Laboratories Ltd Research Articles

Correlation of CD10 immunoexpression and eosinophil count in stromal tissue as predictors of prognosis of squamous cell carcinoma of the oral cavity

Introduction: CD10 can also facilitate the ability of tumour metastasis by its ability to invade blood vessel walls.Correlation of CD10 immunoexpression and eosinophil count in stromal tissue as predictors of prognosis of squamous cell carcinoma of the oral cavity. Methods: This study was preceded by collecting data on squamous cell carcinoma of the oral cavity in the medical record and paraffin blocks in the Anatomy Pathology section of 15 samples with enlarged regional lymph nodes and 15 samples without enlargement of regional lymph nodes. Then two preparations were made, the first preparation was stained with Hematoxylin Eosin (HE) to establish the diagnosis and determine the histopathological gradation, then count the number of eosinophils, and the other development was smeared with CD10. Outward immunohistochemistry uses the Labelled streptavidin biotin immunoperoxidase complex (LSAB) method using the Starr Trek Universal HRP Detection system. (Biocare Medical, USA). The primary antibody used was CD10 (Novocastra Laboratories Ltd, Newcastle upon Tyne, UK) with a dilution of 1:25 - 1: 50. Results: There was no statistically significant correlation between CD10 and enlargement of the lymph node regionally tested statistically using the Spearman rank test rs = -0.236, meaning that the increase in CD10 immunoexpression is inversely proportional to the enlargement of regional lymph node, but the correlation is not statistically significant The results of the analysis using the Spearman rank test found significance of p = 0.043 (significant) with a correlation strength of 37.2% Conclusion: There was no correlation between CD10 and enlargement of the lymph node regionally and correlation strength eosinophil distribution in squamous cell carcinoma of the oral cavity without regional lymph node enlargement.

Presence of free radicals in intracameral agents commonly used during cataract surgery

BackgroundFree radicals are known to cause cellular damage and are present in ophthalmic preparations. Corneal defence mechanisms are bypassed in intra-ocular surgery. We evaluated commonly used intracameral agents to ascertain...

The expression of E-cadherin-catenin complex in patients with advanced gastric cancer: role in formation of metastasis.

The E-cadherin-catenin complex plays an important role in the process of cell adhesion. Its dysfunction is associated with a decrease in cell differentiation and with increased invasiveness and metastasis. Our aim was to evaluate the expression of E-cadherin and B-catenin in advanced gastric cancer in relation to selected clinico-pathomorphological parameters. Formalin-fixed, paraffin-embedded tissue specimens were immunohistochemically stained with monoclonal antibodies E-cadherin (NCL-E-Cad, Novocastra Laboratiries Ltd; dilution 1:50), beta-catenin (NCL-B-CAT, Novocastra Laboratories Ltd; dilution 1:100), alpha-catenin (alpha-E-caten, Santa Cruz Biotechnology; dilution 1:300) and gamma-catenin (gamma-catenin, Santa Cruz Biotechnology; dilution 1:100). The expressions of E-cadherin and alpha-, beta-, gamma-catenins in the main mass of tumor and lymph node metastasis were investigated in 91 patients with gastric cancer. No statistically significant correlation was observed between the expressions of E-cadherin, alpha-, beta-catenins and histological differentiation and between the expressions of E-cadherin, alpha-, gamma-catenins and location or depth of invasion. Moreover, the expression of alpha-, gamma-catenins in the main mass of tumor was not associated with lymph node metastasis. However, we found a relationship between the expression of beta-catenin in the main mass of tumor and lymph node metastasis and tumor location. The depth of invasion was correlated with positive expression of beta-catenin in the main mass of gastric cancer. A statistically significant association was observed between the expressions of E-cadherin and beta-catenin in the main mass of tumor and lymph node involvement. The expression of alpha-catenin in the main mass of tumor was also associated with histological differentiation and Lauren's classification. Statistical analysis showed an association between the expression of E-cadherin and postoperative survival time. No significant correlation was found between the expression of alpha-, beta-, gamma-catenins and survival time. Our results may suggest that the E-cadherin-catenin complex is the factor indicative of metastasis and disease progression in gastric cancer. Also the expression of E-cadherin may play a role as a prognostic factor.

