Circulating cell-free DNA as predictor of treatment failure after neoadjuvant chemo-radiotherapy before surgery in patients with locally advanced rectal cancer.

Background: Treatment of patients with locally advanced rectal cancer (LARC) is based on a combination of chemo-radiotherapy (CRT) and surgery. The rate of distant recurrences remains over 25%. Circulating cell-free DNA (cfDNA) in plasma is a mixture of normal and cancer-specific DNA segments and is a promising biomarker in patients with colorectal cancer. The aim of our study was to investigate plasma cfDNA as a prognostic marker for outcome in patients with LARC treated with neoadjuvant CRT and surgery. Patients and methods: In total, 123 patients with LARC were included in 2 biomarker studies. Patients were treated with neoadjuvant CRT before TME surgery. Fifty-two (42%) of the patients received induction chemotherapy with capecitabine + oxaliplatin. Total cfDNA was measured by direct fluorescent assay in EDTA plasma samples obtained at baseline, after induction chemotherapy, and after CRT. Serial samples 5 years after surgery were collected in 51 patients (41%). Results: Median follow-up was 55 months. Distant or local recurrence was seen in 30.9% of the patients. Patients with baseline cfDNA levels above the 75th quartile had a higher risk of local or distant recurrence and shorter time to recurrence compared with patients with plasma cfDNA below the 75th percentile (HR = 2.48, 95% CI: 1.3-4.8, P = 0.007). The same applied to disease-free survival (DFS) (HR = 2.43, 95% CI: 1.27-4.7, P = 0.015). In multivariate analysis, a high cfDNA level was significantly associated with time to progression and DFS. During follow-up, the association remained significant regardless of time point for sample analysis. Conclusion: We have demonstrated an association between a high baseline plasma level of cfDNA and increased risk of recurrence, shorter time to recurrence, and shorter DFS in patients with LARC. Consequently, cfDNA could potentially improve pre- and post-treatment risk assessment and facilitate individualized therapy for patients with LARC.

Prolonged labour as indication for emergency caesarean section: a quality assurance analysis by criterion-based audit at two Tanzanian rural hospitals.

OBJECTIVE: To audit the quality of obstetric management preceding emergency caesarean sections for prolonged labour. DESIGN: A quality assurance analysis of a retrospective criterion-based audit supplemented by in-depth interviews with hospital staff. SETTING: Two Tanzanian rural mission hospitals. POPULATION: Audit of 144 cases of women undergoing caesarean sections for prolonged labour; in addition, eight staff members were interviewed. METHODS: Criteria of realistic best practice were established, and the case files were audited and compared with these. Hospital staff were interviewed about what they felt might be the causes for the audit findings. MAIN OUTCOME MEASURES: Prevalence of suboptimal management and themes emerging from an analysis of the transcripts. RESULTS: Suboptimal management was identified in most cases. Non-invasive interventions to potentially avoid operative delivery were inadequately used. When deciding on caesarean section, in 26% of the cases labour was not prolonged, and in 16% the membranes were still intact. Of the women with genuine prolonged labour, caesarean sections were performed with a fully dilated cervix in 36% of the cases. Vacuum extraction was not considered. Amongst the hospital staff interviewed, the awareness of evidence-based guidelines was poor. Word of mouth, personal experience, and fear, especially of HIV transmission, influenced management decisions. CONCLUSION: The lack of use and awareness of evidence-based guidelines led to misinterpretation of clinical signs, fear of simple interventions, and an excessive rate of emergency caesarean sections.

Muscle bleeds in professional athletes--diagnosis, classification, treatment and potential impact in patients with haemophilia.

