YY1 modulates the radiosensitivity of esophageal squamous cell carcinoma through KIF3B-mediated Hippo signaling pathway.

Radiotherapy is an important strategy in the comprehensive treatment of esophageal squamous cell carcinoma (ESCC). However, effectiveness of radiotherapy is still restricted by radioresistance. Herein, we aimed to understand the mechanisms underlying ESCC radioresistance, for which we looked into the potential role of YY1. YY1 was upregulated in radioresistant tissues and correlated with poor prognosis of patients with ESCC. YY1 depletion enhanced the radiosensitivity of ESCC in vitro and in vivo. Multi-group sequencing showed that downregulation of YY1 inhibited the transcriptional activity of Kinesin Family Member 3B (KIF3B), which further activated the Hippo signaling pathway by interacting with Integrin-beta1 (ITGB1). Once the Hippo pathway was activated, its main effector, Yes-associated protein 1 (YAP1), was phosphorylated in the cytoplasm and its expression reduced in the nucleus, thus enhancing the radiosensitivity by regulating its targeted genes. Our study provides new insights into the mechanisms underlying ESCC radioresistance and highlights the potential role of YY1 as a therapeutic target for ESCC.

SLC43A2 and NFκB signaling pathway regulate methionine/cystine restriction-induced ferroptosis in esophageal squamous cell carcinoma via a feedback loop.

Studies have indicated dietary restriction of methionine/cystine provided a therapeutic benefit in diseases such as cancer. However, the molecular and cellular mechanisms that underlie the interaction between methionine/cystine restriction (MCR) and effects on esophageal squamous cell carcinoma (ESCC) have remained elusive. Here, we discovered the dietary restriction of methionine/cystine has a large effect on cellular methionine metabolism as assayed in a ECA109 derived xenograft model. RNA-seq and enrichment analysis suggested the blocked tumor progression was affected by ferroptosis, together with the NFκB signaling pathway activation in ESCC. Consistently, GSH content and GPX4 expression were downregulated by MCR both in vivo and in vitro. The contents of Fe2+ and MDA were negatively correlated with supplementary methionine in a dose-dependent way. Mechanistically, MCR and silent of SLC43A2, a methionine transporter, diminished phosphorylation of IKKα/ß and p65. Blocked NFκB signaling pathway further decreased the expression of SLC43A2 and GPX4 in both mRNA and protein level, which in turn downregulated the methionine intake and stimulated ferroptosis, respectively. ESCC progression was inhibited by enhanced ferroptosis and apoptosis and impaired cell proliferation. In this study, we proposed a novel feedback regulation mechanism underlie the correlation between dietary restriction of methionine/cystine and ESCC progression. MCR blocked cancer progression via stimulating ferroptosis through the positive feedback loop between SLC43A2 and NFκB signaling pathways. Our results provided the theoretical basis and new targets for ferroptosis-based clinical antitumor treatments for ESCC patients.

Diclofenac exhibits cytotoxic activity associated with metabolic alterations and p53 induction in ESCC cell lines and decreases ESCC tumor burden in vivo.

Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive forms of human malignancy, often displaying limited therapeutic response. Here, we examine the non-steroidal anti-inflammatory drug diclofenac (DCF) as a novel therapeutic agent in ESCC using complementary in vitro and in vivo models. DCF selectively reduced viability of human ESCC cell lines TE11, KYSE150, and KYSE410 as compared with normal primary or immortalized esophageal keratinocytes. Apoptosis and altered cell cycle profiles were documented in DCF-treated TE11 and KYSE 150. In DCF-treated TE11, RNA-Sequencing identified differentially expressed genes and Ingenuity Pathway Analysis predicted alterations in pathways associated with cellular metabolism and p53 signaling. Downregulation of proteins associated with glycolysis was documented in DCF-treated TE11 and KYSE150. In response to DCF, TE11 cells further displayed reduced levels of ATP, pyruvate, and lactate. Evidence of mitochondrial depolarization and superoxide production was induced by DCF in TE11 and KYSE150. In DCF-treated TE11, the superoxide scavenger MitoTempo improved viability, supporting a role for mitochondrial reactive oxygen species in DCF-mediated toxicity. DCF treatment resulted in increased expression of p53 in TE11 and KYSE150. p53 was further identified as a mediator of DCF-mediated toxicity in TE11 as genetic depletion of p53 partially limited apoptosis in response to DCF. Consistent with the anticancer activity of DCF in vitro, the drug significantly decreased tumor burdene in syngeneic ESCC xenograft tumors and 4-nitroquinoline 1-oxide-mediated ESCC lesions in vivo. These preclinical findings identify DCF as an experimental therapeutic that should be explored further in ESCC.

