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Background: The optimal type of esophagectomy and extent of lymphadenectomy for patients after neoadjuvant chemoradiotherapy (nCRT) for esophageal squamous cell carcinoma remain controversial. We hypothesized that a more radical resection is associated with better survival., Methods: Data of patients who received nCRT followed by resection for esophageal squamous cell carcinoma between 2012 and 2021 were analyzed. Modified en bloc esophagectomy (mEBE) involves total mediastinal lymphadenectomy and resection of all periesophageal node-bearing tissues. Perioperative outcomes and survival rates of mEBE were compared with those of conventional esophagectomy (CE)., Results: A total of 238 patients were included. Compared with CE, mEBE was associated with a longer operative time, higher total number of resected lymph nodes, fewer complications, and less anastomotic leakage; length of stay was similar between the 2 groups. There was no difference in overall survival rates between patients with ypT0 N0 stage in the mEBE and CE groups; however, in patients with non-ypT0 N0 stage in the mEBE and CE groups, the 3-year overall survival rates were 58.5% and 28.5%, respectively (P < .001). On disease-free survival analysis, no difference was observed in patients with ypT0 N0 stage, whereas patients with non-ypT0 N0 stage after nCRT had significantly better disease-free survival after mEBE compared with CE (49.7% vs 27.2%; P = .017)., Conclusions: Survival after mEBE was significantly better than that after CE. The mEBE did not increase postoperative hospital stay and complication rates., (Copyright © 2023 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

The prognostic impact of circumferential resection margin (CRM) in surgically resected esophageal squamous cell carcinoma (ESCC) has been controversial. This investigation assessed the prognostic impact of CRM in surgically resected pathologic T3 ESCC patients with or without neoadjuvant chemoradiotherapy (nCRT). We reviewed consecutive p/yp T3 ESCC patients undergoing esophagectomy from two medical centers between January 2009 and December 2016. The cohort was divided into two groups: upfront esophagectomy (upfront surgery) and nCRT followed by esophagectomy (nCRT + surgery). CRM status was assessed and divided into CRM > 1 mm, 0 < CRM < 1 mm, and tumor at CRM. A total of 217 p/yp T3 ESCC patients undergoing esophagectomy (138 patients in the upfront surgery group and 79 in the nCRT + surgery group) were enrolled. In the upfront surgery group, patients with 0 < CRM < 1 mm showed equivalent overall survival to those with CRM > 1 mm (log-rank P = 0.817) and significantly outlived those with tumor at CRM (log-rank P < 0.001). However, in the nCRT + surgery group, CRM > 1 mm failed to show survival superiority to CRM between 0 and 1 mm or involved by cancer (log-rank P = 0.390). In conclusion, a negative CRM, even though being <1 mm, is adequate for pT3 ESCC patients undergoing upfront esophagectomy. In contrast, the CRM status is less prognostic in ypT3 ESCC patients undergoing nCRT followed by esophagectomy., (© The Author(s) 2020. Published by Oxford University Press on behalf of International Society for Diseases of the Esophagus. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Studies addressing both lymphovascular invasion (LVI) and perineural invasion (PNI) in patients with esophageal squamous cell carcinoma (ESCC) treated with or without neoadjuvant therapy are limited. We aimed to analyze the incidence and prognostic significance of LVI and PNI in patients with thoracic ESCC., Methods: This retrospective study included 520 patients with ESCC: 174 patients after neoadjuvant treatment followed by surgery and 346 after primary esophagectomy, from two medical centers. The relationships between LVI, PNI, and other histological factors were evaluated. The Cox regression model was used for survival analysis., Results: Positive LVI and PNI were noted in 35.6% and 22.4% of patients with residual primary tumor after neoadjuvant treatment and in 39.6% and 24.0% of patients who underwent primary esophagectomy, respectively. In patients with neoadjuvant treatments, the 5-year overall survival rates were 12.7% and 28.3% in patients with positive LVI and negative LVI, respectively (p = 0.001). The 5-year overall survival rates were 6.4% and 29.9% in patients with positive PNI and negative PNI, respectively (p < 0.001). In patients who did not receive neoadjuvant treatment, the 5-year overall survival rates were 28.2% and 61.1% in patients with positive LVI and negative LVI, respectively (p < 0.001). The 5-year overall survival rates were 30.2% and 52.5% in patients with positive PNI and negative PNI (p < 0.001). In subgroup analysis, the presence of PNI was an independent prognostic factor in patients with neoadjuvant treatments, whereas the presence of LVI had more significant prognostic impact in patients with node-negative ESCC after primary esophagectomy., Conclusions: Both LVI and PNI statuses are significant prognostic factors for patients with ESCC. However, the prognostic impact of LVI was majorly in the subgroup of node-negative patients who received primary esophagectomy.