DR REDDY'S LABORATORIES (UK) LTD v ELI LILLY AND CO LTD

DR REDDY’S LABORATORIES (UK) LTD v ELI LILLY AND CO LTD Get access Reports of Patent, Design and Trade Mark Cases, Volume 127, Issue 3, March 2010, Pages 222–255, https://doi.org/10.1093/rpc/rcq020 Published: 01 March 2010

Evaluation of 5 Different Labeled Polymer Immunohistochemical Detection Systems

Immunohistochemical staining is important for diagnosis and therapeutic decision making but the results may vary when different detection systems are used. To analyze this, 5 different labeled polymer immunohistochemical detection systems, REAL EnVision, EnVision Flex, EnVision Flex+ (Dako, Glostrup, Denmark), NovoLink (Novocastra Laboratories Ltd, Newcastle Upon Tyne, UK) and UltraVision ONE (Thermo Fisher Scientific, Fremont, CA) were tested using 12 different, widely used mouse and rabbit primary antibodies, detecting nuclear, cytoplasmic, and membrane antigens. Serial sections of multitissue blocks containing 4% formaldehyde fixed paraffin embedded material were selected for their weak, moderate, and strong staining for each antibody. Specificity and sensitivity were evaluated by subjective scoring and digital image analysis. At optimal primary antibody dilution, digital image analysis showed that EnVision Flex+ was the most sensitive system (P < 0.005), with means of 8.3, 13.4, 20.2, and 41.8 gray scale values stronger staining than REAL EnVision, EnVision Flex, NovoLink, and UltraVision ONE, respectively. NovoLink was the second most sensitive system for mouse antibodies, but showed low sensitivity for rabbit antibodies. Due to low sensitivity, 2 cases with UltraVision ONE and 1 case with NovoLink stained false negatively. None of the detection systems showed any distinct false positivity, but UltraVision ONE and NovoLink consistently showed weak background staining both in negative controls and at optimal primary antibody dilution. We conclude that there are significant differences in sensitivity, specificity, costs, and total assay time in the immunohistochemical detection systems currently in use.

Non-specific interference in the measurement of plasma ammonia: importance of using a sample blank

Enzymatic assays using glutamate dehydrogenase (GLDH) to monitor the transformation of NAD(P)H to NAD(P)(+) by a spectrophotometric technique are the most common methods to measure plasma ammonia (PA) in routine laboratories worldwide. However, these assays can potentially be subject to interference by substances in plasma able to oxidize NAD(P)H at a substantial rate, thereby providing falsely high results. To study this potential interference, we spiked a plasma pool with a liver homogenate and measured the ammonia concentration using a dry chemistry system (Vitros 250, Ortho Clinical Diagnostic, Raritan, NJ, USA), an enzymatic assay without a sample blanking step (Infinity Ammonia Liquid Stable Reagent, Thermo Fisher Scientific, Waltham, USA) and an enzymatic assay that corrects for the non-specific oxidation of NADPH (Ammonia kit, RANDOX Laboratories Ltd, Crumlin, UK). This experiment shows that the Infinity ammonia reagent kit is subject to a clinically significant interference and explains the discrepancies previously reported between these methods in patients with acute liver failure (ALF). When using enzymatic methods for the assessment of PA, we recommend including a sample blanking correction and this should be mandatory when monitoring patients with ALF.

Clinical Evaluation of An in Vitro Assay for Rapid Assessment of Cytosine Arabinoside Response in Patients with Acute Leukaemia and Chronic Lymphocytic Leukaemia.

