Inhibition of the ATR kinase enhances 5-FU sensitivity independently of nonhomologous end-joining and homologous recombination repair pathways.

The anticancer agent 5-fluorouracil (5-FU) is cytotoxic and often used to treat various cancers. 5-FU is thought to inhibit the enzyme thymidylate synthase, which plays a role in nucleotide synthesis and has been found to induce single- and double-strand DNA breaks. ATR Ser/Thr kinase (ATR) is a principal kinase in the DNA damage response and is activated in response to UV- and chemotherapeutic drug-induced DNA replication stress, but its role in cellular responses to 5-FU is unclear. In this study, we examined the effect of ATR inhibition on 5-FU sensitivity of mammalian cells. Using immunoblotting, we found that 5-FU treatment dose-dependently induced the phosphorylation of ATR at the autophosphorylation site Thr-1989 and thereby activated its kinase. Administration of 5-FU with a specific ATR inhibitor remarkably decreased cell survival, compared with 5-FU treatment combined with other major DNA repair kinase inhibitors. Of note, the ATR inhibition enhanced induction of DNA double-strand breaks and apoptosis in 5-FU-treated cells. Using gene expression analysis, we found that 5-FU induced the activation of the intra-S cell-cycle checkpoint. Cells lacking BRCA2 were sensitive to 5-FU in the presence of ATR inhibitor. Moreover, ATR inhibition enhanced the efficacy of the 5-FU treatment, independently of the nonhomologous end-joining and homologous recombination repair pathways. These findings suggest that ATR could be a potential therapeutic target in 5-FU-based chemotherapy.

[18F]fluoro-2-deoxyglucose-positron emission tomography for the assessment of histopathological response after preoperative chemoradiotherapy in advanced oral squamous cell carcinoma.

BACKGROUND: [(18)F]fluoro-2-deoxyglucose-positron emission tomography (FDG-PET) is widely used to evaluate tumor metabolic activity. The aim of this study was to evaluate the usefulness of FDG-PET in assessing the histopathological response to preoperative concurrent chemoradiotherapy (CRT) in patients with oral squamous cell carcinoma (OSCC). METHODS: Forty-five patients with resectable advanced OSCC who had received preoperative CRT followed by tumor ablative surgery between January 2004 and December 2011 were included in the study. All patients underwent FDG-PET before and after preoperative CRT. The maximum standardized uptake value (SUVmax) before (pre-SUV) and after preoperative CRT (post-SUV) and the SUVmax reduction rate (ΔSUV %) were used to evaluate the response to preoperative CRT. Correlations among SUVmax, histopathological response, and expression of cancer antigen Ki-67 and hypoxia-inducible factor-1α (HIF-1α) were analyzed. RESULTS: Preoperative CRT significantly reduced intratumoral FDG uptake (P < 0.001). The pre-SUV and post-SUV were significantly lower in patients with a pathological complete response (pCR) than in those with a non-pCR (pre-SUV P = 0.037; post-SUV P = 0.001). ΔSUV % was higher in patients with pCR than in those with non-pCR (P = 0.029). The pre-SUV was significantly correlated with Ki-67 and HIF-1α expression in pretreatment biopsy specimens (Ki-67 P = 0.046, R = 0.292; HIF-1α P = 0.007, R = 0.385). The expression of both Ki-67 and HIF-1α was significantly lower in patients with pCR than in those with non-pCR (Ki-67 P < 0.001; HIF-1α P < 0.001). CONCLUSIONS: Low pre-SUV and post-SUV and high ΔSUV % may predict a good histopathological response to preoperative CRT. Ki-67 and HIF-1α expression in pretreatment biopsy specimens were predictors of histopathological response to preoperative CRT.

Volume-based 18F-fluorodeoxyglucose positron emission tomography/computed tomography parameters correlate with delayed neck metastasis in clinical early-stage oral squamous cell carcinoma.

