Replication fork blocking deficiency leads to a reduction of rDNA copy number in budding yeast.

The ribosomal RNA genes are encoded as hundreds of tandem repeats, known as the rDNA, in eukaryotes. Maintaining these copies seems to be necessary, but copy number changes in an active manner have been reported in only frogs, flies, Neurospora, and yeast. In the best-studied system, yeast, a protein (Fob1) binds to the rDNA and unidirectionally blocks the replication fork. This block stimulates rDNA double-strand breaks (DSBs) leading to recombination and copy number change. To date, copy number maintenance and concerted evolution mediated by rDNA repeat turnover were the proposed benefits of Fob1-dependent replication fork arrest. In this study, we tested whether Fob1 provides these benefits and found that rDNA copy number decreases when FOB1 is deleted, suggesting that Fob1 is important for recovery from low copy number. We suppose that replication fork stalling at rDNA is necessary for recovering from rDNA copy number loss in other species as well.

Use of a novel screw-type dilator for endoscopic ultrasonography-guided hepaticogastrostomy via 22-Gauge needle and 0.018-inch guidewire.

Tract dilation is an essential step in completing endoscopic ultrasonography-guided hepaticogastrostomy (EUS-HGS). EUS-HGS using a 22-gauge needle is currently attracting attention; however, the complexity of subsequent dilation and guidewire exchange is problematic. A-60-year-old man with duodenal cancer was referred to our center for the drainage of obstructive jaundice. As endoscopic retrograde cholangiopancreatography was not feasible because of duodenal obstruction, EUS-HGS was performed.

Utility of a Passive Bending Colonoscope for Endoscopic Retrograde Cholangiopancreatography in Patients with Surgically Altered Anatomy.

BACKGROUND AND AIM: The utility of a passive bending colonoscope (PBCS) in ERCP for patients with surgically altered anatomy has not been established. This study compared the outcome of PBCS-ERCP and balloon-assisted enteroscope (BAE)-ERCP. METHODS: This multicenter observational study included 343 patients with surgically altered anatomy who underwent ERCP. Among these, 110 underwent PBCS-ERCP and 233 underwent BAE-ERCP. Propensity score matching was applied, and a final cohort of 210 (105 in each group) with well-balanced backgrounds was analyzed. The primary outcome was the success rate of reaching anastomosis or ampulla of Vater. Secondary endpoints included the cannulation success rate, completion rate, procedure time (to reach, cannulate, complete), and adverse events. RESULTS: The success rate for reaching the target was 91.4% (96/105) with PBCS and 90.5% (95/105) with BAE (odds ratio [95% CI] 1.12, [0.44-2.89], P = 0.809). The mean time required to reach the target was significantly shorter in PBCS: 10.04 min (SD, 9.62) with PBCS versus 18.77 min (SD, 13.21) with BAE (P < 0.001). There were no differences in the success of cannulation or procedure completion, although the required times for cannulation and procedure completion were significantly shorter in PBCS. The incidence of adverse events was significantly higher in BAE (19.0%) than in PBCS (4.8%; P < 0.001). CONCLUSIONS: In patients with surgically altered anatomy, PBCS-ERCP showed promising results with shorter time to reach, cannulate, and a lower incidence of adverse events compared with BAE-ERCP. The success rate of reaching was favorable through PBCS compared with BAE. CLINICAL TRIAL REGISTRATION: UMIN000045546.

The risk of weekend biopsy: Impact of specimen source and fixation status on HER2 assessment in the treatment of advanced gastric cancer (The HER_WEEKEND study).

Accurate evaluation of human epidermal growth factor receptor type 2 (HER2) expression is crucial for determining chemotherapy regimens in gastric cancer. However, formalin fixation status has been identified as an important factor affecting HER2 assessment reliability. This retrospective cohort study aimed to investigate the correlation between sample collection day (weekday vs. weekend) and source (biopsy vs. surgical specimens) in assessing HER2 expression in patients with unresectable advanced/recurrent gastric cancer. Data were collected from gastric cancer patients who received chemotherapy at a single public hospital in Japan from 2008 to 2021. The analysis included 177 patients (109 men, 68 women) with a median age of 68.0 (21-88) years, and the primary outcome was the HER2 positivity rate. The overall HER2 positivity rate was 18.1%, with higher rates on weekdays (20.0%) compared to weekends (12.8%). Biopsies had higher positivity rates on weekdays (23.9%) but lower rates on weekends (11.1%) than surgical specimens. Significant differences were observed in formalin fixation times between weekdays and weekends for both biopsies and surgical samples. The study findings suggest that longer formalin fixation times on weekends may lead to underestimating HER2 expression, particularly in biopsies. Therefore, it is crucial to be cautious of excessive formalin fixation when collecting samples, especially during weekend biopsies.