Exploring the hub genes and mechanisms of Daphne altaica treating esophageal squamous cell carcinoma based on network pharmacology and bioinformatics analysis.

PURPOSE: Esophageal squamous cell carcinoma (ESCC), is a frequent digestive tract malignant carcinoma with a high fatality rate. Daphne altaica (D. altaica), a medicinal plant that is frequently employed in Kazakh traditional medicine, and which has traditionally been used to cure cancer and respiratory conditions, but research on the mechanism is lacking. Therefore, we examined and verified the hub genes and mechanism of D. altaica treating ESCC. METHODS: Active compounds and targets of D. altaica were screened by databases such as TCMSP, and ESCC targets were screened by databases such as GeneCards and constructed the compound-target network and PPI network. Meantime, data sets between tissues and adjacent non-cancerous tissues from GEO database (GSE100942, GPL570) were analyzed to obtain DEGs using the limma package in R. Hub genes were validated using data from the Kaplan-Meier plotter database, TIMER2.0 and GEPIA2 databases. Finally, AutoDock software was used to predict the binding sites through molecular docking. RESULTS: In total, 830 compound targets were obtained from TCMSP and other databases. In addition, 17,710 disease targets were acquired based on GeneCards and other databases. In addition, we constructed the compound-target network and PPI network. Then, 127 DEGs were observed (82 up-regulated and 45 down-regulated genes). Hub genes were screened including TOP2A, NUF2, CDKN2A, BCHE, and NEK2, and had been validated with the help of several publicly available databases. Finally, molecular docking results showed more stable binding between five hub genes and active compounds. CONCLUSIONS: In the present study, five hub genes were screened and validated, and potential mechanisms of action were predicted, which could provide a theoretical understanding of the treatment of ESCC with D. altaica.

Integrative analysis of bulk and single-cell gene expression profiles to identify tumor-associated macrophage-derived CCL18 as a therapeutic target of esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is a common gastrointestinal malignancy with poor patient prognosis. Current treatment for ESCC, including immunotherapy, is only beneficial for a small subset of patients. Better characterization of the tumor microenvironment (TME) and the development of novel therapeutic targets are urgently needed. METHODS: In the present study, we hypothesized that integration of single-cell transcriptomic sequencing and large microarray sequencing of ESCC biopsies would reveal the key cell subtypes and therapeutic targets that determine the prognostic and tumorigenesis of ESCC. We characterized the gene expression profiles, gene sets enrichment, and the TME landscape of a microarray cohort including 84 ESCC tumors and their paired peritumor samples. We integrated single-cell transcriptomic sequencing and bulk microarray sequencing of ESCC to reveal key cell subtypes and druggable targets that determine the prognostic and tumorigenesis of ESCC. We then designed and screened a blocking peptide targeting Chemokine C-C motif ligand 18 (CCL18) derived from tumor associated macrophages and validated its potency by MTT assay. The antitumor activity of CCL18 blocking peptide was validated in vivo by using 4-nitroquinoline-1-oxide (4-NQO) induced spontaneous ESCC mouse model. RESULTS: Comparative gene expression and cell-cell interaction analyses revealed dysregulated chemokine and cytokine pathways during ESCC carcinogenesis. TME deconvolution and cell interaction analyses allow us to identify the chemokine CCL18 secreted by tumor associated macrophages could promote tumor cell proliferation via JAK2/STAT3 signaling pathway and lead to poor prognosis of ESCC. The peptide Pep3 could inhibit the proliferation of EC-109 cells promoted by CCL18 and significantly restrain the tumor progression in 4-NQO-induced spontaneous ESCC mouse model. CONCLUSIONS: For the first time, we discovered and validated that CCL18 blockade could significantly prevent ESCC progression. Our study revealed the comprehensive cell-cell interaction network in the TME of ESCC and provided novel therapeutic targets and strategies to ESCC treatment.

