Determination of pepsin in human saliva using pepsin-susceptible peptide reporter and colorimetric dipstick assay: a prospective, cross-sectional study.

A simple and effective pepsin detection assay is reported based on a pepsin-susceptible peptide (PSP) reporter degradation strategy. PSP, which can be specifically cleaved by pepsin, was modified with fluorescein isothiocyanate (FITC) and biotin at the N- and C-terminals to be used as a reporter for colorimetric detection of dipsticks. A universal lateral flow dipstick consisting of a streptavidin test line for biotin binding and a sample pad immobilized with a gold-labeled polyclonal (rabbit) anti-FITC antibody was used to verify PSP-based pepsin detection. When the PSP reporter reacts with pepsin in a tube, it cleaves into two fragments, and the cleaved fragments do not display any color on the test line. Therefore, the higher the concentration of pepsin is, the greater is the decrease in test line intensity (IT-line) and the higher is the control line intensity (IC-line). First, the PSP cleavage and dipstick assay conditions for pepsin detection was optimized. The ratio of color intensity (IT-line/IC-line) of PSP-based dipstick assay showed a linear relationship with log concentration of pepsin ranging between 4 and 500 ng/mL (R2 = 0.98, n = 6), with a limit of detection of 1.4 ng/mL. It also exhibited high specificity and good reproducibility. Finally, pepsin levels were quantified in saliva samples from healthy controls (n = 34) and patients with laryngopharyngeal reflux (LPR, n = 61). Salivary pepsin levels were higher in patients with LPR than in healthy controls. The salivary pepsin levels correlated with those measured using a conventional enzyme-linked immunosorbent assay kit. Therefore, this PSP-based dipstick assay is a convenient tool for assessing salivary pepsin levels.

Clinically conserved genomic subtypes of gastric adenocarcinoma.

Gastric adenocarcinoma (GAC) is a lethal disease characterized by genomic and clinical heterogeneity. By integrating 8 previously established genomic signatures for GAC subtypes, we identified 6 clinically and molecularly distinct genomic consensus subtypes (CGSs). CGS1 have the poorest prognosis, very high stem cell characteristics, and high IGF1 expression, but low genomic alterations. CGS2 is enriched with canonical epithelial gene expression. CGS3 and CGS4 have high copy number alterations and low immune reactivity. However, CGS3 and CGS4 differ in that CGS3 has high HER2 activation, while CGS4 has high SALL4 and KRAS activation. CGS5 has the high mutation burden and moderately high immune reactivity that are characteristic of microsatellite instable tumors. Most CGS6 tumors are positive for Epstein Barr virus and show extremely high levels of methylation and high immune reactivity. In a systematic analysis of genomic and proteomic data, we estimated the potential response rate of each consensus subtype to standard and experimental treatments such as radiation therapy, targeted therapy, and immunotherapy. Interestingly, CGS3 was significantly associated with a benefit from chemoradiation therapy owing to its high basal level of ferroptosis. In addition, we also identified potential therapeutic targets for each consensus subtype. Thus, the consensus subtypes produced a robust classification and provide for additional characterizations for subtype-based customized interventions.

Association between cancer stem cell gene expression signatures and prognosis in head and neck squamous cell carcinoma.

BACKGROUND: Various cancer stem cell (CSC) biomarkers and the genes encoding them in head and neck squamous cell carcinoma (HNSCC) have been identified and evaluated. However, the validity of these factors in the prognosis of HNSCC has been questioned and remains unclear. In this study, we examined the clinical significance of CSC biomarker genes in HNSCC, using five publicly available HNSCC cohorts. METHODS: To predict the prognosis of patients with HNSCC, we developed and validated the expression signatures of CSC biomarker genes whose mRNA expression levels correlated with at least one of the four CSC genes (CD44, MET, ALDH1A1, and BMI1). RESULTS: Patients in The Cancer Genome Atlas (TCGA) HNSCC cohort were classified into CSC gene expression-associated high-risk (CSC-HR; n = 285) and CSC gene expression-associated low-risk (CSC-LR; n = 281) subgroups. The 5-year overall survival and recurrence-free survival rates were significantly lower in the CSC-HR subgroup than in the CSC-LR subgroup (p = 0.04 and 0.02, respectively). The clinical significance of the CSC gene expression signature was validated using four independent cohorts. Analysis using Cox proportional hazards models showed that the CSC gene expression signature was an independent prognostic factor of non-oropharyngeal HNSCC which mostly indicates HPV (-) status. Furthermore, the CSC gene expression signature was associated with the prognosis of HNSCC patients who received radiotherapy. CONCLUSION: The CSC gene expression signature is associated with the prognosis of HNSCC and may help in personalized treatments for patients with HNSCC, especially in cases with HPV (-) status who were classified in more detail.

