Effects of Daprodustat, a Novel Hypoxia-Inducible Factor Prolyl Hydroxylase Inhibitor on Anemia Management in Japanese Hemodialysis Subjects.

BACKGROUND: Daprodustat (GSK1278863) is an oral hypoxia-inducible factor prolyl hydroxylase inhibitor being developed for treatment of anemia associated with chronic kidney disease (CKD). The effect of daprodustat in Japanese CKD patients with anemia has not been previously investigated. METHODS: We evaluated the relationship between daprodustat dose and hemoglobin response in Japanese patients on hemodialysis (HD) with anemia in a 4-week, phase II, double-blind, placebo-controlled study. After interrupting their erythropoiesis-stimulating agent for between 2 and 8 weeks, subjects with hemoglobin 8.5-10.5 g/dL were randomized to placebo or daprodustat 4, 6, 8, or 10 mg orally once daily. Hemoglobin, erythropoietin (EPO), and vascular endothelial growth factor (VEGF) levels during therapy were evaluated. RESULTS: Eighty-six of 97 randomized subjects completed the study. Mean baseline hemoglobin ranged from 9.68 to 9.92 g/dL across groups. After 4-week administration, mean hemoglobin changes were -0.28, -0.01, 0.54, and 0.97 g/dL in the 4, 6, 8, and 10 mg groups, respectively, as compared to -1.41 g/dL for placebo. Dose-dependent increase in plasma EPO concentration were observed up to 8 mg, with the 10 mg dose responses being similar to 8 mg. Plasma VEGF concentrations were minimally changed, even though 5 subjects treated with 6-10 mg reached EPO >500 mIU/mL. CONCLUSION: Daprodustat 4-10 mg once-daily produced dose-dependent increase in hemoglobin relative to placebo in Japanese HD subjects. The doses evaluated in the study have moderately increased endogenous EPO without changes in circulating VEGF levels.

Occurrence of Deep Vein Thrombosis among Hospitalized Non-Surgical Japanese Patients.

OBJECTIVE: To estimate the frequency of deep vein thrombosis (DVT) among non-surgical inpatients, and to evaluate the D-dimer assay as a screening tool for DVT. METHODS: Subjects were non-surgical inpatients aged 20 years or older who had been bedridden for at least 24 hours and had moderate-to-high risk factors for DVT. We assessed the presence of DVT by venous ultrasonography. Patients who received a diagnosis of venous thromboembolism (VTE) before admission, who had symptoms or findings of VTE at admission, or who had surgery or trauma within the past 3 months before admission were excluded. RESULTS: DVT was confirmed in 96 of 525 patients (18.3%). In a logistic regression analysis, longer duration of hospitalization, higher D-dimer value, and history of cancer surgery were significantly associated with the occurrence of DVT. The D-dimer assay showed high sensitivity (96.1%) and high negative predictive value (97.6%). CONCLUSION: Non-surgical inpatients with a long-term hospitalization or history of cancer surgery have a risk for DVT, and need to be considered for added DVT preventive measures as recommended in the prevention guidelines. In addition, the D-dimer assay is beneficial for the screening of DVT in medical practice.

Frequency of deep vein thrombosis among hospitalized non-surgical Japanese patients with congestive heart failure.

PURPOSE: Congestive heart failure (CHF) is one of the risk factors for deep vein thrombosis (DVT) according to the Japanese guidelines for DVT treatment and prevention. The purpose of this study is to estimate the frequency of DVT among hospitalized CHF patients, since there have been only limited DVT data in Japanese CHF patients. METHODS: Patients enrolled in the study were with risk factors for DVT listed in the guidelines as well as with acute exacerbation of CHF, bed rest for at least 4 days, and aged 60 or above. Patients treated by physical prophylaxis or anti-platelet medication were included, while patients treated by any anticoagulant medicines were excluded. Patients with surgery or injury within 3 months before the hospitalization or diagnosed clinically or with obvious past history as having DVT at hospitalization were excluded. The presence of DVT in the eligible patients was determined by ultrasonography and the images were evaluated by an independent central evaluation committee. RESULTS: Forty-four patients were enrolled in the study including 19 males and 25 females. The mean age was 79.0±10.6 years, and the mean duration of bed rest was 8.9±3.2 days. Out of these 44 patients, DVT was detected in 15 (34%) patients. Eight patients were on treatment with physical prophylaxis but DVT was still detected in two patients. Furthermore, 12 out of the rest of the patients were treated by anti-platelet agents and were still with DVT in 3 patients. CONCLUSION: When evaluated ultrasonographically, the frequency of DVT in hospitalized non-surgical Japanese patients with CHF was approximately 35%. DVT occurred in 25% of patients treated by physical prophylaxis or anti-platelet agents. The results suggest that Japanese hospitalized patients with CHF have a high risk of DVT and thus can be recognized to have potential benefit by preventing and treating DVT according to the guidelines.