Abstract 1608▪▪This icon denotes an abstract that is clinically relevant.Poster Board I-634Cytosine arabinoside (Ara-C) is the mainstay of treatment for acute non lymphoblastic leukaemia (ANLL). Currently there is no rapid, inexpensive test to assess patient sensitivity to Ara-C prior to treatment. We have previously reported a bioluminescent 8-hour assay which assesses Ara-CTP levels in KG-1a and THP-1 cell lines (Smith et al., Blood 2005; 106(11): p695a). Here we present results of this assay using six further ANLL cell lines, 45 patients with ANLL, 2 patients with Ph+ acute lymphoblastic leukaemia (ALL) and 12 with B-CLL. Over 30% of patients with ANLL fail to respond to Ara-C. Potential causes of resistance include lack of hENT-1 transporter activity, over expression of cytidine deaminase (cdd) and over expression of Y-family DNA polymerases. The 8-hour assay was initially tested on eight ANLL cell lines with known response to Ara-C (CCRF-CEM, K562, MV-4-11, HEL, KG-1a, HL-60, THP-1, Mo7e) and this correlated with a commercially available 3-day cytotoxicity assay (CellTiterGlo‘, Promega). ANLL patient samples were sourced at presentation from patients with blast burdens >80%. The distribution of FAB sub-types was M4 (26%), M2 (20%), secondary AML (20%), M0 (14%), M1 (6%), M3 (6%), JAK2+ (3%) and Ph+ ALL (5%). Patient ages ranged from 24 to 94 (median 67.5 years) and included 25 peripheral blood and 22 bone marrow samples. The Ara-C concentration used in vitro was 25 μM which is equivalent to 2g/m2 in the clinical setting. The luminescence cut-off for sensitivity was an increase in light production of more than 10%. The 8-hour luminescence assay showed 100% correlation with the 3-day CellTiterGlo” test results, both indicating that 51% of patients were Ara-C sensitive and the remainder resistant. In the 32 patients for whom clinical outcome data was available, assay results correlated with clinical outcome in 30 patients (p<0.05). The remaining two patients showed results which conflicted with clinical outcome and are therefore under further investigation to examine Y-family DNA polymerase expression. Confirmation of the predictive capacity of the 8-hour assay was observed in two patients with M0 ANLL, initially resistant to Ara-C, now corroborated by the assay, who only achieved remission with FLAG-Ida. The bioluminescence assay was therefore modified to allow a 4-hour pre-incubation with fludarabine and showed marked improvement in blast cell sensitivity to combination treatment compared with Ara-C alone. A further 2 patients with Ph+ ALL and 12 patients with B-CLL were assessed using the 8-hour assay. For the Ph+ ALL patients, one entered complete remission and the second failed to respond to treatment. In both cases this was corroborated by the 8-hour assay and the 3-day CellTiterGlo” assay. The 12 B-CLL patients, all exhibited a highly sensitive profile with the 8-hour assay but in contrast showed resistant results with the 3-day CellTiterGlo” assay. This may indicate the ability of B-CLL cells to incorporate and metabolise Ara-C to Ara-CTP yet this may not translate into cytotoxic effect due to over expression of low fidelity DNA polymerases This new assay system may enable response prediction prior to patients receiving Ara-C based chemotherapy. This may be of importance in high risk patients with adverse cytogenetics/immunophenotype who may require fludarabine with Ara-C to achieve response. Similarly patients with a highly sensitive profile may benefit from a lower dose of Ara-C in vivo thus minimising unwanted side effects. In the event of relapse, repeat analysis may give insight into true clonal evolution and acquisition of chemo-insensitivity. DisclosuresRuddock:Randox Laboratories Ltd: Employment. Martin:Randox Laboratories Ltd: Employment. Lamont:Randox Laboratories Ltd: Employment.

Influence of the placebo effect on oxidative stress in healthy older adults of Mexico City

The placebo effect has been widely recognized in the randomized clinical trials; nevertheless, this effect has not been evaluated in terms of antioxidant therapy on oxidative stress (OxS). The objective of this study was to determine the influence of the placebo effect on OxS in healthy older adults of Mexico City. We carried out a double-blind controlled clinical assay with the participation of 75 healthy older adults residents for the past 10 years of Mexico City; randomly distributed into three groups of 25 subjects each after previous informed consent; control group not received any treatment, placebo group received a placebo with a pharmaceutical presentation similar to that of the treatment, whereas treatment group were administered 1000 mg of ascorbic acid and 400 IU of alpha-tocopherol. All subjects ingested the treatment daily according to study group for 12 months. We measured before and after 12 months of treatment, lipoperoxides levels (LPO), erythrocyte superoxide dismutase, glutathione peroxidase and total plasma antioxidant status with Randox Laboratories Ltd kits. The concentration of ascorbic acid and alpha-tocopherol were measured using high-performance liquid chromatography. The placebo group subjects showed a statistically significant decrease in LPO concentration, in the same way as the treatment group subjects (P < 0.01), both in comparison with a control group. Our findings suggest that the placebo has a significant effect on OxS.

Glucosamine sulfate—environmental antibacterial activity

We have recently showed antibacterial activity against E. coli in vitro of a trademark Mega-Gluflex-containing glucosamine sulfate (GS) and chondroitin sulfate (CS). The purpose of this study was to examine the antibacterial activity of GS as a new trademark Arthryl (Manufacturer Rottapharm Ltd, Ireland; Distributor in Israel Rafa Laboratories Ltd) in vitro. We used cabbage and chicken broths and milk (every media of 20 ml) left opened for 1 week with and without Arthryl supplements 1,500 mg, the content of one package of the medication. A similar volume (20 ml) is ingested in taking the medication. Experiments with three repeatable results were taken for consideration. Arthryl inhibited environmental bacterial colonies' growth in every media but fungi growth was not impaired. Milk stayed liquid for the whole week with supplement of the Arthryl compared with sour milk transformation without Arthryl. Sample B showed inhibitory properties of the bacterial colonies on the fungi growth. The sample with Arthryl showed progressive growth of fungi without bacterial growth after 10 days of follow up compared with bacterial growth on media without Arthryl. Glucosamine sulfate as a new trademark Arthryl has environmental antibacterial properties but does not inhibit growth of fungal colonies.