Stabilization of HDAC1 via TCL1-pAKT-CHFR axis is a key element for NANOG-mediated multi-resistance and stem-like phenotype in immune-edited tumor cells.

Cancer immunoediting enriches NANOG expression in tumor cells, resulting in multi-drug resistance and stem-like phenotypes. We previously demonstrated that these NANOG-associated phenotypes are promoted through HDAC1 transcriptional upregulation. In this study, we identified that NANOG also contributes to the stabilization of HDAC1 protein through the AKT signaling pathway. NANOG-AKT axis leads to phosphor-dependent inactivation of CHFR, an E3 ligase for HDAC1 protein, and thereby inhibiting the ubiquitin-mediated degradation of HDAC1. Furthermore, AKT inhibition disrupts HDAC1 WT-mediated phenotypes but had no effect on the phenotypes mediated by HDAC1 FM, a mutant that is unable to interact with CHFR. Critically, we applied a catalytic dead mutant, HDAC1-H141A, to uncover that HDAC1 confers immune-resistance, drug-resistance and stem-like phenotype in tumor cells through its catalytic activity. Collectively, our results establish a firm molecular link in immune-edited tumor cells among NANOG, AKT, CHFR, and HDAC1, identifying HDAC1 as a molecular target in controlling NANOGHIGH immune-refractory cancer.

Recurrence-associated pathways in hepatitis B virus-positive hepatocellular carcinoma.

BACKGROUND: Despite the recent identification of several prognostic gene signatures, the lack of common genes among experimental cohorts has posed a considerable challenge in uncovering the molecular basis underlying hepatocellular carcinoma (HCC) recurrence for application in clinical purposes. To overcome the limitations of individual gene-based analysis, we applied a pathway-based approach for analysis of HCC recurrence. RESULTS: By implementing a permutation-based semi-supervised principal component analysis algorithm using the optimal principal component, we selected sixty-four pathways associated with hepatitis B virus (HBV)-positive HCC recurrence (p < 0.01), from our microarray dataset composed of 142 HBV-positive HCCs. In relation to the public HBV- and public hepatitis C virus (HCV)-positive HCC datasets, we detected 46 (71.9%) and 18 (28.1%) common recurrence-associated pathways, respectively. However, overlap of recurrence-associated genes between datasets was rare, further supporting the utility of the pathway-based approach for recurrence analysis between different HCC datasets. Non-supervised clustering of the 64 recurrence-associated pathways facilitated the classification of HCC patients into high- and low-risk subgroups, based on risk of recurrence (p < 0.0001). The pathways identified were additionally successfully applied to discriminate subgroups depending on recurrence risk within the public HCC datasets. Through multivariate analysis, these recurrence-associated pathways were identified as an independent prognostic factor (p < 0.0001) along with tumor number, tumor size and Edmondson's grade. Moreover, the pathway-based approach had a clinical advantage in terms of discriminating the high-risk subgroup (N = 12) among patients (N = 26) with small HCC (<3 cm). CONCLUSIONS: Using pathway-based analysis, we successfully identified the pathways involved in recurrence of HBV-positive HCC that may be effectively used as prognostic markers.

SIRT1 deacetylates and stabilizes hypoxia-inducible factor-1α (HIF-1α) via direct interactions during hypoxia.