Presence of minute cancer cell dissemination in peritoneal lavage fluid detected by reverse transcription PCR is an independent prognostic factor in patients with resectable pancreatic cancer.

BACKGROUND: Presence of minute cancer cell dissemination in peritoneal lavage fluid detected by reverse transcription polymerase chain reaction (RT-PCR) has been reported to be a reliable predictor of the prognosis in several kinds of cancers, but has not been determined in pancreatic cancer. METHODS: Peritoneal lavage fluid was harvested just after a laparotomy in 83 patients with adenocarcinoma of the pancreas. Half of the fluid was examined by cytology and the remaining half was used to measure carcinoembryonic antigen/beta-2-microglobulin (beta2M) mRNA expression. Patients were followed after surgery to evaluate its clinical significance. RESULTS: Among 83 patients, 3 were cytologically positive (CY+), while 23 were positive by RT-PCR (PCR+). Seventy-one patients underwent a surgical resection whereas 12 were unresectable. Because 2 were CY+ among the 71 operated patients, the remaining 69 CY- patients were further investigated. Among those 69 patients, PCR+ was observed in 15 patients, whose incidence of postoperative peritoneal recurrence was significantly higher than that in PCR- patients (21% vs 4% at 3 years; P = .039). Moreover, both the recurrence-free rate in the abdominal cavity (peritoneal or local recurrence, excluding liver metastases) and the overall survival rate were better in PCR- patients than PCR+ patients (78% vs 33%, P = .0045 and 67% vs 46%, P = .0151). A multivariate analysis revealed positive lymph node metastases (hazard ratio; 5.18) and positive RT-PCR (hazard ratio; 3.65) were independent prognostic factors. CONCLUSION: The RT-PCR-based cancer cell detection was an independent prognostic factor in patients with resectable adenocarcinoma of the pancreas and had close association with local or peritoneal recurrence.

Molecular detection of lymph node metastases in breast cancer patients: results of a multicenter trial using the one-step nucleic acid amplification assay.

PURPOSE: Accurate assessment of metastasis in sentinel lymph nodes (SLN) of breast cancer is important but involves a heavy workload for the pathologist. We conducted a multicenter clinical trial in Japan to evaluate a new automated assay system for cytokeratin 19 mRNA, the one-step nucleic acid amplification (OSNA) assay (Sysmex), to detect lymph node metastasis of breast cancer. EXPERIMENTAL DESIGN: Surgically obtained axillary lymph nodes were sectioned into four pieces, two of which were examined with the OSNA assay. The other two adjacent pieces were examined with H&E and immunohistochemical staining for cytokeratin 19. Serial sections at 0.2-mm intervals were used in trial 1 to determine the specificity of the OSNA assay, and three pairs of sections cut from the sliced surfaces of the pieces were used in trial 2 to compare the accuracy of the OSNA assay with that of a routine pathologic examination for SLNs in Japan. RESULTS: In trial 1, the sensitivity and specificity were 95.0% [95% confidence interval (95% CI), 75.1-99.9%] and 97.1% (95% CI, 91.8-99.4%), respectively, for 124 axillary lymph nodes obtained from 34 patients. In trial 2, the agreement between findings of the assay and of the pathologic examination was 92.9% (95% CI, 90.1-95.1%) for 450 axillary lymph nodes obtained from 164 patients. CONCLUSION: The OSNA assay can detect lymph node metastasis as accurately as can conventional pathology and thus can be an effective addition to or alternative for rapid intraoperative examination of SLNs.