Background: This study aimed to compare survival between standard lobectomy and surgeons' preference sublobar resection among patients with stage I non-small cell lung cancer (NSCLC)., Methods: Medical records of patients undergoing pulmonary resection between 2006 and 2016 were reviewed retrospectively. Differences in disease-free survival (DFS) and DFS-associated factors between patients receiving lobectomy and surgeons' preference sublobar resection were analyzed after 1-1 propensity score-matching (n = 119 per group)., Results: In total, 1064 pathological stage I NSCLC patients were identified, including 816 (76.7%) who underwent lobectomy, 111 (10.4%) who underwent sublobar resection as a compromised procedure (medically unfit), and 137 (12.9%) who underwent surgeons' preference sublobar resection. Rates of five-year DFS for patients undergoing lobectomy, medically unfit, and surgeons' preference sublobar resection were 88.7%, 71.0%, and 93.4%, respectively (P < 0.001). Multivariable Cox regression analysis demonstrated that radiological solid-appearance (adjusted hazard [aHR] = 2.908, P = 0.003), PL2 invasion (aHR = 1.970, P = 0.024), and angiolymphatic invasion (aHR = 2.202, P = 0.005) were significantly associated with lower DFS after adjusting for surgeons' preference sublobar resection (aH = 1.031, P = 0.939). Subgroup analysis of all 403 solid-dominant patients demonstrated equivalent five-year DFS between surgeons' preference sublobar resection and lobectomy (87.7% and 84.1%, respectively, P = 0.721). Propensity-matched analysis showed no differences in five-year DFS in stage I NSCLC patients undergoing lobectomy or surgeons' preference sublobar resection (90.5% vs. 93.4% P = 0.510), and DFS for surgeons' preference sublobar resection remained an insignificant factor (aHR = 0.894, P = 0.834)., Conclusions: Carefully selected patients who have undergone surgeons' preference sublobar resection have comparable outcomes to those receiving lobectomy for stage I NSCLC <3 cm., Key Points: Significant findings of the study Intended sublobar resection has a good outcome. What this study adds Sublobar resection is applicable for stage I NSCLC <3 cm., (© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Tumor progression with chemoresistance and local recurrence is commonly happened during treatment of esophageal squamous cell carcinoma (ESCC). Cancer stem cells (CSC) may respond for tumor progression. However, there are few reports regarding metabolism of esophageal CSCs with clinical correlation. In this work, we demonstrated that ESCC cell lines in spheroid culture display CSC phenotypes, including increased ALDH activity, chemoresistance and tumor initiation, which are dependent on Hsp27 activation. Esophageal CSCs also exhibit reprogrammed metabolic features particularly higher glycolysis and oxidative phosphorylation, which are regulated via the Hsp27-AKT-HK2 pathway. Moreover, HK2 is required for maintenance of CSC phenotypes. Inhibition of CSC metabolism reduces cell growth and tumor formation. Clinically, patients who underwent surgical resection for esophageal cancer, and displayed overexpression of both Hsp27 and HK2, had the worst prognosis of all expression types. In conclusion, stem cells features and aberrant metabolic reprogramming of esophageal CSCs depend on the Hsp27-AKT-HK2 pathway. Targeting Hsp27 and HK2 could be novel therapeutic strategy for treating esophageal cancer and warrants further investigation., (© 2019 UICC.)

Autologous fat grafting procedures have noted a markedly increased frequency, not only for cosmetic purposes, but also for deformities after head and neck cancer and breast cancer surgery. Carcinogenesis is always a major concern in cell therapy-related issues. However, there is no literature discussing this issue in head and neck squamous cell carcinoma patients. To evaluate the interaction of tongue cancer cells and adipose-derived stem cells, we performed a series of in vitro experiments. Our results demonstrated that cisplatin significantly reduced the viabilities of SCC‑25 and CAL‑27 cells in a concentration-dependent manner, but it had low cytotoxicity in cisplatin-resistant CAL‑27 (CAR) cells. There was no significant difference in terms of viability among the SCC‑25, CAL‑27, and CAR cells in the adipose-derived stem cell conditioned medium and control groups. There was also no significant difference in terms of cell migration as determined by wound healing assay of SCC‑25, CAL‑27, and CAR cells between the adipose-derived stem cell conditioned medium treatment and control treatment. Importantly, the adipose-derived stem cell conditioned medium attenuated cisplatin-triggered cell death in the SCC‑25 and CAL‑27 cells. Moreover, adipose-derived stem cell conditioned medium markedly inhibited cisplatin-induced apoptotic cell death (sub‑G1 phase) in the CAL‑27 cells. Western blot analyses indicated that cisplatin-induced reductions in pro‑caspase‑3, pro‑caspase‑9, phospho-BAD, phospho-IGF-1R, phospho-AKT, and phospho-ERK in CAL‑27 cells were reversed by adipose-derived stem cell conditioned medium supplement. Taken together, we provide evidence that adipose-derived stem cell conditioned medium protects CAL‑27 cells from cisplatin-induced cell death, possibly through upregulation of the IGF-1R/AKT/ERK signaling pathway.