Upon shift to a hypoxic environment, cellular HIF-1α protein is stabilized, with a rapid decline in oxygen-sensitive hydroxylation. Several additional post-translational modifications of HIF-1α are critical in controlling protein stability during hypoxia. In the present study, we showed that SIRT1 stabilizes HIF-1α via direct binding and deacetylation during hypoxia. SIRT1 depletion or inactivation led to reduced hypoxic HIF-1α accumulation, accompanied by an increase in HIF-1α acetylation. Impaired HIF-1α accumulation was recovered upon inhibition of 26S proteasome activity, indicating that SIRT1 is essential for HIF-1α stabilization during hypoxia. Consistently, HIF-1α accumulation was enhanced upon overexpression of wild-type SIRT1, but not its dominant-negative form. SIRT1-mediated accumulation of HIF-1α protein led to increased expression of HIF-1α target genes, including VEGF, GLUT1 and MMP2, and ultimate promotion of cancer cell invasion. These findings collectively imply that hypoxic HIF-1α stabilization requires SIRT1 activation. Furthermore, SIRT1 protection of HIF-1α from acetylation may be a prerequisite for stabilization and consequent enhancement of cell invasion.

Prognostic significance of SASP-related gene signature of radiation therapy in head and neck squamous cell carcinoma.

In this study,we developed and validated the clinical significance of senescence SASP related gene signature and explored its association with radiation therapy (RT) in patients with head and neck squamous cell carcinoma (HNSCC). First, we searched the three published review literature associated with senescence associated secretory phenotype (SASP) and selected all 81 genes to develop SASP related gene signature. Then, 81 SASP related genes were adapted to gene expression dataset from TCGA. HNSCC patients of TCGA were classified into clusters 1 and 2 via unsupervised clustering according to SASP related gene signature. Kaplan-Meier plot survival analysis showed that cluster 1 had a poorer prognosis than cluster 2 in 5-years overall survival and recurrence-free survival. Similarly, cluster 1 showed a worse prognosis than cluster 2 in three validation cohorts. (E-MTAB-8588, FHCRC and KHU). Cox proportional hazards regression observed that the senescence SASP related signature was an independent prognostic factor for HNSCC patients. We also established a nomogram using a relevant clinical parameter and a risk score. Time-dependent receiver operating characteristic (ROC) analysis was carried out to assess the accuracy of the prognostic risk model and nomogram. Senescence SASP related gene signature was associated with the response to RT. Therefore, subsequent, in vitro experiments further validated the association between senescence SASP related gene signature and RT in HNSCC. In conclusion, we developed a senescence SASP related gene signature, which could predict survival of HNSCC patients, and this gene signature provides new clinical evidence for the accurate diagnosis and targeted RT of HNSCC.

Cell Death Induced by the Combination of Ephedra sinica Extract and Radiation in HNSCC is Positively Related to BAX and p-MLKL Expression.

BACKGROUND: Numerous studies have proven the efficacy and safety of natural products, and are widely used as attractive cancer treatments. The investigation of effective natural products for improving cancer treatment is a promising strategy. Combination treatment with radiosensitizers and radiotherapy (RT) is considered necessary for therapeutic improvement in head and neck squamous cell carcinoma(HNSCC). OBJECTIVE: This study aims to investigate whether Ephedra sinica (ES) extract could induce selective cell death in cancer cells and serve as a radiosensitizer for HNSCC. METHODS: HNSCC cells were pretreated with ES extract before radiation, and the radiosensitizing activity was assessed using a colony formation assay. Radiation-induced cell death was evaluated using an annexinV-FITC assay. Western blotting was performed to confirm cell death-related gene expression, including apoptosis and necrosis markers. RESULTS: ES extract significantly inhibited HNSCC cell viability (FaDu and SNU1076), while having minimal effect on normal HaCaT cells. When HNSCC cells were irradiated with 2, 4, or 8 Gy and cultured with ES extract (25 µg/mL), they exhibited increased radiation sensitivity compared to non-treated cells. The combination of ES extract and radiation resulted in increased cell death compared to non-treated, ES-treated, or irradiated cells. The apoptosis marker BAX and necrosis marker p-MLKL expression levels were also elevated following the combination treatment. CONCLUSION: ES extract demonstrated significant cytotoxic potential in HNSCC cells without affecting normal cells. It enhanced the radiosensitivity of HNSCC cells by upregulating BAX and p-MLKL expression, leading to increased cell death. These results suggest ES extract exhibits a potential radiosensitizing capacity in HNSCC.