Implication of changes in PD-L1 expression during neoadjuvant chemotherapy with docetaxel, cisplatin, and 5-fluorouracil (DCF) regimen in esophageal squamous cell carcinoma.

BACKGROUND: Neoadjuvant docetaxel plus cisplatin and 5-FU (NAC-DCF) and adjuvant nivolumab monotherapy are the standard care for locally advanced resectable esophageal squamous cell carcinoma (ESCC). However, no effective biomarkers have been found in perioperative setting. We investigated how programmed death-ligand 1 (PD-L1) changes before and after NAC-DCF and how it relates to the therapeutic effect of NAC-DCF in resectable ESCC. METHODS: PD-L1 expression in paired diagnostic biopsy and surgically resected tissues from ESCC patients who underwent surgical resection after receiving two or three NAC-DCF cycles was evaluated. PD-L1 positivity was defined as a combined positive score (CPS) of 10% ≤ . Gene expression analysis was conducted using samples before NAC-DCF. RESULTS: Sixty-six paired samples from 33 patients were included in PD-L1 expression analysis, and 33 Pre-NAC samples acquired by diagnostic biopsy were included in gene expression analysis. Pretreatment, 3 (9%), 13 (39%), and 17 (52%) patients harbored tumors with CPS ranges of < 1%, 1%-10%, and 10% ≤ , respectively. After NAC-DCF, 5 (15%), 15 (45%), and 13 (39%) tumors presented CPS ranges of < 1%, 1%-10%, and 10% ≤ , respectively. The concordance rate between Pre-and Post-NAC-DCF samples was 45%. Patients with PD-L1-negative tumors both before and after NAC-DCF (n = 9) had shorter survival and different gene expression profile characterized by upregulation in WNT signaling or neutrophils. CONCLUSIONS: A substantial PD-L1 expression alteration was observed, resulting in low concordance rate before and after NAC-DCF. Tumors persistently lacking PD-L1 had distinct gene expression profile with worse clinical outcomes, raising the need for further investigation.

N-linked glycoproteomic profiling in esophageal squamous cell carcinoma.

BACKGROUND: Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing. AIM: To characterize the N-linked glycoproteomic profile in esophageal squamous cell carcinoma (ESCC) via two complementary approaches. METHODS: Using tandem multilectin affinity chromatography for enrichment of N-linked glycoproteins, we performed N-linked glycoproteomic profiling in ESCC tissues by two-dimensional gel electrophoresis (2-DE)-based and isobaric tags for relative and absolute quantification (iTRAQ) labeling-based mass spectrometry quantitation in parallel, followed by validation of candidate glycoprotein biomarkers by Western blot. RESULTS: 2-DE-based and iTRAQ labeling-based quantitation identified 24 and 402 differentially expressed N-linked glycoproteins, respectively, with 15 in common, demonstrating the outperformance of iTRAQ labeling-based quantitation over 2-DE and complementarity of these two approaches. Proteomaps showed the distinct compositions of functional categories between proteins and glycoproteins with differential expression associated with ESCC. Western blot analysis validated the up-regulation of total procathepsin D and high-mannose procathepsin D, and the down-regulation of total haptoglobin, high-mannose clusterin, and GlcNAc/sialic acid-containing fraction of 14-3-3ζ in ESCC tissues. The serum levels of glycosylated fractions of clusterin, proline-arginine-rich end leucine-rich repeat protein, and haptoglobin in patients with ESCC were remarkably higher than those in healthy controls. CONCLUSION: Our study provides insights into the aberrant N-linked glycoproteome associated with ESCC, which will be a valuable resource for future investigations.

microRNA-497-mediated Smurf2/YY1/HIF2α axis in tumor growth and metastasis of esophageal squamous cell carcinoma.

Aberrant expression of microRNA-497 (miR-497) is associated with tumor progression, but the molecular mechanisms in tumorigenesis remain largely unknown. Here, we report that miR-497 expression is downregulated in esophageal squamous cell carcinoma (ESCC) clinical samples. Consistently, upregulation of miR-497 inhibits ESCC cell malignant properties and tumor growth in vivo. Importantly, we uncovered that miR-497 upregulation suppressed ESCC cell growth and tumor growth by inhibiting Smurf2. Mechanistically, we showed that Smurf2 was a target of miR-497, and mediated YY1 expression to elevate HIF2α expression, thereby enhancing the malignancy of ESCC cells. Together, our study uncovered the role of the miR-497-mediated Smurf2/YY1/HIF2α axis in tumor growth and metastasis, which might provide potential therapeutic targets for human ESCC.