OBJECTIVE: There is no known preoperative marker that can effectively predict the risk of delayed neck metastasis (DNM), which is an important factor that determines the prognosis of early-stage oral cancer. In this study, we examined whether 18F-fluorodeoxyglucose positron emission tomography (18F-FDG-PET)/computed tomography (CT) uptake parameters of primary cancer can predict the risk of DNM in early-stage oral squamous cell carcinoma (OSCC). METHODS: Data from patients with stage I-II OSCC who underwent surgical resection of the primary tumor without elective neck dissection between January 2009 and December 2016 were retrospectively reviewed. Patient characteristics, histopathological factors, and PET/CT parameters (maximum standardized uptake value [SUVmax], metabolic tumor volume [MTV], and total lesion glycolysis [TLG]) were evaluated for their association with DNM. DNM rates were calculated, and the parameters that were statistically significant in the univariate analysis were used as explanatory variables. Independent factors associated with DNM were identified using multivariate analysis. For all statistical analyses, p-values < 0.05 were considered statistically significant. RESULTS: Data from 71 patients were analyzed in the study. The overall DNM rate among all patients was 21.8%. The univariate analysis showed that the T classification, depth of invasion, pattern of invasion, lymphovascular invasion, SUVmax, MTV, and TLG were significant predictors of DNM. However, the multivariate analysis revealed that only the depth of invasion, MTV, and TLG were independent predictors of DNM. CONCLUSION: This study suggests that, in addition to conventional predictors, volume-based PET parameters are useful predictors of DNM in those with early-stage OSCC.

[A case of advanced upper gingival carcinoma responding completely to concurrent chemoradiotherapy with S-1].

We report a case of advanced upper gingival carcinoma with a contralateral metastatic lymph node invading the maxillary sinus (T4aN2cM0). An 83-year-old man was treated concurrently with chemoradiotherapy and S-1. S-1 (80 mg/body/day) was administered for 2 weeks followed by a 1-week rest period as one course. Radiation therapy involved a total of 60 Gy (2 Gy/day; 5 days/week). There were side effects of mild leucopenia and a grade 2 stomatitis. After the completion of 2 courses and radiation therapy, the primary tumor disappeared, and the patient achieved a pathologically complete response. The metastatic lymph node also completely disappeared. S-1 was then administered in the same regimen for 1 year. Neither local recurrence nor distant metastasis has been detected 2 years after the completion of the concurrent chemoradiotherapy with S-1.

Relationship between soluble MICA and the MICA A5.1 homozygous genotype in patients with oral squamous cell carcinoma.

NK and cytotoxic T cells play an important role in the elimination of virus-infected and tumor cells through NKG2D activating receptors, which can promote the lysis of target cells by binding to the major histocompatibility complex class I-related chain A (MICA) proteins. Polymorphisms in MICA may influence its binding to the NKG2D. The soluble form of MICA is released from the surface of tumor cells of epithelial origin. Whereas MICA expressed on the cell surface stimulates the immunoreceptor natural killer group 2, member D (NKG2D), the secreted form down-regulates NKG2D activity, thus allowing the tumor to escape immunosurveillance by NKG2D-expressing cells. In this study, we examined the association between MICA gene microsatellite polymorphisms and serum levels of soluble MICA in patients with oral squamous cell carcinoma (OSCC). We found that patients with OSCC were more likely to have the A5.1 allele when compared to healthy subjects and also more likely to be homozygous for this allele (p=0.041). Patients with the homozygous A5.1 genotype had higher levels of soluble MICA (p=0.031) and a lower survival rate (p=0.026).

Relationship Between EGFR Expression in Oral Cancer Cell Lines and Cetuximab Antibody-dependent Cell-mediated Cytotoxicity.

BACKGROUND/AIM: Cetuximab treatment targets the epidermal growth factor receptor (EGFR) overexpressed in oral cancer. This study aimed to investigate the anti-tumour activity of cetuximab against oral cancer cell lines with respect to antibody-dependent cell-mediated cytotoxicity (ADCC), and determine the correlation between ADCC and EGFR expression. MATERIALS AND METHODS: EGFR expression in oral cancer cells was measured by quantitative flow cytometric analysis and immunohistochemistry. ADCC activity was measured by 4-h calcein release assays. RESULTS: Cetuximab-mediated ADCC against oral cancer cells was detectable at a concentration of 0.1 µg/ml. A high correlation was observed between the number of EGFR molecules on the surface of oral cancer cells and ADCC (correlation coefficient: 0.847; p=0.032). CONCLUSION: ADCC is an important mechanism underlying the therapeutic effect of cetuximab, and EGFR expression in tumour cells might serve as a predictive marker to evaluate the effect of cetuximab treatment.

Association between soluble MICA levels and disease stage IV oral squamous cell carcinoma in Japanese patients.