Objectives: Although preoperative chemoradiation followed by surgery has been recognized as an efficient strategy for esophageal cancer treatments, several studies demonstrate survival benefits of postoperative chemoradiation for those undergoing upfront resection. The optimal sequence of surgery and chemoradiation remains unclear., Methods: Data of 1647 patients with clinical stage II/III esophageal squamous cell carcinoma (ESCC), including 1245 receiving preoperative chemoradiation followed by esophagectomy (pre-OP CRT group) and 402 receiving primary esophagectomy followed postoperative chemoradiation (post-OP CRT group), were obtained from a nationwide database. Propensity score matching identified 286 well-balanced pairs for outcome comparison., Results: In matched patients, the 3-year overall survival (OS) rates/median survival were not significantly different between the 2 groups (44.0% 3-year OS/26.0 months; 95% confidence interval [CI], 18.9-89 38.0 months) in the pre-OP CRT group, versus 37.9% 3-year OS/23.5 months (95% CI, 18.5-29.9 months) in the post-OP CRT group, P = .3152). The 3-year disease-free survival rates (DFS)/median survival after surgery were 38.7% 3-year DFS/16.7 months (95% CI, 11.9-29.6 months) in the pre-OP CRT group, compared with 30.2% 3-year DFS/10.4 months (95% CI, 7.6-14.0 months) in the post-OP CRT group (P = .0674). In patients who had complete resection, the freedom from recurrence rate at 1 year after surgery was 74.8% and 67.6% in pre-OP CRT and post-OP CRT groups, respectively (P = .2696). In the multivariable analysis, treatment modality (pre- or post-OP CRT) was not a significant factor for OS (P = .258) or disease-free survival (P = .521)., Conclusions: Similar outcome can be achieved with postoperative chemoradiotherapy compared with preoperative chemoradiotherapy in patients with locally advanced ESCC. There is little difference between these 2 strategies., (Copyright © 2017 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.)

Background: Pathological response is an important marker for tumor aggressiveness in patients with esophageal squamous cell carcinoma (ESCC) who receive preoperative chemoradiation followed by esophagectomy. We aim to evaluate the prognostic value of histological factors after trimodality treatments., Methods: 91 patients who received preoperative chemoradiation followed by transthoracic esophagectomy between 2009 and 2014 were included. The pathological examination was reviewed. Overall survival and disease free survival were recorded. Survival analysis was performed using the Cox regression model, and the survival curves were compared by the log-rank test., Results: Survival analysis showed lymphovascular invasion (LVI, hazard ratio [HR]: 2.009, p = 0.029), perineural invasion (PNI, HR: 2.226, p = 0.019), ypN stage (HR: 2.041, p = 0.019), extracapsular invasion (ECI, HR: 2.804, p = 0.003), and incomplete resection (HR: 1.897, p = 0.039) as unfavorable prognostic factors affecting overall survival (OS). Moreover, tumor regression grade (TRG, HR: 1.834, p = 0.038), LVI (HR: 1.975, p = 0.038), ECI (HR: 2.836, p = 0.003), and incomplete resection (HR: 2.254, p = 0.007) adversely affected disease-free survival (DFS). Prognostic classification based on poor primary tumor (TRG2/3, LVI(+), and PNI (+)), lymph node (ypN(+) and ECI(+)), and surgical (incomplete resection) factors significantly predicts OS (p = 0.013) and DFS (p = 0.017). However, the use of postoperative adjuvant therapy was not a significant prognostic factor even in medium- and high-risk ESCC patients who underwent trimodality treatments., Conclusions: Histological factors, including primary tumor, lymph node, and surgical factors has high prognostic value for predicting outcomes in ESCC patients receiving preoperative chemoradiation followed by surgery.

Background: Patterns of recurrence after surgery with postoperative chemoradiotherapy (S-CCRT) or surgery alone in patients with oesophageal squamous cell carcinoma (SCC) may differ. This might influence the nature and timing of subsequent management strategies., Methods: Patients with SCC who had undergone R0 resection were included. Propensity score matching was used to select matched groups. Survival and recurrence were compared by Kaplan-Meier analysis. Univariable and multivariable Cox regression analyses were used to identify prognostic factors for overall and disease-free survival., Results: A total of 1390 patients were included, of whom 1000 had surgery alone and 390 underwent S-CCRT. Propensity score matching yielded 213 well balanced pairs. The 3-year overall survival rate and median survival time in the S-CCRT group were 0·50 and 36·5 (95 per cent c.i. 25·1 to 52·6) months respectively, compared with 0·38 and 22·8 (18·2 to 29·0) months in the surgery-alone group (P = 0·006). The 3-year disease-free survival rate and median disease-free survival time in the S-CCRT group were 0·46 and 30·6 (22·2 to 39·3) months respectively, compared with 0·36 and 17·6 (11·3 to 23·9) months in the surgery-alone group (P = 0·006). The 2-year freedom from locoregional recurrence rate was 0·87 and 0·77 in the S-CCRT and surgery-alone groups respectively (P = 0·003). In multivariable analysis, independent prognostic factors for disease-free survival included age over 56 years, pT3-4 category, pN category, poor differentiation, tumour length exceeding 4·0 cm, and receiving postoperative chemoradiotherapy (hazard ratio 0·62, 95 per cent c.i. 0·47 to 0·81; P < 0·001)., Conclusion: Oesophagectomy with postoperative chemoradiotherapy was associated with longer survival and lower recurrence rates, especially at a locoregional level, compared with surgery alone., (© 2016 BJS Society Ltd Published by John Wiley & Sons Ltd.)