An Autopsy Case of Rapidly Aggravated Clostridium perfringens Septicemia with Colorectal Cancer.

This report presents a case of a 60-year-old man who was diagnosed with ascending colon cancer with metastases of the lymph nodes and multiple liver metastases. Three days before the introduction of the first chemotherapy, he visited our hospital due to high fever. The blood test revealed an increase in the inflammatory response, hepatobiliary enzyme level, lactate dehydrogenase (LDH) level, and renal function deterioration. Contrast-enhanced computed tomography (CT) showed a rapid progression of primary lesion and liver metastatic lesions. Treatment with 5-fluorouracil, leucovorin, and oxaliplatin and cetuximab (FOLFOX/Cmab) was initiated, and the patient was admitted to our hospital after the first day of chemotherapy. At midnight, he had chills, red urine, and rapid hypoxemia. The second blood test showed progression of anemia; increased total bilirubin, aspartate aminotransferase, and LDH levels; and decreased platelet and fibrinogen levels. The serum was red wine in color, indicating marked hemolysis. The respiratory condition rapidly deteriorated, and tracheal intubation was performed and transferred into the intensive care unit. However, blood oxygenation did not increase, and the patient died the next morning, 19 h after admission, despite intensive care. Postmortem CT showed intraperitoneal free air and gas retention in the liver tumor and portal vein system. Pathological autopsy revealed perforation in ascending colon cancer, many Gram-positive rods in the perforation site, dissemination of bacteria throughout the body, and diffuse pulmonary edema. Subsequently, blood cultures reported Clostridium perfringens (CP), which is a product of alpha-toxin. CP infection can cause rapid aggravation and sudden death. The physicians should be aware of this highly fatal infection, leading to immediate diagnosis and treatment.

Incidence of cancer-associated thromboembolism in Japanese gastric and colorectal cancer patients receiving chemotherapy: a single-institutional retrospective cohort analysis (Sapporo CAT study).

OBJECTIVE: Few data regarding the incidence of cancer-associated thromboembolism (TE) are available for Asian populations. We investigated the incidence of TE (TEi) and its risk factors among gastric and colorectal cancer (GCC) patients received chemotherapy in a daily practice setting. DESIGN: A retrospective cohort study. SETTING: A single-institutional study that used data from Sapporo City General Hospital, Japan, on patients treated between January 2008 and May 2015. PARTICIPANTS: Five hundred Japanese GCC patients who started chemotherapy from January 2008 to May 2015. PRIMARY AND SECONDARY OUTCOME MEASURES: TE was diagnosed by reviewing all the reports of contrast-enhanced CT performed during the follow-up period. All types of thrombosis detected by CT or additional imaging tests, such as venous TE, arterial TE and cerebral infarction, were defined as TE. Medical records of all identified patients were reviewed and potential risk factors for TE, including clinicopathological backgrounds, were collected. We defined the following patients as 'active cancer'; patients with unresectable advanced GCC, cancer recurrence during or after completing adjuvant chemotherapy and/or presence of other malignant tumours. RESULTS: Of the 500 patients, 70 patients (14.0%) developed TE during the follow-up period. TEi was 9.2% and 17.3% in GCC patients, 18.1% and 3.5% in active and non-active cancer patients, and 24.0% and 12.9% in multiple and single primary, respectively. Multivariate logistic regression analysis showed that colorectal cancer (CRC) (OR 2.371; 95% CI 1.328 to 4.233), active cancer (OR 7.593; 95% CI 2.950 to 19.543) and multiple primary (OR 2.527; 95% CI 1.189 to 5.370) were independently associated with TEi. CONCLUSION: TEi was 14.0% among Japanese GCC patients received chemotherapy, and was significantly higher among patients with CRC, active cancer and multiple primary than among those with gastric cancer, non-active cancer and single primary, respectively. TRIAL REGISTRATION NUMBER: UMIN000018912.

A Prospective Observational Study on Effect of Short-Term Periodic Steroid Premedication on Bone Metabolism in Gastrointestinal Cancer (ESPRESSO-01).