Cost of cataract surgery after implantation of three intraocular lenses

Posterior capsule opacification is one of the most frequent adverse events following cataract surgery. This manuscript reports the lifetime cost of complications linked to posterior capsule opacification using three types of intraocular lens with square edges. Costs were estimated from a retrospective study of patients who underwent cataract surgery and data from the literature. The lenses studied were hydrophobic acrylic (SA60AT and AR40E) and hydrophilic acrylic (XL-Stabi) lenses with square edges. The frequency of Nd-Yag laser capsulotomies after 4 years' survival was estimated by two methods: the first involved linear adjustment of the rate at 5 and 6 years follow-up and then application of a constant rate after 6 years; the second involved linear adjustment after 5 years follow-up. The economic perspective was that of the French Sickness Fund. After 3 years' follow-up the percentage of patients who had not undergone laser Nd-Yag capsulotomy was 86.9% with SA60AT, 76.6% with AR40E and 54.6% with XL-Stabi lenses (p < 0.001). The total cost of capsulotomy and management of complications per patient lifetime was estimated to be euro90.5 for SA60AT, euro189.5 for AR40E and euro288.0 for XL-Stabi lenses by the first extrapolation method. With the second method of extrapolation the costs were euro94.8, euro200.0 and euro300.2, respectively. Lower costs for cataract surgery and management of related complications were observed with the two hydrophobic acrylic lenses; the lowest costs were observed with SA60AT lenses as they were associated with fewer Nd-Yag laser capsulotomies.

Lesions in the Larynx of Wistar RccHanTM: WIST Rats

Specific regions in the rat larynx exhibit cellular changes in response to inhaled xenobiotics. These regions include the base of the epiglottis, ventral pouch, and medial surfaces of the vocal processes of the arytenoid cartilages. 1 , 2 In order to collect information on the usefulness of trimming techniques, the influence of different vehicles, the impact of different application routes in toxicity studies, and differences between induced vs. spontaneous lesions, the data obtained from a large number of inhalation and non-inhalation studies performed in Wistar RCCHanTM: Wist rats at Harlan Laboratories Ltd Switzerland, all evaluated or reviewed by the same pathologist, were compiled for a detailed review. The value of different trimming techniques was deemed to be greatest for transverse and sagittolongitudinal section techniques, as compared to horizontolongitudinally section techniques. The comparison of lesions encountered in control rats of inhalation studies treated with different vehicles did not reveal differences in the type, distribution pattern, incidence and/or severity of spontaneous lesions. The types of lesions were also independent of different application routes in non-inhalation studies compared to inhalation studies. The pattern of spontaneous lesions in the rodent larynx was determined by degenerative and inflammatory lesions starting most often in the submucosal glands by desiccated secretion followed by mineralization and local inflammation or were induced by impacted foreign bodies. Squamous metaplasia was recorded in the respiratory epithelium overlaying the ventral gland as a spontaneous lesion in male Wistar rats from inhalation studies with a maxim of 20.0% in an inhalation oncogenicity study. Induced metaplastic changes recorded in the larynx were reversible. Other induced lesions in inhalation studies consisted of submucosal edema, necrosis, inflammation and/or granuloma. Induced lesions in non-inhalation studies were found to be exclusively related to reflux laryngitis or food impaction. It is concluded, that in rodents induced lesions of the larynx differ in type, distribution pattern, severity and incidence from spontaneous lesions.