Identification of tissue-specific microbial profile of esophageal squamous cell carcinoma by full-length 16S rDNA sequencing.

It was previously believed that the microbial community in the esophagus was relatively stable, but it has been reported that different esophageal diseases have different microbial community characteristics. In this study, we recruited patients with esophageal squamous cell carcinoma (ESCC) and collected 51 pairs of tumor and adjacent non-tumor tissues for full-length 16S rDNAsequencing and qPCR to compare the differences in microbial community structure. The results of sequencing in 19 pairs of tissues showed that Proteobacteria, Firmicutes, Bacteroidetes, Deinococcus-Thermus, and Actinobacteria were the main bacteria in tumor and adjacent non-tumor tissues. At the genus level, the bacteria with the highest relative proportion in tumor and adjacent non-tumor tissues were Streptococcus and Labrys, respectively. At the same time, it was observed that the complexity of microbial interactions in tumor tissues was weaker than that of adjacent non-tumor tissues. The results also found that the relative abundance of 24 taxa was statistically different between tumor and adjacent non-tumor tissues. The findings of qPCR in 32 pairs of tissues further evidence that the relative proportions of Blautia, Treponema, Lactobacillus murinus, Peptoanaerobacter stomatis, and Fusobacteria periodonticum were statistically different in tumor and adjacent non-tumor tissues. The findings of PIRCUSt2 indicated the lipopolysaccharide biosynthesis and biotin metabolism in the microbiome of cancer tissues are more significant. This study supplements the existing information on the structure, function, and interaction of microorganisms in the esophagus in situ and provides a direction for the further exploration of the relationship between esophageal in situ microorganisms and esophageal squamous cell carcinoma. KEY POINTS: • The structure of the microbial community in esophageal cancer tissue and adjacent non-tumor tissues at the phylum level is similar • Streptococcus and Labrys are the most important bacteria in esophageal tumor tissues and adjacent non-tumor tissues, respectively • Microbial interactions in tumor tissues are stronger than in adjacent non-tumor tissues.

Genomic and epigenomic evolution of acquired resistance to combination therapy in esophageal squamous cell carcinoma.

BACKGROUNDTargeted arterial infusion of verapamil combined with chemotherapy (TVCC) is an effective clinical interventional therapy for esophageal squamous cell carcinoma (ESCC), but multidrug resistance (MDR) remains the major cause of relapse or poor prognosis, and the underlying molecular mechanisms of MDR, temporal intratumoral heterogeneity, and clonal evolutionary processes of resistance have not been determined.METHODSTo elucidate the roles of genetic and epigenetic alterations in the evolution of acquired resistance during therapies, we performed whole-exome sequencing on 16 serial specimens from 7 patients with ESCC at every cycle of therapeutic intervention from 3 groups, complete response, partial response, and progressive disease, and we performed whole-genome bisulfite sequencing for 3 of these 7 patients, 1 patient from each group.RESULTSPatients with progressive disease exhibited a substantially higher genomic and epigenomic temporal heterogeneity. Subclonal expansions driven by the beneficial new mutations were observed during combined therapies, which explained the emergence of MDR. Notably, SLC7A8 was identified as a potentially novel MDR gene, and functional assays demonstrated that mutant SLC7A8 promoted the resistance phenotypes of ESCC cell lines. Promoter methylation dynamics during treatments revealed 8 drug resistance protein-coding genes characterized by hypomethylation in promoter regions. Intriguingly, promoter hypomethylation of SLC8A3 and mutant SLC7A8 were enriched in an identical pathway, protein digestion and absorption, indicating a potentially novel MDR mechanism during treatments.CONCLUSIONOur integrated multiomics investigations revealed the dynamics of temporal genetic and epigenetic inter- and intratumoral heterogeneity, clonal evolutionary processes, and epigenomic changes, providing potential MDR therapeutic targets in treatment-resistant patients with ESCC during combined therapies.FUNDINGNational Natural Science Foundation of China, Science Foundation of Peking University Cancer Hospital, CAMS Innovation Fund for Medical Sciences, Major Program of Shenzhen Bay Laboratory, Guangdong Basic and Applied Basic Research Foundation, and the third round of public welfare development and reform pilot projects of Beijing Municipal Medical Research Institutes.

