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Esophageal squamous cell carcinoma (ESCC) is one of the deadliest cancers, and it requires novel treatment approaches and effective drugs. In the present study, we found that treatment with plumbagin, a natural compound, reduced proliferation and survival of the KYSE150 and KYSE450 ESCC cell lines in a dose-dependent manner in vitro. The drug also effectively inhibited the viability of primary ESCC cells from fresh biopsy specimens. Furthermore, plumbagin-induced mitotic arrest and massive apoptosis in ESCC cells. Notably, the drug significantly suppressed the colony formation capacity of ESCC cells in vitro and the growth of KYSE150 xenograft tumors in vivo. At the molecular level, we found that exposure to plumbagin decreased both polo-like kinase 1 (PLK1) and phosphorylated protein kinase B (p-AKT) expression in both ESCC cell lines. Enforced PLK1 expression in ESCC cells not only markedly rescued cells from plumbagin-induced apoptosis and proliferation inhibition but also restored the impaired AKT activity. Furthermore, signal transducer and activator of transcription 3 (STAT3), a transcription factor of PLK1, was also inactivated in plumbagin-treated ESCC cells; however, the overexpression of a constitutively activated STAT3 mutant, STAT3C, reinstated the plumbagin-elicited blockade of PLK1-AKT signaling in ESCC cells. Taken together, these findings indicate that plumbagin inhibits proliferation and potentiates apoptosis in human ESCC cells in vitro and in vivo. Plumbagin may exert these antitumor effects by abrogating STAT3-PLK1-AKT signaling, which suggests that plumbagin may be a novel, promising anticancer agent for the treatment of ESCC.

Background: The purpose of the present paper was to study the expression of cyclooxygenase-2 (COX-2) in normal squamous epithelium, squamous dysplasia and squamous cell carcinoma (SCC) of the esophagus, to elucidate the role of COX-2 in esophageal carcinogenesis, and to evaluate the in vitro effect and mechanism of a COX-2 inhibitor, NS-398, in inducing growth inhibition and apoptosis of human esophageal cancer cells., Methods: Biopsy specimens of esophageal dysplasia (n = 21), and surgical resections of SCC (n = 37) were compared with normal esophagus (n = 37) and analyzed by RT-PCR. Human esophageal cells were used for the study. Anti-proliferative effect was measured by MTT, apoptosis was determined by DNA fragmentation assay., Results: Marked COX-2 expression was shown in SCC and esophageal squamous dysplasia, and no marked COX-2 expression was observed in the normal squamous epithelium, respectively. NS-398 could inhibit esophageal cells growth in a dose-dependent manner, induce apoptosis, and elevate caspase-3 activity in vitro., Conclusions: This study provides evidence that COX-2 is upregulated in the majority of cases of squamous dysplasia and SCC of esophagus, and that NS-398 can inhibit growth and induce apoptosis via activating caspase-3 activity in vitro. These results suggest that selective inhibitors of COX-2 may be an effective preventive and therapeutic option for esophageal carcinoma.

Background: Telomerase maintains telomere length and is considered to be necessary for the indefinite proliferation of human cells. Activation of telomerase plays a key role in the malignant transformation process. The aim of this study was to study the regulation of telomerase, and to explore the possibility of telomerase as a biomarker in squamous carcinogenesis of the esophagus., Methods: Twenty-nine esophageal squamous cell carcinomas (ESCC) and its corresponding adjacent normal tissues, and 47 epithelial squamous dysplasia tissues were analyzed by the reverse transcriptase-polymerase chain reaction (RT-PCR) technique for the mRNA expression of three major telomerase subunits: human telomerase RNA (hTR), telomerase protein component 1 (TP1), and human telomerase reverse transcriptase (hTERT) and by telomeric repeat amplification protocol assay (TRAP) for telomerase activity., Results: For the expression of hTR and TP1 mRNA, there were no significant differences among ESCC, dysplasia and normal tissues (P > 0.05). In contrast, hTERT mRNA expression was detected in 28 of 29 ESCC (96.6%), in 23 of 47 dysplasia (48.9%), and only in two of 29 normal tissues (7.5%). Telomerase activity was positive in 25 of 29 ESCC (86.2%), in 21 of 47 (44.7%) epithelial dysplasia tissues, and in none of normal tissue. All together, 95 of 105 cases (90.48%) were concordant for both results, i.e., telomerase activity positive and hTERT positive or telomerase activity negative and hTERT negative tissues, and telomerase activity correlated with hTERT mRNA expression (P < 0.001)., Conclusions: Higher telomerase activity and hTERT mRNA expression were shown during an early stage in the esophageal carcinogenesis. Activation of telomerase activity was strongly correlated with hTERT mRNA expression, suggesting hTERT is a major regulator of telomerase activity, and telomerase activation may play a critical role in esophageal carcinogenesis. Therefore, telomerase, especially hTERT can be used as a potential molecular biomarker of ESCC., (Copyright 2004 Wiley-Liss, Inc.)

To investigate the effect of X-ray repair cross complementing 1 (XRCC1) genetic polymorphisms on esophageal cancer risk, we determined XRCC1 polymorphisms at codon 194 (Arg --> Trp) and codon 399 (Arg --> Gln) in 135 patients with esophageal squamous cell carcinoma (ESCC) and 152 normal controls from hospitals. Although polymorphism at codon 194 was not associated with risk for ESCC, we found that the frequency of XRCC1 399 Gln/Gln genotype in ESCC patients (14.1%) was significantly higher than that in normal controls (3.3%), and that XRCC1 399 Gln/Gln genotype was associated with an increased risk of ESCC (odds ratio (OR) = 5.15, 95% confidence interval (CI): 2.42-0.93). In addition, we found that the risk for smoker increased 4.2-fold than non-smokers in the 399 Gln/Gln genotype (OR = 4.20, 95% CI: 2.37-7.44). These results suggest that XRCC1 399 Gln/Gln genotype may contribute to the risk of ESCC and modify risk associated with smoking.