Circulating cell-free DNA (cfDNA) is a potential biomarker for cancer progression but its role is unclear in patients with esophageal squamous cell carcinoma (ESCC) after esophagectomy. We investigated relationships between plasma cfDNA levels and clinicopathological parameters in ESCC patients. Eighty-one ESCC patients who received esophagectomy were enrolled. Plasma samples from these patients and 95 normal controls were collected. DNA copy numbers were measured by real-time quantitative PCR. Subjects were divided into two groups by cfDNA level. Clinicopathological data were collected retrospectively and relationships between cfDNA levels and clinical parameters were evaluated. The cfDNA level in normal controls ranged from 0-4157 copies/mL. The cfDNA level of 96.3% ESCC patients was higher than the cutoff value (2447.26 copies/mL) with a specificity of 94.1%. The mean cfDNA concentration was 5918 copies/mL in lower and 53,311 copies/mL in higher cfDNA groups. No correlations were found between clinicopathological factors and cfDNA levels except for lymphovascular invasion. Higher cfDNA levels were associated with tumor relapse ( p = 0.018). Five-year disease-free survival (DFS) and overall survival (OS) rates were 34.7% and 33.8%, respectively. Patients with higher cfDNA levels had poorer DFS ( p = 0.013). Patients with higher cfDNA levels had poorer OS, but not significantly ( p = 0.164). Circulating cfDNA could be a biomarker for tumor relapse of ESCC with high sensitivity and specificity. Higher cfDNA levels were associated with tumor relapse and shorter DFS after esophagectomy in ESCC patients., Competing Interests: The authors declare no conflict of interest.

Objectives: Rapamycin inhibits products of molecular pathways in esophageal squamous cell carcinoma and limits tumor cell growth by targeting 4E-BP1- and eIF4E-dependent gene translation. In this study, we investigate the influence of 4E-BP1-to-eIF4E ratio on rapamycin response in esophageal squamous cell carcinoma cells, and the underlying mechanism is discussed., Methods: The response to rapamycin treatment was examined in 6 esophageal cancer cell lines. Adjustment of the 4E-BP1/eIF4E ratio was carried out by knockdown or overexpression of 4E-BP1 and eIF4E. The relationship between Egr-1 and 4E-BP1 expression in esophageal cancer cells was also studied., Results: The 4E-BP1/eIF4E ratio was adjusted to evaluate the response to rapamycin treatment in TE1 and TE2 esophageal cancer cells. TE2 cells are sensitized to rapamycin treatment after overexpression of 4E-BP1 or knockdown of eIF4E; TE1 cells become resistant to rapamycin after knockdown of 4E-BP1 or overexpression of eIF4E. These data suggest that the 4E-BP1/eIF4E ratio is a determinant for the response of TE1 and TE2 cells to rapamycin treatment. Egr-1 expression was higher in TE2 cells compared with other esophageal cancer cell lines, and its knockdown increased 4E-BP1 expression in TE2 cells, which became sensitive to rapamycin treatment., Conclusions: The 4E-BP1/eIF4E ratio is a determinant of the response of rapamycin treatment in esophageal cancer cells. Egr-1 can reduce 4E-BP1 gene expression and render esophageal squamous cell carcinoma cells resistant to rapamycin with a relatively low 4E-BP1/eIF4E ratio. Thus, the 4E-BP1/eIF4E ratio may represent a therapeutic index for the prediction of clinical outcome of rapamycin treatment in patients with esophageal squamous cell carcinoma., (Copyright © 2015 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.)

Background: Esophageal squamous cell carcinoma (ESCC) is a lethal malignancy, but only limited molecular markers can predict its prognosis. Recently, a DNA double-strand break repair protein Nijmegen breakage syndrome 1 (NBS1) was reported to induce Snail expression and predict poor prognosis in head and neck cancers. However, the clinicopathologic roles of NBS1 and Snail in ESCC remain unclear., Methods: From January 1995 to September 1999, tissue samples from 153 patients with ESCC who underwent esophagectomies at our institutions were collected and made into tissue core arrays for study. Expression of NBS1 and Snail was examined by immunohistochemical staining. The clinicopathologic data were analyzed, and some additional studies were performed to explore the relationship between NBS1 and Snail., Results: NBS1 overexpression was observed in 28.1% (43/153) of ESCC, whereas Snail overexpression was observed in 26.1% (40/153) of ESCC. Overexpression of NBS1 correlated inversely with nodal status (P = 0.009) and was associated with better overall survival (P = 0.002). On the other hand, overexpression of Snail correlated positively with lymphovascular invasion (P = 0.034) and was associated with worse overall survival (P = 0.036). Meanwhile, NBS1 overexpression correlated inversely with Snail overexpression marginally (P = 0.084). Using the Cox regression analysis, T status (P = 0.006), M status (P = 0.008), and NBS1 overexpression (P = 0.007) were the independent factors of overall survival., Conclusions: Our results showed that NBS1 overexpression was an independent factor of better survival and Snail overexpression predicted a worse survival in ESCC. Combination of NBS1 plus Snail expression status could be used as a predictor of prognosis in ESCC.