Overexpression of Romo1 promotes production of reactive oxygen species and invasiveness of hepatic tumor cells.

BACKGROUND & AIMS: Chronic oxidative stress from reactive oxygen species (ROS) produced by the mitochondria promotes hepatocarcinogenesis and tumor progression. However, the exact mechanism by which mitochondrial ROS contributes to tumor cell invasion is not known. We investigated the role of ROS modulator 1 (Romo1) in hepatocellular carcinoma (HCC) development and tumor cell invasiveness. METHODS: We performed real-time, semi-quantitative, reverse transcriptase polymerase chain reaction; invasion and luciferase assays; and immunofluorescence and immunohistochemical analyses. The formation of pulmonary metastatic nodules after tumor cell injection was tested in severe combined immunodeficient mice. We analyzed Romo1 expression in HCC cell lines and tissues (n = 95). RESULTS: Expression of Romo1 was increased in HCC cells, compared with normal human lung fibroblast cells. Exogenous expression of Romo1 in HCC cells increased their invasive activity, compared with control cells. Knockdown of Romo1 in Hep3B and Huh-7 HCC cells reduced their invasive activity in response to stimulation with 12-O-tetradecanoylphorbol-13-acetate. Levels of Romo1 were increased compared with normal liver tissues in 63 of 95 HCC samples from patients. In HCC samples from patients, there was an inverse correlation between Romo1 overexpression and patient survival times. Increased levels of Romo1 also correlated with vascular invasion by the tumors, reduced differentiation, and larger tumor size. CONCLUSIONS: Romo1 is a biomarker of HCC progression that might be used in diagnosis. Reagents that inhibit activity of Romo1 and suppress mitochondrial ROS production, rather than eliminate up-regulated intracellular ROS, might be developed as cancer therapies.

SIRT1 suppresses cellular accumulation of ß-TrCP E3 ligase via protein degradation.

ß-Transducin repeat-containing protein (ß-TrCP), an E3 ligase, promotes the degradation of substrate proteins in response to various stimuli. Even though several ß-TrCP substrates have been identified to date, limited information of its upstream regulators is available. Here, we showed that SIRT1 suppresses ß-TrCP protein synthesis via post-translational degradation. SIRT1 depletion led to a significant increase in the ß-TrCP accumulation without affecting the mRNA level. Consistently, ß-TrCP protein accumulation induced by resveratrol was further enhanced upon SIRT1 depletion. Rescue of SIRT1 reversed the effect of resveratrol, leading to reduced ß-TrCP protein levels. Proteasomal inhibition led to recovery of ß-TrCP in cells with SIRT1 overexpression. Notably, the recovered ß-TrCP colocalized mostly with SIRT1. Thus, SIRT1 acts as a negative regulator of ß-TrCP synthesis via promoting protein degradation.

Gene signature predicting recurrence in oral squamous cell carcinoma is characterized by increased oxidative phosphorylation.

Although numerous studies have used systemic approaches to identify prognostic predictors in oral squamous cell carcinoma (OSCC), the effectiveness of these approaches has not been assessed clinically. Further, the mechanism underlying malignant behaviors in OSCC is poorly characterized. This study aimed to develop and verify accurate prognostic predictors for OSCC patients and assess the associated biology. We identified an OSCC-recurrence-related gene signature (ORGS) using a Cox regression analysis. Functional enrichment analysis was used to identify enriched pathways and biological processes to reveal the underlying mechanism of OSCC malignant behavior. The ORGS successfully divided OSCC patients into low- and high-risk groups with significantly different overall survivals. Pathway analysis revealed oxidative phosphorylation (OXPHOS) as a signaling pathway associated with the ORGS in OSCC. Interestingly, high OXPHOS status was strongly associated with poor overall survival in OSCC patients. Mediator complex subunit 30 (MED30) was a predicted upstream regulator of OXPHOS, and knockdown of MED30 reduced histone acetylation. We identified that the ORGS was strongly correlated with OXPHOS regulatory processes, suggesting OXPHOS as a key mechanism leading to poor prognosis in OSCC.

NADH elevation during chronic hypoxia leads to VHL-mediated HIF-1α degradation via SIRT1 inhibition.