Evaluation of the lateral sentinel node by indocyanine green for rectal cancer based on micrometastasis determined by reverse transcriptase-polymerase chain reaction.

The significance of dissecting the lateral pelvic lymph node (LN) for lower rectal cancer remains controversial. We detected the lateral sentinel node (SN) by indocyanine green (ICG) and micrometastases using carcinoembryonic antigen (CEA)-specific reverse transcriptase-polymerase chain reaction (RT-PCR). Twenty-five patients who underwent curative surgery with a dissection of the lateral pelvic LNs between 2003 and 2005 were examined. We investigated the existence of lateral SNs and any associations between pathological metastases and micrometastases by RT-PCR. Lateral SNs were detected in 7 (28%) of the 25 patients. The number of lateral SNs was 13 LNs, or 1.9 nodes per case. Of the 25 cases, 7 had lateral LN metastases based on pathological examinations in dissected lateral LNs. Three cases had massive lateral LN swelling by pre-operative pelvic CT and the SNs were not detected in them. The SNs were detected in two cases and were negative based on pathological examinations and positive according to a genetic diagnosis. SNs were detected in one case, which was positive based on pathological examinations and a genetic diagnosis. SN was not detected in one case. There were five SNs in which CEA was positive by RT-PCR, though only one of them was positive based on pathological examinations. No SNs were observed that were negative based on a genetic diagnosis, but were positive according to the pathological diagnosis. We detected the lateral SNs using ICG. The sensitivity of identifying lateral LN metastasis was improved by the use of a genetic diagnosis. However, the detection rate was still low, therefore we need to develop a new method for detecting SNs.

One-step nucleic acid amplification for intraoperative detection of lymph node metastasis in breast cancer patients.

PURPOSE: Detection of sentinel lymph node (SLN) metastasis in breast cancer patients has conventionally been determined by intraoperative histopathologic examination of frozen sections followed by definitive postoperative examination of permanent sections. The purpose of this study is to develop a more efficient method for intraoperative detection of lymph node metastasis. EXPERIMENTAL DESIGN: Cutoff values to distinguish macrometastasis, micrometastasis, and nonmetastasis were determined by measuring cytokeratin 19 (CK19) mRNA in histopathologically positive and negative lymph nodes using one-step nucleic acid amplification (OSNA). In an intraoperative clinical study involving six facilities, 325 lymph nodes (101 patients), including 81 SLNs, were divided into four blocks. Alternate blocks were used for the OSNA assay with CK19 mRNA, and the remaining blocks were used for H&E and CK19 immunohistochemistry-based three-level histopathologic examination. The results from the two methods were then compared. RESULTS: We established CK19 mRNA cutoff values of 2.5 x 10(2) and 5 x 10(3) copies/muL. In the clinical study, an overall concordance rate between the OSNA assay and the three-level histopathology was 98.2%. Similar results were obtained with 81 SLNs. The OSNA assay discriminated macrometastasis from micrometastasis. No false positive was observed in the OSNA assay of 144 histopathologically negative lymph nodes from pN0 patients, indicating an extremely low false positive for the OSNA assay. CONCLUSION: The OSNA assay of half of a lymph node provided results similar to those of three-level histopathology. Clinical results indicate that the OSNA assay provides a useful intraoperative detection method of lymph node metastasis in breast cancer patients.

Ghrelin reduction after esophageal substitution and its correlation to postoperative body weight loss in esophageal cancer patients.