Independent and opposing associations of dietary phytosterols intake and PLCE1 rs2274223 polymorphisms on esophageal squamous cell carcinoma risk.

PURPOSE: This study was to evaluate the associations of dietary intake of total and specific phytosterols and risk of esophageal squamous cell carcinoma (ESCC) and to explore their joint effects with PLCE1 rs2274223 polymorphisms. METHODS: A population-based case-control study was conducted in a Chinese rural population and 856 eligible incident ESCC cases and 856 controls were included. A validated food frequency questionnaire was used to collect dietary consumption and PLCE1 rs2274223 polymorphisms were genotyped. Unadjusted and adjusted odds ratios (ORs) with 95% confidence interval (CI) were assessed via logistic regression model. RESULTS: When comparing the highest with lowest intake quartiles, ß-sitosterol, campesterol, stigmasterol, ß-sitostanol, campestanol, and total phytosterols were all associated with a decreased risk of ESCC, with adjusted ORs being 0.32 (95% CI 0.20-0.48), 0.18 (95% CI 0.11-0.27), 0.45 (95% CI 0.29-0.70), 0.13 (95% CI 0.08-0.20), 0.14 (95% CI 0.09-0.22) and 0.28 (95% CI 0.18-0.43), respectively. An exposure-response relationship was also observed for both total and five specific phytosterols (all P for trend < 0.001). In comparison to rs2274223 AA genotype, both GA genotype (OR: 1.47, 95% CI 1.16-1.85) and GG genotype (OR: 2.13, 95% CI 1.20-3.84) were associated with an increased risk of ESCC. However, no interaction was observed between total/specific phytosterols intake and rs2274223 polymorphisms. CONCLUSION: Higher dietary intake of total and five specific phytosterols was associated with a lower risk of ESCC, and the risk of ESCC increased with the increment of rs2274223 G allele. The negative association between phytosterols and ESCC risk was not modified by rs2274223 polymorphisms. Foods or supplements rich in phytosterols are a promising source for chemoprevention of ESCC, and still, clinical trials will be required in any specific case.

Deoxypodophyllotoxin, a Lignan from Anthriscus sylvestris, Induces Apoptosis and Cell Cycle Arrest by Inhibiting the EGFR Signaling Pathways in Esophageal Squamous Cell Carcinoma Cells.

Deoxypodophyllotoxin (DPT) derived from Anthriscus sylvestris (L.) Hoffm has attracted considerable interest in recent years because of its anti-inflammatory, antitumor, and antiviral activity. However, the mechanisms underlying DPT mediated antitumor activity have yet to be fully elucidated in esophageal squamous cell carcinoma (ESCC). We show here that DPT inhibited the kinase activity of epidermal growth factor receptor (EGFR) directly, as well as phosphorylation of its downstream signaling kinases, AKT, GSK-3ß, and ERK. We confirmed a direct interaction between DPT and EGFR by pull-down assay using DPT-beads. DPT treatment suppressed ESCC cell viability and colony formation in a time- and dose-dependent manner, as shown by MTT analysis and soft agar assay. DPT also down-regulated cyclin B1 and cdc2 expression to induce G2/M phase arrest of the cell cycle and upregulated p21 and p27 expression. DPT treatment of ESCC cells triggered the release of cytochrome c via loss of mitochondrial membrane potential, thereby inducing apoptosis by upregulation of related proteins. In addition, treatment of KYSE 30 and KYSE 450 cells with DPT increased endoplasmic reticulum stress, reactive oxygen species generation, and multi-caspase activation. Consequently, our results suggest that DPT has the potential to become a new anticancer therapeutic by inhibiting EGFR mediated AKT/ERK signaling pathway in ESCC.

YY1-induced upregulation of FOXP4-AS1 and FOXP4 promote the proliferation of esophageal squamous cell carcinoma cells.