BACKGROUND: Under conditions of hypoxia, cancer cells with hypoxia inducible factor-1α (HIF-1α) from heterogeneous tumor cells show greater aggression and progression in an effort to compensate for harsh environmental conditions. Extensive study on the stability of HIF-1α under conditions of acute hypoxia in cancer progression has been conducted, however, understanding of its involvement during the chronic phase is limited. METHODS: In this study, we investigated the effect of SIRT1 on HIF1 stability in a typical chronic hypoxic conditon that maintains cells for 24 h under hypoxia using Western blotting, co-IP, measurement of intracellular NAD + and NADH levels, semi-quantitative RT-PCR analysis, invasion assay, gene knockdown. RESULTS: Here we demonstrated that the high concentration of pyruvate in the medium, which can be easily overlooked, has an effect on the stability of HIF-1α. We also demonstrated that NADH functions as a signal for conveyance of HIF-1α degradation via the SIRT1 and VHL signaling pathway under conditions of chronic hypoxia, which in turn leads to attenuation of hypoxically strengthened invasion and angiogenic activities. A steep increase in the level of NADH occurs during chronic hypoxia, leading to upregulation of acetylation and degradation of HIF-1α via inactivation of SIRT1. Of particular interest, p300-mediated acetylation at lysine 709 of HIF-1α is recogonized by VHL, which leads to degradation of HIF-1α via ubiquitin/proteasome machinary under conditions of chronic hypoxia. In addition, we demonstrated that NADH-elevation-induced acetylation and subsequent degradation of HIF-1α was independent of proline hydroxylation. CONCLUSIONS: Our findings suggest a critical role of SIRT1 as a metabolic sensor in coordination of hypoxic status via regulation of HIF-1α stability. These results also demonstrate the involvement of VHL in degradation of HIF-1α through recognition of PHD-mediated hydroxylation in normoxia and p300-mediated HIF-1α acetylation in hypoxia.

SIRT1 interacts with and protects glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from nuclear translocation: implications for cell survival after irradiation.

Upon apoptotic stimulation, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a cytosolic enzyme normally active in glycolysis, translocates into the nucleus and activates an apoptotic cascade therein. In the present work, we show that SIRT1 prevents nuclear translocation of GAPDH via interaction with GAPDH. SIRT1 depletion triggered nuclear translocation of cytosolic GAPDH even in the absence of apoptotic stress. Such translocation was not, however, observed when SIRT1 enzymatic activity was inhibited, indicating that SIRT1 protein per se, rather than the deacetylase activity of the protein, is required to inhibit GAPDH translocation. Upon irradiation, SIRT1 prevented irradiation-induced nuclear translocation of GAPDH, accompanied by interaction of SIRT1 and GAPDH. Thus, SIRT1 functions to retain GAPDH in the cytosol, protecting the enzyme from nuclear translocation via interaction with these two proteins. This serves as a mechanism whereby SIRT1 regulates cell survival upon induction of apoptotic stress by means that include irradiation.

Cells with dysfunctional telomeres are susceptible to reactive oxygen species hydrogen peroxide via generation of multichromosomal fusions and chromosomal fragments bearing telomeres.

During genotoxic stress, reactive oxygen species hydrogen peroxide (H(2)O(2)) is a prime mediator of the DNA damage response. Telomeres function both to assist in DNA damage repair and to inhibit chromosomal end-to-end fusion. Here, we show that telomere dysfunction renders cells susceptible to H(2)O(2), via generation of multichromosomal fusion and chromosomal fragments. H(2)O(2) caused formation of multichromosomal end-to-end fusions involving more than three chromosomes, preferentially when telomeres were erosive. Interestingly, extensive chromosomal fragmentation (yielding small-sized fragments) occurred only in cells exhibiting such multichromosomal fusions. Telomeres were absent from fusion points, being rather present in the small fragments, indicating that H(2)O(2) cleaves chromosomal regions adjacent to telomeres. Restoration of telomere function or addition of the antioxidant N-acetylcysteine prevented development of chromosomal aberrations and rescued the observed hypersensitivity to H(2)O(2). Thus, chromosomal regions adjacent to telomeres become sensitive to reactive oxygen species hydrogen peroxide when telomeres are dysfunctional, and are cleaved to produce multichromosomal fusions and small chromosomal fragments bearing the telomeres.