BACKGROUND: A multicenter prospective observational study evaluated the effect of gastrointestinal cancer chemotherapy with short-term periodic steroid premedication on bone metabolism. PATIENTS AND METHODS: Seventy-four patients undergoing chemotherapy for gastrointestinal cancer were studied. The primary endpoints were changes in bone mineral densities (BMDs) and metabolic bone turnover 16 weeks after initiation of chemotherapy. BMDs, measured by dual-energy x-ray absorptiometry, and serum cross-linked N-telopeptides of type I collagen (sNTX), and bone alkaline phosphatase (sBAP) were assessed for evaluation of bone resorption and formation, respectively. RESULTS: In 74.3% (55/74) of the patients, BMDs were significantly reduced at 16 weeks relative to baseline. The percent changes of BMD were -1.89% (95% confidence interval [CI], -2.67% to -1.11%: p < .0001) in the lumbar spine, -2.24% (95% CI, -3.59% to -0.89%: p = .002) in the total hip, and -2.05% (95% CI, -3.11% to -0.99%: p < .0001) in the femoral neck. Although there was no significant difference in sNTX levels during 16 weeks (p = .136), there was a significant increase in sBAP levels (p = .010). Decreased BMD was significantly linked to number of chemotherapy cycles (p = .02). There were no significant correlations between changes in BMDs and the primary site of malignancy, chemotherapy regimens, total cumulative steroid dose, steroid dose intensity, and additive steroid usage. CONCLUSION: Gastrointestinal cancer chemotherapy with periodic glucocorticoid premedication was associated with reduced BMD and increased sBAP levels, which were linked to number of chemotherapy cycles but independent of primary site, chemotherapy regimen, duration, and additive steroid usage. The Oncologist 2017;22:592-600 IMPLICATIONS FOR PRACTICE: Bone health and the management of treatment-related bone loss are important for cancer care. The present study showed that a significant decrease in bone mineral density (BMD) and an increase in serum bone alkaline phosphatase levels occurred in gastrointestinal cancer patients receiving chemotherapy, which were linked to number of chemotherapy cycles but were independent of primary site, chemotherapy regimen, total steroid dose, and steroid dose intensity. Surprisingly, it seems that the decreasing BMD levels after only 16 weeks of chemotherapy for gastrointestinal cancer were comparable to that of 12-month adjuvant aromatase inhibitor therapy for early-stage breast cancer patients.

Prediction of response to treatment by gene expression profiling of peripheral blood in patients with microscopic polyangiitis.

The JMAAV study was an open-labeled prospective clinical trial, which proposed severity-based treatment protocols for patients with microscopic polyangiitis (MPA). The results suggest that the proposed protocols are useful (remission rate: 89.4%), but are also indicative of relapse or patient demise regardless of the treatment (recurrence rate: 19.0%; mortality rate: 10.6%). The aim of this study is to develop the method to predict response to the treatment in patients with MPA. In the present study, transcriptome analysis was performed using peripheral blood from patients enrolled in the JMAAV study before and 1-week after the beginning of treatment. The gene expression profile before treatment was not directly related to the response to the treatment. However, when the samples from 9 patients with good response (persistent remission for 18 months) were examined, the expression of 88 genes was significantly altered by the treatment. Thirty statistically reliable genes were selected, and then the alteration of expression by the treatment was examined among 22 patients, including 17 with good response, which was defined as persistent remission for 18 months and 5 with poor response, which was defined as relapse after remission or no remission. Discrimination analysis between the alteration of expression of the 30 genes by the treatment and the response identified a combination of 16 genes as the most valuable gene set to predict the response to the treatment. This preliminary study identified IRF7, IFIT1, IFIT5, OASL, CLC, GBP-1, PSMB9, HERC5, CCR1, CD36, MS4A4A, BIRC4BP, PLSCR1, DEFA1/DEFA3, DEFA4, and COL9A2 as the important genes that can predict the response to the treatment in patients with MPA at an early point during the therapy.

Aberrant expressions of HOX genes in colorectal and hepatocellular carcinomas.

HOX genes are known as master regulator genes which give cells positional information in embryogenesis. In this study, we compared the expression patterns of 39 HOX genes among human colorectal carcinomas from the right large intestine (cecum, ascending and transverse colon), those from the left large intestine (discending and sigmoid colon, and rectum) and hepatocellular carcinoma. The expression levels of each HOX gene were quantified by analysis based on the real-time RT-PCR. The expression patterns of HOX genes in colorectal and hepatocellular carcinoma tissues differed from those in their normal or non-cancerous tissues. Between the tumor tissues in the right-side large intestine and those in the left-side, different HOX genes were expressed in association with cancer. Further, the expression levels of HOXD8 in liver-metastatic tissues of colorectal carcinomas were as low as in non-cancerous liver tissues, and were significantly lower than those in the primary tissues. These results suggest that dysregulated expressions of HOX genes play an important role in carcinogenesis and malignant progression of colorectal and hepatocellular carcinomas.

PAX4 has the potential to function as a tumor suppressor in human melanoma.