Esophageal squamous cell carcinoma (ESCC) belongs to one of the most common malignant tumors worldwide and possesses high mortality. Long non-coding RNAs (lncRNAs) have been demonstrated to be essential biological participants in the progression of ESCC. On the basis of bio-informatics prediction, forkhead box P4 antisense RNA 1 (FOXP4-AS1) and forkhead box P4 (FOXP4) were upregulated in esophageal carcinoma samples and were positively correlated with each other. The present study aimed to explore the function of FOXP4-AS1 and FOXP4 in ESCC cells. Function assays disclosed that knockdown of FOXP4-AS1 or FOXP4 efficiently suppressed cell proliferation and induced cell apoptosis. Moreover, FOXP4-AS1 positively regulated FOXP4 by interacting with insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) to stabilize FOXP4 messenger RNA. In addition, FOXP4-AS1 could upregulate the expression of FOXP4 by sponging miR-3184-5p. Finally, we found that Yin Yang 1 (YY1) is a transcription factor that can transcriptionally activate both FOXP4-AS1 and FOXP4 in ESCC cells. In a word, YY1-induced upregulation of FOXP4-AS1 and FOXP4 promote the proliferation of ESCC cells.

A Novel Micropeptide Encoded by Y-Linked LINC00278 Links Cigarette Smoking and AR Signaling in Male Esophageal Squamous Cell Carcinoma.

Long noncoding RNAs (lncRNA) have been shown to play critical roles in many diseases, including esophageal squamous cell carcinoma (ESCC). Recent studies have reported that some lncRNA encode functional micropeptides. However, the association between ESCC and micropeptides encoded by lncRNA remains largely unknown. In this study, we characterized a Y-linked lncRNA, LINC00278, which was downregulated in male ESCC. LINC00278 encoded a Yin Yang 1 (YY1)-binding micropeptide, designated YY1BM. YY1BM was involved in the ESCC progression and inhibited the interaction between YY1 and androgen receptor (AR), which in turn decreased expression of eEF2K through the AR signaling pathway. Downregulation of YY1BM significantly upregulated eEF2K expression and inhibited apoptosis, thus conferring ESCC cells more adaptive to nutrient deprivation. Cigarette smoking decreased m6A modification of LINC00278 and YY1BM translation. In conclusion, these results provide a novel mechanistic link between cigarette smoking and AR signaling in male ESCC progression. SIGNIFICANCE: Posttranscriptional modification of a micropeptide-encoding lncRNA is negatively impacted by cigarette smoking, disrupting negative regulation of the AR signaling pathway in male ESCC. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/13/2790/F1.large.jpg.See related commentary by Banday et al., p. 2718.

Abrogation of esophageal carcinoma development in miR-31 knockout rats.

MicroRNA-31 (miR-31) is overexpressed in esophageal squamous cell carcinoma (ESCC), a deadly disease associated with dietary Zn deficiency and inflammation. In a Zn deficiency-promoted rat ESCC model with miR-31 up-regulation, cancer-associated inflammation, and a high ESCC burden following N-nitrosomethylbenzylamine (NMBA) exposure, systemic antimiR-31 delivery reduced ESCC incidence from 85 to 45% (P = 0.038) and miR-31 gene knockout abrogated development of ESCC (P = 1 × 10-6). Transcriptomics, genome sequencing, and metabolomics analyses in these Zn-deficient rats revealed the molecular basis of ESCC abrogation by miR-31 knockout. Our identification of EGLN3, a known negative regulator of nuclear factor κB (NF-κB), as a direct target of miR-31 establishes a functional link between oncomiR-31, tumor suppressor target EGLN3, and up-regulated NF-κB-controlled inflammation signaling. Interaction among oncogenic miR-31, EGLN3 down-regulation, and inflammation was also documented in human ESCCs. miR-31 deletion resulted in suppression of miR-31-associated EGLN3/NF-κB-controlled inflammatory pathways. ESCC-free, Zn-deficient miR-31-/- rat esophagus displayed no genome instability and limited metabolic activity changes vs. the pronounced mutational burden and ESCC-associated metabolic changes of Zn-deficient wild-type rats. These results provide conclusive evidence that miR-31 expression is necessary for ESCC development.