Integrated analysis of prognostic gene expression profiles from hepatitis B virus-positive hepatocellular carcinoma and adjacent liver tissue.

BACKGROUND: The tissue environment in the region of hepatocellular carcinoma (HCC) influences both vascular invasion and recurrence. Thus, HCC patient prognosis depends on the characteristics not only of the tumor but also those of adjacent surrounding liver tissue. MATERIALS AND METHODS: Expression profiles of both tumor and adjacent liver tissue following curative resection were measured to discriminate 56 hepatitis B virus-positive HCC patients into subgroups based on survival risk. This approach was further tested in 40 patients. RESULTS: Expression profiles of both tumor and adjacent liver tissue successfully discriminated 56 training samples into 2 subgroups, those at low- or high-risk for survival and recurrence. However, the prognostic gene set selected for tumor tissue was quite different from that for adjacent tissues. This variation in prognostic genes resulted in a change in allocation of patients within each low- or high-risk group. Combination of survival subgroups from tumor and adjacent liver tissue significantly improved the prediction of prognostic outcome. This integrative approach was confirmed to be effective in a further 40 test patients. A clinicopathological study showed that survival subgroups divided by tumor and adjacent liver tissue gene expression were also statistically associated with UICC stage and extent of cell differentiation, respectively. CONCLUSIONS: Variation in gene expression during the nontumor stage as well as the tumor stage may affect the prognosis of HCC patients, and integration of the gene expression profiles of HCC and adjacent liver tissue increases discriminatory effectiveness between patient groups, predicting clinical outcomes with enhanced statistical reliability.

Prognostic Significance of the BIRC2-BIRC3 Gene Signature in Head and Neck Squamous Cell Carcinoma.

BACKGROUND/AIM: Head and neck squamous cell carcinoma (HNSCC) has poor prognosis, with survival rates that have not significantly improved over the past several decades. Therefore, prediction of HNSCC prognosis is of clinical importance. Baculoviral IAP Repeat containing 2 (BIRC2) and Baculoviral IAP Repeat containing 3 (BIRC3) are involved in oncogenic activity by modulating cell proliferation, apoptosis and invasion in HNSCC. This study aimed to develop and validate a predictive gene signature for BIRC2 and BIRC3. MATERIALS AND METHODS: The genomic copy number and gene expression for BIRC2 and BIRC3 were systematically explored in patients with HNSCC to investigate the clinical relevance of BIRC2 and BIRC3 activation. A prognostic signature was developed based on correlations associated with BIRC2 and BIRC3 mRNA expression and copy number alterations. Hierarchical clustering was used to classify the clusters (Clusters 1 and 2). Moreover, independent validation of the BIRC2-BIRC3 gene signature was performed using the Leipzig, MDACC, FHCRC, and KHU datasets. To explore the biological functions of the BIRC2-BIRC3 gene signature, string analysis and pathway annotation were also performed. RESULTS: BIRC2-BIRC3 gene signature-derived cluster 2 patients exhibited significantly poor survival. This signature also predicted survival in three independent cohorts. Interestingly, the BIRC2-BIRC3 gene signature additionally permitted the identification of survival in advanced tumor stages with excellent accuracy in all three cohorts. Multivariate Cox regression analysis identified that the BIRC2-BIRC3 signature was an independent predictor associated with the survival of patients with HNSCC. Moreover, Inhibition of BIRC2 modulated the NF-B signaling pathway via upregulation of CBR1 expression. CONCLUSION: The BIRC2-BIRC3 gene signature was found to be associated with the prognosis of HNSCC. Thus, BIRC2 and BIRC3 could be potential targets for improving HNSCC prognosis.

NANOG confers resistance to complement-dependent cytotoxicity in immune-edited tumor cells through up-regulating CD59.