The soluble form of major histocompatibility complex class I-related chain A (MICA) is released from the surface of tumor cells of epithelial origin. Although MICA expressed on the cell surface stimulates the immunoreceptor natural killer (NK) group 2, member D (NKG2D), the secreted form downregulates NKG2D activity, thus allowing the tumor to escape immunosurveillance by NKG2D-expressing cells. In this study, we examined the association between serum levels of soluble MICA and the severity of disease in patients with oral squamous cell carcinoma (OSCC). We used enzyme-linked immunoabsorbent assay to measure serum levels of soluble MICA in OSCC patients and normal control individuals. Among patients categorized according to most disease parameters tested (tumor size, location, grade of differentiation, regional lymph node status, disease stage), soluble MICA levels in sera did not statistically differ from those in normal control individuals. Patients with stage IV disease and/or regional lymph node metastasis did, however, exhibit significantly higher serum levels of soluble MICA than control individuals (95% confidence interval (CI), 0.65-2.45, p = 0.021, and 95% CI, 0.62-4.42, p = 0.031, respectively). Overall survival rates were significantly higher for OSCC patients with low soluble MICA levels (<50 pg/ml) than for those with high soluble MICA levels (>50 pg/ml) (95% CI, 0.43-2.75, p = 0.03). Serum levels of soluble MICA may be useful in the diagnosis of advanced stage OSCC and as an indicator of regional lymph node metastasis.

[The influence of intermittent hemodialysis (HD) on bispectral index (scale) (BIS) and serum propofol concentration under postoperative intravenous sedation].

We experienced a case of postoperative intravenous sedation with propofol during intermittent hemodialysis (HD), and investigated the correlation between the clinical sedation level based on bispectral index scale (BIS) together with Ramsay score, and the serum concentration of propofol. One adult oral cancer patient (carcinoma of the lower gingiva) with end-stage renal dysfunction needing HD was selected for this study. The day after operation, HD was commenced under intravenous sedation with propofol. Clinical sedation level was assessed using BIS and Ramsay score, and serum propofol concentrations were determined in arterial blood samples. Serum concentrations were measured every 15 times until 51 hours after operation. The initial dose of propofol was set at 3.5 mg x kg(-1) x h(-1) on the basis of clinical symptoms. According to BIS and Ramsay score, sedation level decreased lineally for 1 hour after commencement of HD. In contrast, serum propofol concentration incresed from 1.71 microg x ml(-1) to 2.21 microg x ml(-1). Total serum concentration of propofol was enhanced during HD because of dialytic dehydration, but, according to BIS and Ramsay scores, the possibility was suggested that the fraction of albmin-unbound propofol with pharmacological activities was eliminated or absorbed by membrane during HD.

Asphyxial death related to postextraction hematoma in an elderly man.

We here report an autopsy case of a man in his seventies who died from asphyxia due to compression of the trachea caused by postextraction bleeding after extraction of his left mandibular third molar by a dentist in private practice. On the morning after the tooth extraction, he had complained of dyspnea and became unconscious at home. Although he was brought to the emergency room by ambulance, he died 7 days later without regaining consciousness. Autopsy examination revealed that the lingual side of the alveolar bone was fractured at the extraction socket. Moreover, subcutaneous bleeding that extended from the extraction socket to the thyrohyoid ligament in the cervical region and deviation of the epiglottis due to the bleeding were observed. Histological findings revealed liver cirrhosis; there were no significant findings in other organs. On the basis of these findings, we concluded that alveolar bone fracture occurred during the extraction and that the bleeding spread to the cervical region. Thus, the patient had died from asphyxia resulting from airway obstruction caused by cervical subcutaneous bleeding derived from postextraction bleeding. We emphasize that tooth extraction may cause fatal complications in patients with bleeding tendencies, particularly in the elderly.

Autopsy case of a patient with 13q- syndrome.

Partial deletion of the long arm of chromosome 13 results in 13q(-) syndrome, and phenotypes of affected patients vary widely. We describe an autopsy case of the sudden, unexpected death of a 17-year-old boy with 13q(-) syndrome. He had severe psychomotor retardation and had been receiving follow-up care. One day he was found dead in his house and autopsy was performed to elucidate the cause of death. Autopsy findings revealed lobulation anomalies of the lungs, hypoplasia of the adrenal and thyroid glands, and apituitarism due to hyperplasia of bone in the hypophyseal fossa. No other pathological lesions were observed. Chromosomal analysis confirmed interstitial deletion from the long arm of chromosome 13. Karyotype was 46,XY, del(13)(q14.3q32). We concluded that the patient died of multi-organ dysfunction due to apituitarism. Autopsy cases of 13q(-) syndrome are rare. Furthermore, lobulation anomalies and apituitarism associated with 13q(-) syndrome have not previously been described. This case report offers novel clues to elucidating critical regions of chromosome 13 associated with malformations of the lungs and pituitary gland.