Metastasis is the major cause of high mortality in head and neck squamous cell carcinoma (HNSCC), in which HNSCC-derived cancer stem cells (CSCs) may be involved. Several reports have coupled non-viral gene delivery with RNA interference (RNAi) to target specific genes in cancer cells. However, the delivery efficiency of RNAi is limited and remained to be improved. Moreover, the therapeutic effect of non-viral gene delivery approaches on HNSCC-derived CSCs is still uncertain. In this study, we found that EZH2 and Oct4 are upregulated in HNSCC-derived ALDH1+/CD44+ CSC-like cells. Polyurethane-short branch PEI (PU-PEI)-based administration of double-stranded DNA (dsDNA) encoding small interfering RNA (siRNA) against EZH2 and Oct4 (siEZH2/siOct4) led to partial anti-cancer capacity and mild suppression of CSC-like properties. By pre-conjugation of nuclear localization signal (NLS) to siRNA-expressing dsDNA, the anti-cancer efficacy was enhanced due to elevated nuclear delivery. Notably, the NLS-preconjugated siEZH2/siOct4 constructs remarkably repressed epithelial-mesenchymal transdifferentiation (EMT) and radioresistance in ALDH1+/CD44+ CSC-like cells, in which Wnt5A and CyclinD1 may be involved respectively. We furthermore demonstrated that this improved method was capable of reducing tumor growth and metastasis in vivo. Our findings may provide a feasible non-viral gene delivery method to eradicate HNSCC-derived CSCs and improve HNSCC therapy., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: We investigated the association between the increased eukaryotic translation initiation factor 4E (eIF4E) level and MDM2 overexpression in the esophageal cancer tissue and cells., Methods: This was a retrospective study of specimens from esophageal cancer patients treated over a 5-year period in a Taiwan university hospital. The predictor variable was eIF4E level in esophageal tumors and CE48T/VGH and TE6 esophageal carcinoma cell lines. The main outcome variable was MDM2 overexpression. Appropriate descriptive and univariate statistics were computed, and a P value of <0.05 was considered statistically significant., Results: There were two study sample groups. Immunohistochemistry analyses of the first sample group (51 esophageal tumors) revealed that 19 specimens demonstrated MDM2 elevation and 20 specimens had eIF4E overexpression. eIF4E elevation was evidenced by accumulation of the protein in the cytoplasm. There was a significant association between the eIF4E and MDM2 expression (P < 0.001). Western blot analysis and semiquantitative reverse transcriptase-polymerase chain reaction of the second specimen group (20 pairs of tumors and normal tissues) revealed the co-elevation of MDM2 and eIF4E (P = 0.008). There was no increased mdm2 transcript in most of the specimens. Without significant alterations in the mdm2 mRNA level and subcellular distribution, MDM2 protein was upregulated in CE48T/VGH cultured cells expressing ectopic eIF4E. Conversely, reduction of eIF4E by specific siRNA enabled TE6 cells synthesizing reduced amounts of MDM2., Conclusions: Our findings indicate that MDM2 protein levels are strongly associated with and regulated by eIF4E in a posttranscriptional mechanism in esophageal cancer.

Bmi-1, a member of the Polycomb family of transcriptional repressors, is essential for maintaining the self-renewal abilities of adult stem cells. Bmi-1 has been demonstrated to play a role in tumorigenesis in head and neck squamous cell carcinomas (HNSCCs). A recent study has further suggested that ALDH1 may be considered to be a putative marker for HNSCC-derived cancer stem cells. However, the role that Bmi-1 plays in HNSCC-derived ALDH1-positive cells (HNSCC-ALDH1(+)) has yet to be determined. In this study, we demonstrated that HNSCC-ALDH1(+) cells possess tumor initiating properties, are capable of self-renewal, and express higher levels of Bmi-1 as compared to HNSCC-ALDH1(-) cells. To further explore the functional role of Bmi-1 in HNSCC-ALDH1(+) cells, we used a lentiviral vector expressing shRNA to knock down Bmi-1 expression (sh-Bmi-1) in HNSCC-ALDH1(+) cells. Silencing of Bmi-1 significantly enhanced the sensitivity of HNSCC-ALDH1(+) cells to chemoradiation and increased the degree of chemoradiation-mediated apoptosis that occurred. Importantly, knockdown of Bmi-1 increased the effectiveness of radiotherapy and led to the inhibition of tumor growth in nude mice transplanted with HNSCC-ALDH1(+) cells. Kaplan-Meier survival analysis indicated that the mean survival rate of HNSCC-ALDH1(+) tumor-bearing immunocompromised mice treated with radiotherapy was significantly improved by treatment with sh-Bmi-1 as well. In summary, these results suggest that Bmi-1 is a potential target for increasing the sensitivity of HNSCC cancer stem cells to chemoradiotherapy., ((c) 2009 Elsevier Ltd. All rights reserved.)

Purpose: To determine the interrelationships of p53, MDM2, and p14(ARF) protein expression in primary esophageal squamous cell carcinoma (ESCC) and their prognostic value in ESCC., Methods: In total, 119 patients treated for ESCC with esophagectomy were enrolled in this study. Demographic and clinical data including gender, age, depth of tumor invasion, lymph node involvement, and 5-year survival rate were collected by chart review. p53, MDM2, and p14(ARF) were detected immunohistochemically in the resected tumors to evaluate their usefulness as biomarkers of clinical outcome., Results: p53, MDM2, and p14(ARF) were expressed in 61 (51.3%), 34 (28.6%), and 22 (18.5%) of 119 tumor specimens, respectively. Overall, p53 protein expression was positively correlated with MDM2 (P = 0.024) and p14(ARF) expression (P = 0.026). In addition, p14(ARF) expression was most often found in specimens that were positive for both p53 and MDM2. Changes in the p53, MDM2, and p14(ARF) protein levels were not correlated with 5-year survival rate., Conclusions: Expression of p53 protein correlates with increased MDM2 and p14(ARF) protein levels in ESCC. In addition, status of p53 (wild-type versus mutant) rather than expression level of p53, MDM2, or p14(ARF) is likely to be the more critical determinant of clinical outcome.