Peripartum Haemostatic Changes in Women in Labour

Abstract Introduction: Obstetric haemorrhage (OH) is a major cause of maternal mortality and morbidity worldwide. Appropriate management requires understanding of normal peripartum haemostatic changes during labour, and yet there are few studies. Aim: In this pilot study we aimed to evaluate peripartum haemostatic changes in healthy women. Methods: We collected blood samples from 20 women peripartum, into 0.102 M trisodium citrate tubes (ratio9:1) (Becton Dickinson) from the antecubital fossa using flawless venepuncture at the following time-points: antepartum; after delivery of the placenta; 24hrs post-partum and 48hrs post-partum. Plasma was aliquoted and stored in −70°C until the following ELIZA assays were performed: D-Dimer levels (Dade Behring, Sysmex UK), prothrombin fragments 1+ 2 (PF 1+2) (Dade Behring, Sysmex, UK) and tissue plasminogen activator antigen (t-PA:Ag) (Biopool, Trinity Biotech, UK). Prothrombin time (PT) (PT-Fib HS), activated partial thromboplastin time (APTT) (APTT micronized silica) and clauss fibrinogen (Fib-C), reagents were from Instrumentation Laboratories Ltd UK. Statistical analysis was performed using XLSTAT 2008. Results: The median (range) age was 28 (19–36) years and the body mass index (BMI) 28.9 (21.7– 35.3). Table 1: To show the median (range) for the haemostatic assays Assay (normal ranges) D-dimers (4–52ug/l) PF1+2 (0.2–1.2 pmol/l) t-PA:Ag: 3–11 ng/ml) PT (12–16s) APTT (26–38s) Fibrinogen (1.52–4.12 g/l) * p&amp;lt;0.05 using the Mann Whitney U test between the 1st and each of the other three samples Antepartum 231 (122–900) 570 (352–1533 12 (7.2–23) 12 (10–14) 31 (24–36) 5.2 (3.1–8.7) Post Labour 482 (117–1044) 968 (480–1858)* 18 (8.6–40.6)* 13 (10–15) 30 (24–36) 5.3 (2.2–9.8) 24h 309 (65–931) 568 (264–1080)* 7 (4–23)* 13 (10–16)* 33 (27–41)* 5.3 (4–11) 48hrs 235 (85–614) 648 (371–2287) 7 (3–18)* 12 (10–16) 31 (23–43) 5.7 (3.5–11) Conclusion: These findings demonstrate that peripartum haemostasis is prothrombotic with shortened PT, APTT and prolonged fibrinogen. There is increased thrombin generation and fibrinolytic activity. t-PA antigen is most notably elevated immediately post delivery.

Possibilities of nimesulid administration in pediatric rheumatology

Objective. To analyze nimesulid efficacy and safety in children with reactive and juvenile chronic arthritis. Material and methods. 108 children aged 1,5 to 18 years receiving niomesulid (Doctor Reddi’s Laboratories Ltd, India) from 3,0 to 5,0 mg/kg/day as the main anti-inflammatory drug were included. Results. Duration of continuous treatment with nimesulid varied from 10 days in reactive arthritis to 1 year in juvenile chronic arthritis. Efficacy of the drug was quite comparable with other NSAID in 99,1% of pts with joint diseases. Adverse events appeared in 5,4% of cases (mainly gastrointestinal damage). Allergic skin rush was seen only during suspension administration. Nimesulid monotherapy did not induce hepatotoxic effect. Conclusion. Nimesulid can be recommended as anti-inflammatory drug in children with reactive and juvenile chronic arthritis. The drug has sufficient anti-inflammatory effect and minimum adverse events during long term administration and in combination with disease modifying anti-rheumatic drugs. Age restriction for nimesulid administration can be decreased to 1,5 years.

Endometrial-Peritoneal Interactions during Endometriotic Lesion Establishment

The pathophysiology of endometriosis remains unclear but involves a complex interaction between ectopic endometrium and host peritoneal tissues. We hypothesized that disruption of this interaction would suppress endometriotic lesion formation. We hoped to delineate the molecular and cellular dialogue between ectopic human endometrium and peritoneal tissues in nude mice as a first step toward testing this hypothesis. Human endometrium was xenografted into nude mice, and the resulting lesions were analyzed using microarrays. A novel technique was developed that unambiguously determined whether RNA transcripts identified via microarray analyses originated from human cells (endometrium) or mouse cells (mesothelium). Four key pathways (ubiquitin/proteasome, inflammation, tissue remodeling/repair, and ras-mediated oncogenesis) were revealed, demonstrating communication between host mesothelial cells and ectopic endometrium. Morphometric analysis of nude mouse lesions confirmed that necrosis, inflammation, healing and repair, and cell proliferation occurred during xenograft development. These processes were entirely consistent with the molecular networks revealed by the microarray data. The transcripts detected in the xenografts overlapped with differentially expressed transcripts in a comparison between paired eutopic and ectopic endometria from human endometriotic patients. For the first time, components of the interaction between ectopic endometrium and peritoneal stromal tissues are revealed. Targeted disruption of this dialogue is likely to inhibit endometriotic tissue formation and may prove to be an effective therapeutic strategy for endometriosis.