Soluble MICB serum levels correlate with disease stage and survival rate in patients with oral squamous cell carcinoma.

BACKGROUND: Expression of ligands of natural killer group 2D (NKG2D) immunoreceptors, such as major histocompatibility complex class I-related chain A/B (MICA/B), has been proposed to play an important role in tumour immunosurveillance. Soluble forms of MICA/B are increased in sera of cancer patients and are postulated to impair antitumour immune response by downregulating expression of NKG2D immunoreceptors. Serum levels of soluble MICA have been shown to be of diagnostic significance in malignant diseases. AIMS: The potential of soluble MICB (sMICB) as a marker for oral squamous cell carcinoma (OSCC) was investigated. RESULTS: sMICB levels did not differ significantly from those in normal control individuals. However, the findings indicate that sMICB levels are significantly increased in stage IV OSCC and high sMICB levels are significantly associated with decreased survival rates in patients.

MICA gene polymorphism not associated with nonsyndromic cleft lip with or without cleft palate in the Japanese population?

UNLABELLED: Nonsyndromic cleft lip with or without cleft palate (NSCLP) is one of the most common birth defects. Despite its frequency, the etiology remains largely unknown. Most likely, both genetic and environmental factors contribute to this malformation. A polymorphic gene family, the major histocompatibility complex class I chain-related gene A (MICA), is located about 40 kb centromeric to the HLA-B gene. In this study, we analyzed the association between MICA gene polymorphisms and NSCLP in Japanese patients. METHODS: The (GCT)n polymorphism of the MICA gene was investigated in 94 patients with NSCLP and 180 normal controls using polymerase chain reaction amplification and denaturing polyacrylamide gel electrophoresis. RESULTS: Our results demonstrate that there are no differences in microsatellite allele frequency between NSCLP patients and controls. However, the microsatellite allele frequency of the MICA-A6 (p = 0.045) allele was increased in male patients, as compared with controls. Further, the MICA-A5 (p = 0.359) allele was also increased in female NSCLP patients. CONCLUSION: These results suggest that the microsatellite allele frequencies of the MICA-A6 allele increased in male NSCLP patients. Although the MICA-A5 allele increased in female NSCLP patients, the increase was not statistically significant. These results suggest that the MICA gene could be one of the candidate genes for NSCLP.

An association between the MICA-A5.1 allele and an increased susceptibility to oral squamous cell carcinoma in Japanese patients.

BACKGROUND: Recently, a new polymorphic gene family called the major histocompatibility complex class I chain-related gene A (MICA) was discovered about 40 kb centromeric to HLA-B gene. The MICA protein, expressed on epithelial cells and many kinds of tumor cells, serves to regulate immune function. The MICA protein is thought to activate immune function on mucosal tissue by binding to NKG2D which is expressed on most natural killer cells, CD8 positive T cells, and gamma delta T cells. An association between MICA gene polymorphisms and the development of oral squamous cell carcinoma (OSCC) has also been reported. OBJECTIVE: This study was designed to test this association in Japanese patients with OSCC. METHODS: The (GCT)(n) polymorphisms of the MICA gene was investigated in 123 patients with OSCC and 188 normal controls using polymerase chain reaction amplification and denaturing polyacrylamide gel electrophoresis. RESULTS: Five alleles, namely A4, A5, A6, A9, and A5.1, were found in both groups. The phenotype frequency of the MICA-A5.1 allele was significantly higher in patients with OSCC when compared with normal controls (OR 1.707, 95% CI 0.76-3.45, P=0.042). Also, the microsatellite frequency of the MICA-A5.1 allele was significantly higher in patients with OSCC compared with normal controls (OR 1.664, 95% CI 0.82-3.42, P=0.021). Lastly, the frequency of the MICA-A5.1 allele was significantly higher in those with lymph node metastasis from OSCC compared with normal controls (OR 2.605, 95% CI 1.14-5.27, P=0.026). CONCLUSIONS: These results suggest that the MICA-A5.1 allele may be associated with an increased susceptibility to OSCC in Japan.