Aims: Our study investigates the significance of the expression of Wnt pathway proteins including beta-catenin, Axin, beta-transducin-repeat-containing protein (beta-TrCP), and adenomatous polyposis coli (APC) in squamous cell carcinoma of the esophagus (ESCC)., Methods: Immunohistochemical analysis was performed on paraffin-embedded tissue specimens from 128 resected ESCC tumors to detect the expression of beta-catenin, Axin, beta-TrCP, and APC. Correlation between immunoexpression, clinicopathological parameters, and patient survival was analyzed., Results: Increased beta-catenin expression was noted in 22 (18.2%) of 121 tumor specimens. Reduced expression of Axin, beta-TrCP, and APC was observed in 57 (46.0%) of 124, 29 (24.4%) of 119, and 54 (48.2%) of 119 specimens, respectively. No correlation was found among these protein expressions. Axin protein expression was inversely correlated with tumor invasion depth (P = 0.033). Reduced Axin protein expression, lymph node involvement, and distant metastasis were significant negative predictors for overall survival and disease-free survival on univariate analysis. In multivariate analysis, reduced Axin expression remained a significant prognostic factor for patients with ESCC (P = 0.005)., Conclusions: Reduced Axin expression was observed in 46% of ESCC tumor specimens and was associated with poor prognosis in patients with ESCC. Further study is mandatory to elucidate the underlying mechanism responsible for loss of Axin expression and the role of Axin in ESCC tumorigenesis.

A HIC1-SIRT1-p53 circular loop in which hypermethylation in cancer 1 (HIC1) represses the transcription of SIRT1 that deacetylates and inactivates p53 thus leading to HIC1 inactivation has been identified in cell and animal models. However, the alteration and prognostic effects of HIC1-SIRT1-p53 circular loop have never been demonstrated in human cancer patients. We examine the HIC1-SIRT1-p53 alterations in 118 lung cancer patients to define their etiological roles in tumorigenesis. We found that patients with lung squamous cell carcinoma with low p53 acetylation and SIRT1 expression mostly showed low HIC1 expression, confirming deregulation of HIC1-SIRT1-p53 circular loop in the clinical model. Interestingly, the expression of deleted in breast cancer 1 (DBC1), which blocks the interaction between SIRT1 deacetylase and p53, led to acetylated p53 in patients with lung adenocarcinoma. However, epigenetic alteration of HIC1 promoter by posttranslational modifications of histones and promoter hypermethylation favoring the compacted chromatin production attenuated the transcriptional induction by acetylated p53. Importantly, lung cancer patients with altered HIC1-SIRT1-p53 circular regulation showed poor prognosis. Our data show the first valid clinical evidence of the deregulation of HIC1-SIRT1-p53 loop in lung tumorigenesis and prognosis. Distinct status of p53 acetylation/deacetylation and HIC1 alteration mechanism result from different SIRT1-DBC1 control and epigenetic alteration in lung squamous cell carcinoma and lung adenocarcinoma.

Aldehyde dehydrogenase 1 (ALDH1) has been considered to be a marker for cancer stem cells. However, the role of ALDH1 in head and neck squamous cell carcinoma (HNSCC) has yet to be determined. In this study, we isolated ALDH1-positive cells from HNSCC patients and showed that these HNSCC-ALDH1+ cells displayed radioresistance and represented a reservoir for generating tumors. Based on microarray findings, the results of Western blotting and immunofluorescent assays further confirmed that ALDH1+-lineage cells showed evidence of having epithelial-mesenchymal transition (EMT) shifting and endogenously co-expressed Snail. Furthermore, the knockdown of Snail expression significantly decreased the expression of ALDH1, inhibited cancer stem-like properties, and blocked the tumorigenic abilities of CD44+CD24(-)ALDH1+ cells. Finally, in a xenotransplanted tumorigenicity study, we confirmed that the treatment effect of chemoradiotherapy for ALDH1+ could be improved by Snail siRNA. In summary, it is likely that ALDH1 is a specific marker for the cancer stem-like cells of HNSCC.

Prostaglandin E2 (PGE2), a major cyclooxygenase-2 (COX-2) product, has been shown to affect numerous tumorigenic processes. PGE2 acts through G-protein-coupled receptors designated as EPs. Recently it has been documented that PGE2 promotes colon cancer cell growth via EP2. However, the expression and the prognostic role of EP2 in esophageal squamous cell carcinoma (ESCC) remained unknown. From January 1995 to January 2001, tissue samples from 226 patients with ESCC who underwent esophagectomies at our institutions were collected and made into tissue core arrays for study. EP2 expression was examined by immunohistochemical staining and confirmed by Western blot. The clinicopathologic data were then analyzed. EP2 overexpression was observed in 43.4% (98/226) of ESCC. Overexpression of EP2 correlated positively with depth of tumor invasion (T status) (P = 0.016) and was associated with worse overall survival (P = 0.047). In patients without regional or distant lymph node metastasis (N0 or M0), EP2 overexpression was associated with worse overall survival (P = 0.033 and P = 0.003, respectively). Using Cox regression analysis, T status, N status, and M status were the independent factors of overall survival, but EP2 expression was not. However, when focusing on patients with T1-3N0M0 status, EP2 expression became an independent factor of overall survival (P = 0.048). Our results show that EP2 overexpression was associated with worse prognosis, and correlated positively with T status in ESCC. Meanwhile, among those patients at earlier stages, EP2 overexpression significantly disclosed patients at high risks for poor prognosis.

