clues to the mutagenic processes involved in causing ESCC. METHODOLOGY/PRINCIPAL FINDINGS: Biopsies of 119 confirmed ESCC tumor tissue from subjects enrolled in a case-control study conducted in Golestan Province were analyzed by direct sequencing of TP53 exons 2 through 11. Immunohistochemical staining for p53 was carried out using two monoclonal antibodies, DO7 and 1801. A total of 120 TP53 mutations were detected in 107/119 cases (89.9%), including 11 patients with double or triple mutations. The mutation pattern was heterogeneous with infrequent mutations at common TP53 "hotspots" but frequent transversions potentially attributable to environmental carcinogens forming bulky DNA adducts, including 40% at bases known as site of mutagenesis by polycyclic aromatic hydrocarbons (PAHs). Mutations showed different patterns according to the reported temperature of tea consumption, but no variation was observed in relation to ethnicity, tobacco or opium use, and alcoholic beverage consumption or urban versus rural residence. CONCLUSION/SIGNIFICANCE: ESCC tumors in people from Golestan Province show the highest rate of TP53 mutations ever reported in any cancer anywhere. The heterogeneous mutation pattern is highly suggestive of a causative role for multiple environmental carcinogens, including PAHs. The temperature and composition of tea may also influence mutagenesis.

Tea drinking habits and oesophageal cancer in a high risk area in northern Iran: population based case-control study.

OBJECTIVE: To investigate the association between tea drinking habits in Golestan province, northern Iran, and risk of oesophageal squamous cell carcinoma. DESIGN: Population based case-control study. In addition, patterns of tea drinking and temperature at which tea was drunk were measured among healthy participants in a cohort study. SETTING: Golestan province, northern Iran, an area with a high incidence of oesophageal squamous cell carcinoma. PARTICIPANTS: 300 histologically proved cases of oesophageal squamous cell carcinoma and 571 matched neighbourhood controls in the case-control study and 48 582 participants in the cohort study. MAIN OUTCOME MEASURE: Odds ratio of oesophageal squamous cell carcinoma associated with drinking hot tea. RESULTS: Nearly all (98%) of the cohort participants drank black tea regularly, with a mean volume consumed of over one litre a day. 39.0% of participants drank their tea at temperatures less than 60 degrees C, 38.9% at 60-64 degrees C, and 22.0% at 65 degrees C or higher. A moderate agreement was found between reported tea drinking temperature and actual temperature measurements (weighted kappa 0.49). The results of the case-control study showed that compared with drinking lukewarm or warm tea, drinking hot tea (odds ratio 2.07, 95% confidence interval 1.28 to 3.35) or very hot tea (8.16, 3.93 to 16.9) was associated with an increased risk of oesophageal cancer. Likewise, compared with drinking tea four or more minutes after being poured, drinking tea 2-3 minutes after pouring (2.49, 1.62 to 3.83) or less than two minutes after pouring (5.41, 2.63 to 11.1) was associated with a significantly increased risk. A strong agreement was found between responses to the questions on temperature at which tea was drunk and interval from tea being poured to being drunk (weighted kappa 0.68). CONCLUSION: Drinking hot tea, a habit common in Golestan province, was strongly associated with a higher risk of oesophageal cancer.

Esophageal cancer in Northeastern Iran: a review.

Golestan Province in northeastern Iran has one of the highest rates of esophageal squamous cell cancer in the world. This article reviews the studies conducted on esophageal squamous cell cancer in this area and summarizes the data on epidemiologic patterns, incidence trends, and etiology of esophageal squamous cell cancer in this province.

Flexing the neck relieves tension on cervical esophageal anastomosis.

BACKGROUND: The conventional method of bridging anatomic defects of the upper digestive tract in the neck is by tissue transfer--either gastric or colon pull-through, free jejunal graft, or full-thickness skin flaps. An alternative way of closing such defects is to flex the neck. This moves the remnant proximal esophagus or pharynx a considerable distance downwards--a standard tension-releasing maneuver in tracheal resection and reconstruction. METHODS: Neck flexion was used in 7 patients grouped into three separate surgical conditions: A) in two patients after esophagectomy, where the pulled-up stomach would not reach the remnant proximal esophagus or the pharynx; B) in three patients where the defect after removal of the diseased portion of the cervical esophagus measured 4.5, 5.0, and 8.0 cm, respectively; and C) in 2 patients with 4.5- and 1.5-cm long circumferential postoperative esophageal strictures managed by Heineke-Miculicz repair. RESULTS: No postoperative cervical fistulas were seen. One patient, whose 8-cm long cervical esophageal defect had been closed by end-to-end anastomosis, developed a stricture. CONCLUSION: In special situations, flexing the neck allows for safe anastomosis or closure of esophageal defects in the neck, obviating the need for tissue transfer.