Muscle haematoma represents 10-25% of bleeds in patients with severe haemophilia. There is limited consensus on diagnostic or treatment strategies and little knowledge about the natural history of muscle haematoma and optimal treatment goals. The aim of this review was to perform a systematic description of the natural history of muscle haematoma in healthy athletes, focusing on diagnosis, classification and treatment options. Publications and educational textbooks on management of sports injuries were used as data source. Muscle haematomas occur following contusion, strain, or laceration and can be categorized as mild, moderate, or severe. Muscle haematoma may be inter- or intramuscular. In healthy athletes, the healing process takes 20-40 days. Optimal diagnosis includes history, physical examination (inspection, palpation, active and passive range of motion (ROM) test, muscle length test, isometric strength test, biomechanical examination, full spinal examination, peripheral nerve test and slump test), ultrasound, MRI or CT. Treatment is conducted based on: (i) super-acute stage, control of the bleeding and minimizing the size of the haematoma; (ii) acute stage, restoration of pain-free ROM; (iii) subacute stage, functional rehabilitation; and (iv) gradual return to normal activity. Treatment and preventive strategies include RICE (rest, ice, compression and elevation), protected mobilization, stretching and strengthening exercises, manual therapy (articular, neural and soft tissue mobilization and massage), correction of movement dysfunction, functional rehabilitation and electro-therapeutic interventions. The study reviews the natural history of muscle haematoma and state-of-the-art diagnosis and treatment in healthy athletes. Results may be useful to optimize diagnosis and treatment of muscle haematoma in patients with haemophilia.

Evaluating the conjugated linoleic acid and trans 18:1 isomers in milk fat of dairy cows fed increasing amounts of sunflower oil and a constant level of fish oil.

The objective was to evaluate different levels of sun-flower oil (SFO) in dairy rations to increase vaccenic (trans-11-18:1) and rumenic acids (cis-9,trans-11-18:2) in milk fat, and assess the content and composition of other trans-octadecenoic (trans-18:1) and conjugated linoleic acids (CLA) isomers. Eighty lactating Holstein cows were fed control diets for 4 wk and then placed on 4 diets for 38 d; milk fat was analyzed after 10 and 38 d. The treatments were: control, 1.5% SFO plus 0.5% fish oil (FO), 3% SFO plus 0.5% FO, and 4.5% SFO plus 0.5% FO. The forage-to-concentrate ratio was 50:50 and consisted of barley/alfalfa/hay silage and corn/barley grain concentrate. There were no differences in milk production. Supplementation of SFO/FO reduced milk fat compared with respective pretreatment periods, but milk protein and lactose levels were not affected. There was a linear decrease in all short- and medium-chain saturated fatty acids (SFA) in milk fat after 10 d (25.5, 24.1, 20.2, and 16.7%) and a corresponding linear increase in total trans-18:1 (5.2, 9.1, 14.1, and 21.3%) and total CLA (0.7, 1.9, 2.4, and 3.9%). The other FA in milk fat were not affected. Separation of trans-18:1 isomers was achieved by combination of gas chromatography (GC; 100-m highly polar capillary column) and prior separation of trans FA by silver ion-thin layer chromatography followed by GC. The CLA isomers were resolved by a combination of GC and silver ion-HPLC. The trans-11- and trans-10-18:1 isomers accounted for approximately 50% of the total trans-18:1 increase when SFO/FO diets were fed. On continued feeding to 38 d, trans-11-18:1 increased with 1.5% SFO/FO, stayed the same with 3%, and declined with 4.5% SFO/FO. Rumenic acid showed a similar pattern on continued feeding as trans-11-18:2; levels increased to 0.43, 1.5, 1.9, and 3.4% at 10 d and to 0.42, 2.15, 2.09, and 2.78% at 38 d. Rumenic acid was the major CLA isomer in all 4 diets: 66, 77, 78 and 85%. The CLA isomers trans-7,cis-9-, trans-9,cis-11-, trans-10,cis-12-, trans-11,trans-13-, and trans-9,trans-11-/trans-10,trans-12-18:2 also increased from 0.18 (control) to 0.52% (4.5% SFO/FO). Milk fat produced from 3% SFO/FO appeared most promising: trans-11-18:1 and cis-9,trans-11-18:2 increased 4.5-fold, total SFA reduced 18%, and moderate levels of trans-10-18:1 (3.2%), other trans-18:1 (6.6%) and CLA isomers (0.5%) were observed, and that composition remained unchanged to 38 d. The 4.5% SFO/FO diet produced higher levels of trans-11-18:1 and cis-9,trans-11-18:2, a 28% reduction in SFA, and similar levels of other trans-18:1 (9.2%) and CLA isomers (0.52%), but the higher levels of trans-11-18:1 and cis-9,trans-11-18:2 were not sustained. A stable milk fat quality was achieved by feeding moderate amounts of SFO (3% of DM) in the presence of 0.5% FO that had 4% vaccenic and 2% rumenic acids.