Combination treatment with highly bioavailable curcumin and NQO1 inhibitor exhibits potent antitumor effects on esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is one of the most intractable cancers, so the development of novel therapeutics has been required to improve patient outcomes. Curcumin, a polyphenol from Curcuma longa, exhibits various health benefits including antitumor effects, but its clinical utility is limited because of low bioavailability. Theracurmin® (THC) is a highly bioavailable curcumin dispersed with colloidal submicron particles. METHODS: We examined antitumor effects of THC on ESCC cells by cell viability assay, colony and spheroid formation assay, and xenograft models. To reveal its mechanisms, we investigated the levels of reactive oxygen species (ROS) and performed microarray gene expression analysis. According to those analyses, we focused on NQO1, which involved in the removal of ROS, and examined the effects of NQO1-knockdown or overexpression on THC treatment. Moreover, the therapeutic effect of THC and NQO1 inhibitor on ESCC patient-derived xenografts (PDX) was investigated. RESULTS: THC caused cytotoxicity in ESCC cells, and suppressed the growth of xenografted tumors more efficiently than curcumin. THC increased ROS levels and activated the NRF2-NMRAL2P-NQO1 expressions. Inhibition of NQO1 in ESCC cells by shRNA or NQO1 inhibitor resulted in an increased sensitivity of cells to THC, whereas overexpression of NQO1 antagonized it. Notably, NQO1 inhibitor significantly enhanced the antitumor effects of THC in ESCC PDX tumors. CONCLUSIONS: These findings suggest the potential usefulness of THC and its combination with NQO1 inhibitor as a therapeutic option for ESCC.

Targeting AKT with Oridonin Inhibits Growth of Esophageal Squamous Cell Carcinoma In Vitro and Patient-Derived Xenografts In Vivo.

Overexpression or activation of AKT is very well known to control cell growth, survival, and gene expression in solid tumors. Oridonin, an inflammatory medical and diterpenoid compound isolated from Rabdosia rubescens, has exhibited various pharmacologic and physiologic properties, including antitumor, antibacterial, and anti-inflammatory effects. In this study, we demonstrated that oridonin is an inhibitor of AKT and suppresses proliferation of esophageal squamous cell carcinoma (ESCC) in vitro and in vivo The role of AKT in ESCC was studied using immuno-histochemical analysis of a tumor microarray, the effect of AKT knockdown on cell growth, and treatment of cells with MK-2206, an AKT inhibitor. Oridonin blocked AKT kinase activity and interacted with the ATP-binding pocket of AKT. It inhibited growth of KYSE70, KYSE410, and KYSE450 esophageal cancer cells in a time- and concentration-dependent manner. Oridonin induced arrest of cells in the G2-M cell-cycle phase, stimulated apoptosis, and increased expression of apoptotic biomarkers, including cleaved PARP, caspase-3, caspase-7, and Bims in ESCC cell lines. Mechanistically, we found that oridonin diminished the phosphorylation and activation of AKT signaling. Furthermore, a combination of oridonin and 5-fluorouracil or cisplatin (clinical chemotherapeutic agents) enhanced the inhibition of ESCC cell growth. The effects of oridonin were verified in patient-derived xenograft tumors expressing high levels of AKT. In summary, our results indicate that oridonin acts as an AKT inhibitor to suppress the growth of ESCC by attenuating AKT signaling. Mol Cancer Ther; 17(7); 1540-53. ©2018 AACR.

CYP1A2*1F Gene Variant, Alkaline Salt Tea Intake and Risk of Esophageal Squamous Cell Carcinoma.

Unlike many other cancers, the relationship of CYP1A2*1F (rs762551) polymorphism with esophageal squamous cell carcinoma (ESCC) risk has not been assessed so far. To evaluate its association with ESCC, we conducted a case control study in Kashmir, India, a high risk region. We recruited 404 histopathologically confirmed ESCC cases and 404 controls, individually matched for sex, age and residence to the respective cases. Information was obtained on dietary, lifestyle and environmental factors in face to face interviews using a structured questionnaire from each subject. Genotypes were analyzed by polymerase chain reaction, restriction fragment length polymorphism and sequencing randomly selected samples. Conditional logistic regression models were used to calculate odds ratios (ORs) and 95% confidence intervals (95% CIs). We found that mutant genotype (AA) of CYP1A2*1F polymorphism was associated with ESCC risk (OR = 3.11; 95% CI: 1.72-5.36). A very strong ESCC risk was observed in subjects who drank >1250 ml of salt tea daily and harbored mutant genotype of CYP1A2*1F (OR = 14.51; 95% CI: 5.33-39.47). The study indicates that CYP1A2*1F polymorphism is associated with ESCC risk and the risk is modified in salt drinkers. However, more replicative and mechanistic studies are needed to substantiate the findings.