We hypothesize that dysregulated expression levels of the developmental regulatory genes in the adult body result in tumor development and malignant progression. PAX genes discovered as human orthologous genes of Drosophila 'paired' encode transcription factors, which control the expression of target genes to go on along the program of development. In this study, we first quantified expression of 9 PAX genes in human nevus pigmentosus tissues, melanoma tissues and melanoma cell lines by the real-time reverse transcription-PCR method. As a result, we found that the expression levels of PAX4 and PAX9 were extremely low in melanoma tissues and cell lines compared to nevus pigmentosus tissues. We then established melanoma cells overexpressing PAX4 and examined roles of PAX4 in cell growth. PAX4-overexpression reduced in vitro cell growth of human melanoma C8161 and MeWo cells. BrdU-uptake assay and cell cycle analysis by flow cytometry indicated that the retardation of cell proliferation by PAX4-overexpression was due to decreased DNA synthesis and cell cycle arrest at the G0/G1 phase. Furthermore, treatment of C8161 and MeWo cells with 5-azacytidine, a DNA demethylating agent, induced the expression of PAX4, suggesting that DNA methylation repressed the PAX4 gene expression in human melanoma. These results suggest that PAX4 functions as a potent tumor suppressor.

Genotyping of human papillomaviruses by a novel one-step typing method with multiplex PCR and clinical applications.

We describe here a rapid, high-throughput genotyping procedure that allows the simultaneous detection of 16 high- and low-risk genital human papillomavirus (HPV) types by multiplex PCR in a single reaction tube. Multiplex PCR is based on the amplification of HPV DNA by sets of HPV genotype-specific primers, and the genotypes of HPV are visually identified by the sizes of amplicons after they are separated by capillary electrophoresis. The procedure does not include a hybridization step with HPV-specific probes and is rapid and labor-saving. We detected all 16 HPV genotypes (types 16, 58, 52, 51, 56, 31, 18, 39, 66, 59, 6, 33, 30, 35, 45, and 11) with a high sensitivity and a high degree of reproducibility. By using this newly developed method, we conducted a pilot study to examine the correlation between the prevalence and genotype distributions of HPV and the cytological group classifications for 547 cervical samples. Compared with the group of samples considered normal (14.7%), there was a significant increase in the prevalence of HPV in women with atypical squamous cells of unknown significance (61.3%), low-grade intraepithelial lesions (75.8%), and high-grade intraepithelial lesions (HSILs) (82.2%). The prevalence and distribution of type 58 were correlated with cytological malignancies, with the highest prevalence in women with HSILs. In conclusion, the novel multiplex PCR method described appears to be highly suitable not only for the screening of cervical cancer precursor lesions but also for the characterization of genotype distributions in large-scale epidemiological studies and HPV vaccination trials.

HOXC6 and HOXC11 increase transcription of S100beta gene in BrdU-induced in vitro differentiation of GOTO neuroblastoma cells into Schwannian cells.

HOX genes encode transcription factors that play a key role in morphogenesis and cell differentiation during embryogenesis of animals. Human neuroblastoma cells are known to be chemically induced to differentiate into neuronal or Schwannian cells. In the present study, we investigated the roles of HOX genes in differentiation of GOTO neuroblastoma cells into Schwannian cells. When GOTO cells were grown in the presence of 5-bromo-2'-deoxyuridine (BrdU), they increased the expressions of two HOX genes (HOXC6 and HOXC11) and marker genes for Schwannian cells (S100beta and myelin basic protein). Forced expression of HOXC11 alone or both HOXC6 isoform 1 and HOXC11 induced the expression of S100beta in GOTO cells. In transient transfection experiments, the overexpression of HOXC6 and HOXC11 transactivated the S100beta promoter-reporter construct. Taken together, our results suggest that HOXC6 and HOXC11 are associated with differentiation of GOTO cells into Schwannian cells through the transcriptional activation of S100beta gene.

Aberrant expression of HOX genes in oral dysplasia and squamous cell carcinoma tissues.

Human HOX genes consist of 39 genes and encode transcription factors that function as master developmental regulators. We hypothesized that the misexpression of HOX genes was associated with carcinogenesis and malignant progression. The expression levels of 39 HOX genes in 31 human oral squamous cell carcinoma (SCC), 11 dysplasia, and 10 normal mucosa tissues were quantified by the real-time RT-PCR method. The expression levels of 18 HOX genes in the SCC tissues were significantly higher than those in the normal mucosa tissues. The dysplasia tissues showed higher expression of HOXA2, A3, B3, and D10 than normal mucosa tissues whereas they showed lower expression of HOXA1, B7, B9, and C8 than SCC. The SCC with lymph node metastasis showed high expression of HOXC6 compared to the SCC without it. These results suggest that misexpressions of particular HOX genes are implicated in the development of oral dysplasia and SCC.