Biodegradability of metronidazole, olaquindox, and tylosin and formation of tylosin degradation products in aerobic soil--manure slurries.

The use of veterinary drugs (primarily antibiotics) in animal husbandry harbors the risk that these compounds end up in the farmland when manure is used as fertilizer. The biodegradability of three compounds, olaquindox (OLA), metronidazole (MET), and tylosin (TYL), was simulated in soil--manure slurries with 50 g of soil per liter. Supplemental batch sorption tests revealed that insignificant amounts of OLA and MET were located in the soil phase, whereas only 0.1 to 10% of the added amounts of TYL remained in the liquid phase. This may reduce the bioavailability and thus biodegradation rates of TYL. Unidentified metabolites of OLA and TYL and four known TYL metabolites were detected using HPLC. However, none of these substances were seen to persist in the biodegradation experiments, indicating that OLA and TYL most likely were mineralized in the experiments. Neither the use of sandy or clayey soil nor the use of 0, 1, or 10% (V/V) of manure added to these soils had a significant effect on the degradation rates. Degradation half-lives for the primary degradation were 3.3--8.1 days for TYL, 5.8--8.8 days for OLA, and 13.1--26.9 days for MET. Based on comparisons of results obtained with the benchmark chemical aniline and degradation half-lives of this compound in nature, it was assessed that results obtained with the current test method slightly overestimate real-world biodegradation rates.

Biodegradability of the molluscicidal saponins of Phytolacca dodecandra.

The biodegradability of water-extracted saponins of berries from the Endod plant, Phytolacca dodecandra L'Herit, was evaluated under OECD standardized conditions. Persistence of the saponins was evaluated by determination of saponin concentrations in water over a 30-day period, using a quantitative HPLC method and a semiquantitative hemolytic assay, which is considered a potential field method. The two methods were compared. Bioassays were simultaneously conducted using Biomphalaria glabrata fresh water snails to assess the molluscicidal potency over time. All experiments were carried out in water, which was chemically balanced to sustain snails, and to one set river water was added to the medium to mimic natural conditions. (1) Saponin concentrations in water treated with an aqueous extract of P. dodecandra cultivar E44 were stable for 2 days then rapidly decreased during the third and fourth day. Lethal concentrations to snails were LC50 = 9.6 mg/l (95% CI: 6.3 to 19.4) in one set and LC50 = 6.8 mg/l (95% CI: 5.4 to 12.2) in the other. Adding river water to one set of the experiments had no effect on the mean saponin concentration over time (F = 0.02, P = >0.05). (2) Comparison of the hemolytic assay with the HPLC method showed no significant difference in mean saponin concentrations (t = 0.32, P > 0.05, 95% CI: -2.67 to 3.64), and a correlation of r2 = 0.88 between the two methods. (3) The saponin fraction of an aqueous extract of P. dodecandra was readily biodegraded (t1/2 = 15.8 h), and the complete consumption within a 10-day window indicates ready degradability in aquatic environments under aerobic conditions. (4) These results show that the use of Endod berries for snail control in schistosomiasis-infested water bodies is environmentally acceptable.