Objectives: To investigate the expression of cyclooxygenase-2 (cox-2) protein in normal squamous epithelium, squamous dysplasia and squamous cell carcinoma of the esophagus, and to elucidate the role of cox-2 in esophageal carcinogenesis., Methods: Biopsy specimens of atypical esophageal dysplasia (n = 47) and surgical resection of squamous cell carcinoma (n = 86) were compared with normal esophageal specimens (n = 42) and the expression of cox-2 in those specimens was analyzed by immunohistochemistry and western blotting., Results: A significant elevated cox-2 expression was shown in atypical esophageal squamous dysplasia and squamous cell carcinoma, as compared to that in normal esophageal squamous epithelium, with immunohistochemical stain scores of 2.67 +/- 1.77, 2.19 +/- 1.79 and 0.71 +/- 0.46, respectively. Results of western blotting analysis confirmed those obtained by immunohistochemistry. Cox-2 expression significantly correlated with proliferation activity assessed by proliferating cell nuclear antigen index in dysplastic and carcinomous lesions, respectively, and no such correlation could be found in normal esophageal mucosa. Elevated cox-2 expression was not associated with clinical-pathological features of esophageal squamous carcinoma, including age, gender, tumor size, histological grade, lymph node metastasis and clinical stage., Conclusion: Elevated expression of cox-2 in atypical squamous dysplasia and squamous cell carcinoma of the esophagus, which correlated with cell proliferation activity, indicated that cox-2 may be involved in the early stage of squamous carcinogenesis of the esophagus, and may be a target of prevention and treatment for esophageal squamous cell carcinoma.

Objective: To study the relationship of human telomerase reverse transcriptase (hTRT) and malignant transformation of esophageal dysplasia., Methods: Telomerase activity and hTRT expression in esophageal dysplasia (n = 47), squamous cell carcinoma (n = 29) and normal esophagus (n = 11) were detected by telomeric repeat amplification protocol (TRAP) and in situ hybridization, respectively., Results: Telomerase activity was detected in none of the 11 cases of normal esophageal tissues (0%) but in 21 of 47 cases (44.7%) of dysplasia, and in 25 of 29 cases (86.2%) of esophageal squamous cell carcinoma. There were statistically significant differences among the telomerase activity in normal esophagus, esophageal dysplasia, and in squamous cell carcinoma (chi(2) = 5.89, P < 0.05; chi(2) = 11.35, P < 0.01). hTRT mRNA was expressed in none of the 11 cases of normal esophageal tissues (0%) but in 23 of 47 cases (48.9%) of dysplasia, and in 24 of 29 cases (82.8%) of esophageal squamous cell carcinoma. There were statistically significant differences among the expression of hTRT mRNA in normal esophagus, esophageal dysplasia, and in squamous cell carcinoma (chi(2) = 6.99, P < 0.01; chi(2) = 7.32, P < 0.01). Significant correlation was found between the telomerase activity and the expression of hTRT mRNA (chi(2) = 57.91, P < 0.001)., Conclusion: The mRNA expression of hTRT which paralleled to telomerase activity implied that there was a crucial role to play in regulating the activation of telomerase, and was closely related to the malignant transformation of esophageal dysplasia. hTRT might serve as a new, valuable biomarker to detect esophageal squamous cell carcinoma.

Cyclooxygenase-2 (cox-2) overexpression has been observed in several types of human cancers and has been implicated in carcinogenesis. To elucidate the role of cox-2 in esophageal carcinogenesis, we evaluated the expression of cox-2 in normal squamous epithelium squamous epithelial dysplasia (n=47), and squamous cell carcinoma of the esophagus (n=86) by immunohistochemistry, reverse transcription-PCR assay, and western blotting. A significant overexpression of cox-2 was observed in esophageal squamous dysplasia and squamous cell carcinoma compared with normal squamous epithelium. The immunoreactive score of cox-2 expression, an index determined by intensity and positivity of cox-2 staining, was 0.71 +/- 0.46 (mean +/- SD) in normal squamous esophagus, 2.19 +/- 1.79 in squamous epithelial dysplasia, and 2.67 +/- 1.77 in squamous cell carcinoma. The results of immunohistochemistry were confirmed by a reverse transcription-PCR assay and western blotting analysis. Cox-2 expression level was correlated with proliferation activity assessed by proliferating cell nuclear antigen (PCNA) index and MIB-1 index in dysplastic lesion (r=0.55, P<0.01 with PCNA and r=0.72, P<0.01 with MIB-1) and carcinoma (r=0.56, P<0.01 with PCNA and r=0.72, P<0.01 with MIB-1). Elevated cox-2 expression was associated with high p53 expression (p<0.001) but not with clinicopathological features including age, sex, tumor size, histological grade, lymph node metastasis, and TNM stage. The results indicated that cox-2 may be involved in an early stage of squamous carcinogenesis of the esophagus, and that cox-2 overexpression was related to cell proliferation in esophageal squamous dysplasia and squamous cell carcinoma.