Patterns of global histone modifications have been recently suggested as outcome predictors in cancer patients. To date, there has been no report on the prognostic significance of global histone modifications in esophageal squamous cell carcinoma. We investigated the role of global histone modification as outcome predictor in patients undergoing esophagectomy for esophageal squamous cell carcinoma. A retrospective clinicopathologic analysis was undertaken of 97 patients with esophageal squamous cell carcinoma who recovered from esophagectomy. Immunohistochemical expression of five histone modification markers, acetylated histone 3 lysine 18 (H3K18Ac), acetylated histone 4 lysine 12 (H4K12Ac), dimethylated histone 4 arginine 3 (H4R3diMe), dimethylated histone 3 lysine 4 (H3 K4diMe), and trimethylated histone 3 lysine 27 (H3K27triMe) was assessed in paraffin-embedded tumor samples. Results were analyzed in relation to patients' clinicopathologic parameters. There was a positive relationship between tumor differentiation and H3K18Ac (P<0.001), H4R3diMe (P=0.003), and H3K27triMe (P<0.001). Expression of H3K27triMe correlated positively with nodal (N) status (P=0.012) and stage (P=0.025). Univariate analysis showed that better survival in patients with low expression of H3K18Ac (P=0.038) and H3K27triMe (P=0.003). Multivariate analysis showed that nodal status, metastasis status (M), and expression of H3K27triMe predicted survival independently (P<0.001, P=0.016, and 0.048, respectively). Low expression of H3K18Ac and H3K27triMe correlated with better prognosis of patients with esophageal squamous cell carcinoma, especially for those of early stages. We hypothesize that expression of H3K27triMe may be considered as a significant survival predictor for patients with esophageal squamous cell carcinoma.

Objectives: The aim of this study was to evaluate, by immunohistochemical analysis, the protein expression of beta-catenin and p53 in resected esophageal squamous cell carcinoma specimens. The clinical relevance and prognostic significance of the expression of these proteins were also analyzed., Methods: Immunohistochemistry was performed on paraffin-embedded tissue specimens from 68 resected esophageal squamous cell carcinoma tumor specimens to detect the expression of beta-catenin and p53. The correlation between the results of immunoexpression and the clinicopathologic parameters and patient survival was processed statistically., Results: Reduced membranous beta-catenin expression was noted in 43 (63.2%) of 68 tumor specimens. Increased expression of p53 was observed in 43 (63.2%) of 68 specimens. Reduced membranous beta-catenin protein expression was associated with the presence of distant metastasis (P = .006). Patients with reduced membranous beta-catenin expression had a worse prognosis than patients with normal membranous beta-catenin expression (P = .005). Patients with combined increased p53 and reduced membranous beta-catenin protein expression had the worst prognosis (P = .012). In a multivariate survival analysis, reduced membranous beta-catenin expression and nodal involvement were independent prognostic factors (P = .004 and .019, respectively)., Conclusions: Immunohistochemical analysis revealed that reduced membranous beta-catenin protein expression was associated with the presence of distant metastasis and a poor prognosis in patients with esophageal squamous cell carcinoma. Combined increased p53 and reduced membranous beta-catenin protein expression indicated a very poor prognosis in patients with esophageal squamous cell carcinoma. Further investigation is needed to understand the roles of beta-catenin and p53 in the tumorigenesis and metastasis of esophageal squamous cell carcinoma.

Background: Lung squamous cell carcinoma (SCC) cells frequently exhibit markers of chromosome instability such as high fractional allelic loss (FAL). We postulated that alterations in the p53 damage responsive gene and in the double-strand break (DSB) repair genes, BRCA1 and XRCC5, are involved in patients with high FAL. In addition, chromosomal deletion analysis enables the delineation of the likely locations of tumor suppressor genes (TSG) and could provide molecular markers for disease classification., Patients and Methods: To define the minimal deletion regions (MDRs), we used 92 microsatellites spanning 29 regions identified in our previous genome-wide chromosomal deletion study in 36 lung SCC patients to verify the maximal contiguous deletion loci., Results: Eight MDRs at 2q35, 3p14.1-3p14.3, 3p22.2-p23, 3p25.3-3p26.3, 5q35.1-q35.2, 9p23-p24.1, 13q14.11-q14.2, and 17p13.1-p13.2 were found in lung SCC. The candidate genes GAS7 and OVCA2 in the MDR17pA (17p13.1-p13.2) were further examined for mRNA expression. Low expression of the GAS7 gene in 57% of patients analyzed suggested its importance in lung SCC tumorigenesis. In addition, we found a panel of five microsatellites (D3S1766, D4S2397, D4S2361, D13S175, and D17S974), which can be used as prognostic biomarkers in lung SCC. Furthermore, alteration in more than two genes in DSB repair-related pathways was more apparent in high FAL patients., Conclusions: Our results provide biomarkers that may be used for monitoring tumor progression and for positional cloning of new TSGs. Importantly, our data show direct evidence that alterations in DSB repair-related pathways are involved in the genomic instability verified by intensive microsatellites of lung SCC.