Discovery of a series of pyrrolidine-based endothelin receptor antagonists with enhanced ET(A) receptor selectivity.

Endothelins, ET-1, ET-2, and ET-3 are potent vasoconstricting and mitogenic 21-amino acid bicyclic peptides, which exert their effects upon binding to the ET(A) and ET(B) receptors. The ET(A) receptor mediates vasoconstriction and smooth muscle cell proliferation, and the ET(B) receptor mediates different effects in different tissues, including nitric oxide release from endothelial cells, and vasoconstriction in certain vascular cell types. Selective antagonists of endothelin receptor subtypes may prove useful in determining the role of endothelin in various tissue types and disease states, and hence as therapeutic agents for such diseases. The pyrrolidine carboxylic acid A-127722 has been disclosed as a potent and ET(A)-selective antagonist, and is currently undergoing clinical trials. In our efforts to find antagonists with altered selectivity (ET(A)-selective, ET(B)-selective, or nonselective), we investigated the SAR of the 2-substituent on the pyrrolidine. Compounds with alkyl groups at the 2-position possessed ET(A) selectivity improved over A-127722 (1400-fold selective), with the best of these compounds showing nearly 19,000-fold selectivity.

The ability of hypoxia to modify the gene expression of thymidylate synthase in tumour cells in vivo.

PURPOSE: Hypoxic cells in tumours are resistant to 5-fluorouracil (5-FU). This in vivo study investigated the ability of hypoxia to regulate the gene expression of thymidylate synthase (TS), the target enzyme of 5-FU. MATERIALS AND METHODS: C3H mammary carcinomas, grown in the feet of female CDF1 mice, were used for all experiments. Mice were placed in a 10% oxygen environment for various time periods and the tumour oxygen status was determined with an Eppendorf oxygen electrode. The animals were then injected with BrdU (100 mg/kg, i.p.). Tumours were excised and immediately frozen (-80 degrees C) until isolation of total RNA. The mRNA was reversibly transcribed to complementary DNA and the resulting cDNA amplified in a multiplex PCR reaction, with beta-actin as the internal reference gene. RESULTS: One hour of low oxygen breathing made tumours significantly more hypoxic. This increase was maintained for a maximum incubation period of 48 h. In the same tumours, no change in TS gene expression was seen with up to 3 h of low oxygen breathing. At longer times it decreased, reaching significance at 12-24 h and remaining at this lower level for up to 48 h. BrdU labelling was significantly reduced after breathing low O2 for 24 h (p = 0.001). CONCLUSION: Hypoxia-induced down-regulation of TS gene expression was observed. This would be expected to make hypoxic tumour cells more sensitive to 5-FU. Other mechanisms must be responsible for the previously reported resistance to this drug.

A novel MLL-AF10 fusion mRNA variant in a patient with acute myeloid leukemia detected by a new asymmetric reverse transcription PCR method.

A novel variant of the chimerical MLL-AF10 mRNA transcript was detected in a pediatric patient with acute myeloid leukemia (AML) by a new asymmetric reverse-transcription polymerase chain reaction (ART-PCR) method. Sequence analysis of the fusion region on the amplified cDNA fragment showed an in-frame joining of exon e5 of the MLL gene and position 1931 of the cDNA sequence of the AF10 gene, giving rise to a new MLL-AF10 transcript. The presence of the new chimerical mRNA product in a sample from the patient was confirmed by classical RT-PCR.

The cardioprotector ADR-529 and high-dose epirubicin given in combination with cyclophosphamide, 5-fluorouracil, and tamoxifen: a phase I study in metastatic breast cancer.