Cancer immunoediting drives the adaptation of tumor cells to host immune surveillance. Previously, we have demonstrated that immunoediting driven by cytotoxic T lymphocytes (CTLs) enriches NANOG+ tumor cells with immune-refractory properties. Here, we found that CTL-mediated immune pressure triggered cross-resistance of tumor cells to the complement system, a part of the innate immune system. In this process, NANOG upregulated the membrane-bound complement regulatory protein (mCRP) CD59 through promoter occupancy, thereby contributing to the resistance of tumor cells against complement-dependent cytotoxicity (CDC). Notably, targeting of NANOG sensitized the immune-refractory tumor cells to trastuzumab-mediated CDC. Collectively, our results revealed a possible mechanism through which selection imposed by T-cell based immunotherapy triggered complement-resistant phenotypes in the tumor microenvironment (TME), by establishing a firm molecular link between NANOG and CD59 in immune-edited tumor cells. We believe these results hold important implications for the clinical application of CDC-mediated therapeutic antibody.

Clinical significance of FAT1 gene mutation and mRNA expression in patients with head and neck squamous cell carcinoma.

The FAT1 gene functions as a tumor suppressor or promoter and remains incompletely understood. We examined the clinical significance of FAT1 in head and neck squamous cell carcinoma (HNSCC) using four publicly available HNSCC cohorts and one HNSCC cohort enrolled at a tertiary medical center. We developed FAT1 signatures reflecting FAT1 mutations and mRNA expression using one cohort. Patients with HNSCC were classified into FAT1-associated low risk (FAT1-LR; n = 195) and FAT1-associated high risk (FAT1-HR; n = 371) subgroups. The five-year overall survival and recurrence-free survival rates were significantly lower in the FAT1-HR subgroup than in the FAT1-LR subgroup (P = 0.01 and 0.003, respectively). The clinical significance of FAT1 was validated using four independent cohorts. Cox proportional hazards models showed that the FAT1 signature was an independent prognostic factor for HNSCC patients. In addition, FAT1 signature was associated with the response to radiotherapy, advanced stage, and human papilloma virus (HPV) status in HNSCC patients. In conclusion, the FAT1 gene signature was associated with prognosis of HNSCC and may help to provide personalized treatments for HNSCC patients.

Changes of Pepsin Concentration in Saliva Sample According to Storage Period.

OBJECTIVES: To determine whether the concentration of pepsin in the saliva sample changes according to the storage period of the sample. METHODS: Forty eight patients with suspected laryngopharyngeal reflux were included in this study. Saliva samples were collected from each patient and each sample divided into six and stored for different period of time. Pepsin concentration was measured using enzyme-linked immunosorbent assay. A comparison was made between the pepsin concentration measured immediately and the concentration measured after storage for 1 week, 2 weeks, 1 month, 3 months, and 6 months. RESULTS: No significant difference in pepsin concentrations were detected between the sample analyzed immediately and those analyzed at 1 week, 2 weeks, 1 month, and 3 months after saliva collection. A significant difference in pepsin concentration was observed in the sample analyzed immediately and the sample analyzed 6 months after saliva collection. CONCLUSIONS: Pepsin concentration in saliva samples did not demonstrate a significant difference between the concentration measured immediately and the concentration measured 3 months after saliva collection, although a significant difference was observed in the concentration measured 6 months after collection.

Paclitaxel stimulates chromosomal fusion and instability in cells with dysfunctional telomeres: implication in multinucleation and chemosensitization.

The anticancer effect of paclitaxel is attributable principally to irreversible promotion of microtubule stabilization and is hampered upon development of chemoresistance by tumor cells. Telomere shortening, and eventual telomere erosion, evoke chromosomal instability, resulting in particular cellular responses. Using telomerase-deficient cells derived from mTREC-/-p53-/- mice, here we show that, upon telomere erosion, paclitaxel propagates chromosomal instability by stimulating chromosomal end-to-end fusions and delaying the development of multinucleation. The end-to-end fusions involve both the p- and q-arms in cells in which telomeres are dysfunctional. Paclitaxel-induced chromosomal fusions were accompanied by prolonged G2/M cell cycle arrest, delayed multinucleation, and apoptosis. Telomere dysfunctional cells with mutlinucleation eventually underwent apoptosis. Thus, as telomere erosion proceeds, paclitaxel stimulates chromosomal fusion and instability, and both apoptosis and chemosensitization eventually develop.