Hypoxia-inducible factor (HIF)-1alpha and vascular endothelial growth factor (VEGF) are important angiogenic factors in human cancers. Relative to VEGF-C, prognostic significance of VEGF-D expression and its association with HIF-1alpha expression remain elusive in esophageal squamous cell cancer (ESCC). We studied expression of HIF-1alpha and VEGF-D using immunohistochemistry in 85 resected ESCC specimens and correlated results with patients' clinicopathologic parameters and survival. Association between expression of HIF-1alpha and VEGF-D was investigated using a concordance analysis. High expression of HIF-1alpha and VEGF-D was observed in 52 (61.2%) and 56 (65.9%) patients, respectively. HIF-1alpha expression correlated well with tumor stage (P = 0.041), whereas VEGF-D expression correlated with tumor stage (P = 0.027) and N status (P = 0.019). Groups of high HIF-1alpha and VEGF-D showed worse survivals than those of low expression (P = 0.002 and 0.001, respectively). Multivariate analysis supported expression of HIF-1alpha and VEGF-D as significant survival predictors (P = 0.044 and 0.035, respectively). A concordance rate of 69.5% was observed between expression of HIF-1alpha and VEGF-D. In conclusion, protein expression of HIF-1alpha and VEGF-D are independent prognostic predictors. An association between expression of HIF-1alpha and VEGF-D suggests that these two angiogenic factors are essential in progression of ESCC.

Background: Promoter methylation inactivates expression of some important tumor suppressor genes and may be associated with histone modification. The fragile histidine triad (FHIT) gene is considered a tumor suppressor gene in different human epithelial cancers. We investigated whether FHIT methylation is associated with aberrant expression of Fhit protein and acetylated histone, and whether aberrant expression of Fhit protein and acetylated histone are related to prognosis after resection for esophageal squamous cell cancer., Methods: We analyzed FHIT methylation using methylation-specific polymerase chain reaction and Fhit protein and acetylated histone H4 using immunohistochemistry in 60 resected tumor specimens. Concordance analysis was performed between FHIT methylation and expression of Fhit as well as H4., Results: The FHIT methylation was observed in 33(55%) specimens, and the aberrant expression of Fhit and acetylated H4 was found in 42 (70%) and 40 (67%) specimens, respectively. Expression of aberrant Fhit correlated positively with tumor staging (p < 0.017) and nodal involvement (p = 0.004). Aberrant expression of acetylated H4 correlated positively with tumor staging (p < 0.001), nodal involvement (p < 0.001), and metastasis (p = 0.004). Concordance rates of 75% and 81.7% were present between promoter methylation of FHIT and expression of Fhit (p = 0.035) and acetylated H4 (p = 0.02)., Conclusions: Aberrant expression of Fhit and acetylated histone H4 are frequently associated with the presence of esophageal squamous cell carcinoma, and they are potential prognostic predictors for patients after resection of the tumor.

Objective: Aberrant expression of mismatch repair genes, such as human mutL homolog 1 (hMLH1) and human mutS homolog 2 (hMSH2), are common in some human cancers, and promoter methylation is believed to inactivate expression of hMLH1. We investigated whether promoter methylation is involved in loss of hMLH1 protein and whether aberrant expression of hMLH1 and hMSH2 protein is related to prognosis after resection for esophageal squamous cell cancer., Methods: We analyzed promoter methylation of hMLH1 using methylation-specific polymerase chain reaction and hMLH1 and hMSH2 protein by using immunohistochemistry in 60 resected tumor specimens. The Pearson chi2 test was used to compare expression of hMLH1 and hMSH2 protein among patients with different clinicopathologic parameters. Concordance analysis was performed between hMLH1 methylation and its protein expression., Results: Loss of hMLH1 and hMSH2 protein was found in 43 (72%) and 39 (65%, P = .06) of 60 resected specimens, respectively. hMLH1 protein correlated well with tumor staging (P < .0001), depth of tumor invasion (P = .008), and nodal involvement (P < .0001) but not with distant metastasis, whereas hMSH2 did not show correlation with any of these parameters. A concordance rate of 83.3% was present between expression of hMLH1 protein and its promoter methylation (P < .001)., Conclusions: Aberrant expression of hMLH1 and hMSH2 protein is frequently associated with the presence of esophageal squamous cell carcinoma, and expression of hMLH1 protein is a better prognostic predictor than is expression of hMSH2 protein. Promoter methylation is one of the mechanisms responsible for loss of hMLH1 protein in esophageal squamous cell cancer.

Background: Squamous cell carcinoma of the esophagus (ESCC) is one of the ten most common cancers in Taiwan. Surgical resection is the mainstay of the treatment. Young patients with ESCC are reported to have poor surgical outcome. This retrospective and comparative study is to evaluate the clinicopathologic difference and survival between young and elderly patients., Methods: From January 1985 to December 2000, totally 797 patients with esophageal cancer received esophagectomy with reconstruction at our institution. Among them, 12 patients who received neoadjuvant concurrent chemo-radiotherapy followed by esophagectomy were excluded from this study. The other 785 patients were divided into two groups. Group I included 67 patients aged 50 or younger (8.5%) and group II included 718 patients aged over 50 years (91.5%). The clinical data were collected by chart review and follow-up data were collected through outpatient department record or telephone contact. These two groups were analyzed and compared regarding sex ratio, exposure to tobacco or alcohol, surgical mortality rate and 5-year survival rate., Results: There was no difference in gender distribution between young and old patients groups. More patients in the young patients group had the habits of alcohol and tobacco consumption than in the elder patients group. The surgical (30-day) mortality was 2.98% in the young patient group, and 5.43% in the elder group. The 5-year survival rate was better in young patients group than in elder patients group but there was no statistical significance. Survival in young patients group was significantly related to TNM staging system., Conclusions: More young patients with ESCC had the habit of smoking and drinking. There was no statistical difference between the groups in clinical variables, surgical mortality and survival. The survival rate was significantly related to the TNM tumor stage.