The purpose of this study was to determine the maximal tolerable dose (MTD) of epirubicin and ADR-529 given in combination with cyclophosphamide, 5-fluorouracil, and tamoxifen. A total of 64 breast cancer patients with locally advanced disease or a first metastatic event were included. Using fixed doses of cyclophosphamide, 5-fluorouracil, and tamoxifen, cohorts of ten patients were treated with escalating doses of epirubicin and ADR-529. With the use of protocol criteria specifying evaluation after the first course, the MTD was not reached. Dose reductions carried out due to hematologic toxicity during the first four courses made it impossible to escalate doses of epirubicin beyond 80 mg/m2 given together with ADR-529 600 mg/m2. The vascular toxicity of ADR-529 necessitated central venous access in a number of patients. For phase III evaluation of ADR-529 given together with cyclophosphamide, epirubicin, 5-fluorouracil, and tamoxifen (CEF/TAM) we recommend using epirubicin/ADR-529 at 60/600 mg/m2. Together with evaluation of the cardioprotective properties of ADR-529, we recommend evaluating the impact of ADR-529 on the efficacy of cytotoxic therapy and investigating further the toxicity of ADR-529.

The pharmacokinetics of high-dose epirubicin and of the cardioprotector ADR-529 given together with cyclophosphamide, 5-fluorouracil, and tamoxifen in metastatic breast-cancer patients.

A high-pressure liquid chromatographic method for determination of the bisdioxopiperazine derivative ADR-529 (ICRF-187), a compound proven effective in protection against anthracycline-induced cardiotoxicity, has been developed. The limit of quantitation was 5 ng/ml using a narrow-bore 5-microns silica column and UV detection. The method was used for determination of pharmacokinetic profiles of ADR-529 after a 3-weekly i.v. administration of different doses of ADR-529 (600-1000 mg/m2) together with different doses of epirubicin (E, 60-100 mg/m2), fixed-dose cyclophosphamide (C, 600 mg/m2), fixed-dose 5-fluorouracil (F, 600 mg/m2), and daily administration of tamoxifen (T, 30 mg; CEF-T) in the treatment of patients with metastatic breast cancer. Pharmacokinetic parameters for epirubicin were also determined. The aim of the study was to determine (1) whether the pharmacokinetics of ADR-529 as part of a combination with CEF-T changes with increasing doses of ADR-529 and increasing doses of epirubicin and (2) whether the pharmacokinetics of epirubicin in the same combinations is altered with the administration of increasing doses of ADR-529. A total of 82 patients were included. A crossover study including 16 of the patients showed no significant difference in epirubicin pharmacokinetic parameters when epirubicin was given with or without concomitant administration of ADR-529. Apart from minor changes in the distributional half-lives, the pharmacokinetic parameters of epirubicin were not altered with increasing doses of ADR-529, nor were the pharmacokinetic parameters of ADR-529 itself. Escalating doses of epirubicin did not significantly alter the pharmacokinetic parameters of ADR-529 with the exception of a 30% increase in the terminal half-life and a decrease in total body clearance when the epirubicin dose was raised from 60 to 100 mg/m2. We conclude that concomitant administration of ADR-529 does not alter the distribution and elimination of epirubicin in doses suitable for preventing the anthracycline-induced cardiotoxicity.

Peribulbar anesthesia. Effect of bicarbonate on mixtures of lidocaine, bupivacaine, and hyaluronidase with or without epinephrine.