BACKGROUND: Body weight loss is observed commonly after esophagectomy with gastric tube reconstruction in thoracic esophageal cancer patients. The functional and anatomical alteration of the stomach by this surgery should affect ghrelin secretion, a novel gastric hormone that upregulates body weight through appetite control and metabolic reaction. METHODS: Early-phase postoperative alteration of serum ghrelin was measured before and at day 3 and day 7 after surgery in 9 patients. With 26 other patients, who had previously undergone surgery from 3 months to 67 months (mean, 25 months) before the present study period, the late-phase postoperative alteration of serum ghrelin was investigated along with postoperative body weight loss and serum leptin. RESULTS: Serum ghrelin concentration, which was equivalent to the control group before surgery (88.6 fmol/mL vs 97.5 fmol/mL) significantly decreased by half at 3 and 7 days after surgery. Thereafter, the serum ghrelin decline continued in the outpatients within 1 year after surgery (58.8 fmol/mL), while it was marginal in those from 1 to 3 years after surgery (77.2 fmol/mL). Serum ghrelin was significantly higher than the control after 3 years (185.1 fmol/mL). Thus, a significant positive correlation was observed between ghrelin and time after surgery (P < .0001). Postoperative body weight loss was significant, averaged as DeltaBMI - 2.7 in the outpatients (P < .0001). Until 3 years after surgery, a significant correlation between ghrelin and postoperative body weight loss was observed (P = .0152), with those having higher serum ghrelin showing less body weight loss. Serum leptin correlated well with body weight (P = .0144), but not with postoperative time, the degree of body weight loss, or serum ghrelin concentration. CONCLUSION: Gastric tube replacement for esophagectomy resulted in temporary reduction of ghrelin production, which is associated with body weight loss after surgery. The decline of ghrelin may play some role in the serious body weight loss after esophagectomy, thus encouraging clinical application of exogenous recombinant ghrelin for these patients.

Postoperative ghrelin levels and delayed recovery from body weight loss after distal or total gastrectomy.

BACKGROUND: Body weight loss is a common but one of the most serious sequelae after gastrectomy. Ghrelin, a novel gastric hormone that up-regulates body weight through appetite control and metabolic reaction, seems to be affected by gastric surgery. PATIENTS AND METHODS: Early postoperative alteration of serum ghrelin was evaluated at days 3 and 7 after gastrectomy for 13 hospital patients. In 64 outpatients who had previously undergone total gastrectomy (TG: 26 patients) or distal gastrectomy (DG: 38 patients) 4.6 months to 136 months (average, 41 months) earlier, the association between their serum ghrelin and leptin levels and postoperative body weight was investigated. RESULTS: Serum ghrelin declined immediately and greatly after TG to 12% of the preoperative level (day 3 and day 7), whereas the decline was less significant after DG at 39% (day 3) and 56% (day 7). In outpatients, serum ghrelin after TG was very low compared with the control (18.6 fmol/mL versus 92.1 fmol/mL, P<0.0001), irrespective of the period after surgery, whereas the level after DG recovered and was equivalent to the control (73.4 fmol/mL, P=0.355). Body weight loss was more apparent in TG patients than in DG patients, showing postoperative reduction of body mass index (DeltaBMI) -3.940 versus -1.949 (P<0.0001). Serum leptin concentration, reflecting the systemic fat volume, significantly correlated with BMI in both TG and DG patients, and tended to be lower in TG patients than in DG patients (800 pg/mL versus 1158 pg/mL, P=0.236). CONCLUSION: Persistent decline of serum ghrelin and body weight was observed commonly after total gastrectomy. Further study is needed as to whether or not ghrelin administration can improve the body weight level for these patients.

Cloning and reporter analysis of human mitochondrial phosphoenolpyruvate carboxykinase gene promoter.

Phosphoenolpyruvate carboxykinase (PEPCK) is one of the key regulatory enzymes in gluconeogenesis. In human liver, PEPCK is about equally distributed in both cytosol (PEPCK-1) and mitochondria (PEPCK-2). The human pepck2 gene and cDNA have been reported, but the cloning of the promoter region of the pepck2 gene has not been elucidated yet. We isolated and characterized human genomic P1-artificial chromosome (PAC) clones carrying the human pepck2 gene promoter. The oligocapping method revealed that the transcriptional start point (tsp) of the human pepck2 gene is located at 97 bp upstream of the first adenine residue of the translation start site. We also determined the nucleotide sequence to 1819 bp upstream of tsp. Sequence analysis of this region revealed that it contained several potential regulatory elements, including five GC boxes and three CCAAT boxes. Reporter analysis using transient transfection with firefly luciferase synthetic gene indicated 5' flanking region up to 822 bp, and 317 bp upstream of tsp had transcriptional activity. These results suggest that these regions of the human pepck2 gene play an important role for its expression.