The pH-adjustment of local anesthetic solutions with sodium bicarbonate may shorten onset time and improve spread of neural blockade. The authors undertook a prospective, double-masked, randomized study to see if a pH-adjusted mixture of lidocaine, bupivacaine, and hyaluronidase had faster and more complete onset of neural blockade, when used for peribulbar anesthesia. Eighty patients were randomly assigned to four groups and received a peribulbar block with one of four mixtures: group 1 (L) = 2% lidocaine, group 2 (LPH) = 2% lidocaine with 0.06 meq/ml sodium bicarbonate, group 3 (LE) = 2% lidocaine with 1:100,000 epinephrine (commercially prepared), or group 4 (LEPH) = 2% lidocaine with 1:100,000 epinephrine with 0.06 meq/ml sodium bicarbonate. To 5 ml of each of the preceding groups, 5 ml of 0.75% bupivacaine and 150 units of hyaluronidase was added. After each block, extraocular muscle movement was followed in each quadrant until akinesia developed. In the event of incomplete akinesia, blocks were supplemented at 20 minutes. The LPH group had the fastest onset to complete akinesia (7.0 +/- 2.0 minutes, mean +/- SEM) when compared with the onset time of all other groups (group 1 = 11.5 +/- 1.9 minutes, group 4 = 13.1 +/- 1.4 minutes, and group 3 = 16.0 +/- 1.8 minutes, significance greater than 95% by analysis of variance). Furthermore, when compared with group 3 by analysis of variance, group 4 had a faster onset time. The authors conclude that pH-adjustment of solutions with bicarbonate of either lidocaine/bupivacaine/hyaluronidase or commercially prepared lidocaine with epinephrine/bupivacaine/hyaluronidase decreases the onset time of peribulbar anesthesia.

Management information system for leprosy eradication programme--an alternative information system.

For efficient monitoring of multidrug therapy programmes for leprosy both at microlevel (individual patient monitoring) as well as microlevel (programme monitoring), DANIDA decided to develop an alternative, simple and quick information system using a computer. A patient data base system was designed using dBase III Plus package. The field workers of the National Leprosy Eradication Programme were trained in transcribing data on to coded data sheets. The data of 1750 patients of six leprosy control units from the 4 MDT districts were processed and feedback reports were sent to paramedical workers and programme managers. The initial experience in the field over the past year has shown that a computerized management information system is feasible and well accepted by the field staff for the purpose of improving monitoring.

The Copenhagen case-control study of bladder cancer. II. Effect of coffee and other beverages.

During the years 1979-1981 a population-based case-control study of bladder cancer including papilloma was performed in greater Copenhagen. A total of 371 patients (280 males; 91 females), and a comparable age- and sex-stratified group of 771 controls (577 males; 194 females) remained for logistic regression analysis. Controls were selected at random from the general population of the study area. All persons were questioned about their drinking habits with respect to coffee, tea and other beverages, as well as their exposure to a number of known or suspected risk factors for bladder cancer. After adjustment for tobacco smoking, the relative risk of bladder cancer in relation to coffee drinking was not statistically significant among either men or women. A significant association was found between bladder cancer and tea drinking among men, but with no regular trend for increasing consumption. An association was found between risk of bladder cancer and both total daily liquid intake and non-cola soft drinks. This population-based case-control study provides no evidence of an isolated influence of coffee drinking or caffeine intake on bladder cancer risk.

Cardiac effects of vecuronium and pancuronium during halothane anaesthesia.

The cardiac effects of equipotent doses of pancuronium and vecuronium (Org NC 45) were compared in 20 patients, anaesthetized with halothane. Heart rate and arterial pressure were recorded and the systolic time intervals were evaluated from simultaneous tracings of ECG, phonocardiogram and carotid pulse curve. Pancuronium 0.08 mg kg-1 caused a significant increase in heart rate (20%), insignificant changes in arterial pressure and significant changes in systolic time intervals compatible with a minor positive inotropic action. The equipotent dose of vecuronium (0.057 mg kg-1) caused no significant changes in heart rate, arterial pressure or systolic time intervals.

Supraphysiological glucocorticoid treatment in hypoxic stress in dogs.

Effects of supraphysiologic methylprednisolone succinate doses on systolic time intervals, hemodynamic performance, and metabolism were studied in normal dogs during normoxia, after 60 min of normocapnic hypoxemia (PAO2 = 30 mm Hg), and reoxygenation. Bolus injections of 30 or 100 mg/kg methylprednisolone did not exert any positive inotropic action during normoxia. Cardiovascular and metabolic changes induced by hypoxia were modified only to a modest and statistically insignificant degree by sustained high levels of methylprednisolone.