Abbott Laboratories Ltd v Evysio Medical Devices ULC

Abbott Laboratories Ltd v Evysio Medical Devices ULC

Characterization of tRNA-dependent Peptide Bond Formation by MurM in the Synthesis of Streptococcus pneumoniae Peptidoglycan

MurM is an aminoacyl ligase that adds l-serine or l-alanine as the first amino acid of a dipeptide branch to the stem peptide lysine of the pneumococcal peptidoglycan. MurM activity is essential for clinical pneumococcal penicillin resistance. Analysis of peptidoglycan from the highly penicillin-resistant Streptococcus pneumoniae strain 159 revealed that in vivo and in vitro, in the presence of the appropriate acyl-tRNA, MurM(159) alanylated the peptidoglycan epsilon-amino group of the stem peptide lysine in preference to its serylation. However, in contrast, identical analyses of the penicillin-susceptible strain Pn16 revealed that MurM(Pn16) activity supported serylation more than alanylation both in vivo and in vitro. Interestingly, both MurM(Pn16) acylation activities were far lower than the alanylation activity of MurM(159). The resulting differing stem peptide structures of 159 and Pn16 were caused by the profoundly greater catalytic efficiency of MurM(159) compared with MurM(Pn16) bought about by sequence variation between these enzymes and, to a lesser extent, differences in the in vivo tRNA(Ala):tRNA(Ser) ratio in 159 and Pn16. Kinetic analysis revealed that MurM(159) acted during the lipid-linked stages of peptidoglycan synthesis, that the d-alanyl-d-alanine of the stem peptide and the lipid II N-acetylglucosaminyl group were not essential for substrate recognition, that epsilon-carboxylation of the lysine of the stem peptide was not tolerated, and that lipid II-alanine was a substrate, suggesting an evolutionary link to staphylococcal homologues of MurM such as FemA. Kinetic analysis also revealed that MurM recognized the acceptor stem and/or the TPsiC loop stem of the tRNA(Ala). It is anticipated that definition of the minimal structural features of MurM substrates will allow development of novel resistance inhibitors that will restore the efficacy of beta-lactams for treatment of pneumococcal infection.

Diagnosis of Multidrug‐Resistant Tuberculosis and Extensively Drug‐Resistant Tuberculosis: Current Standards and Challenges

The emergence of multidrug-resistant tuberculosis (MDR-TB) and, more recently, extensively drug-resistant TB (XDR-TB) is widely considered a serious threat to global TB control. Over 400,000 new cases of MDR-TB occur each year and, although their rates are currently unknown, XDR-TB cases have been detected in every country where there is capacity to detect them (including Canada). The present article provides a narrative overview of the various diagnostic options available for XDR-TB, including conventional tools and newer rapid tests for drug resistance. Available data suggest that automated liquid cultures are highly accurate and their use is rapidly expanding. Newly developed phenotypic tests include TK Medium (Salubris Inc, USA), microscopic-observation drug-susceptibility assay, FASTPlaque-Response bacteriophage assay (Biotec Laboratories Ltd, UK), colorimetric redox indicator methods and the microcolony method. These tests are usually cheaper but not always simple to perform, with some requiring high standards of biosafety and quality control. Among the newly developed phenotypic methods, reverse hybridization-based assays, referred to as line probe assays, represent a useful tool because of their superior accuracy and cost-effectiveness. To effectively address the threats of MDR-TB and XDR-TB, global initiatives are required to scale-up culture and drug susceptibility testing capacities, especially in high-burden countries where such capacity is scarce. In parallel, efforts are needed to expand the use of novel and emerging technologies (ie, molecular diagnostics) for the rapid determination of drug resistance.

Pharmacokinetic and Pharmacodynamic Evaluation of a Biosimilar Rituximab in Newly Diagnosed Diffuse Large B-Cell Lymphoma (DLBCL) Treated with R-CHOP (Rituximab, Cyclophosphamide, Adriamycin, Vincristine, Prednisolone).

Addition of rituximab (375 mg/m2) to CHOP has been shown to improve survival in patients with DLBCL. However there is limited data on the pharmacokinetics (PK) and pharmacodynamics (PD) of this drug in this condition. We have evaluated the PK and PD of a biosimilar rituximab (Reditux®, Dr. Reddy's Laboratories Ltd, Hyderabad, India) in 17 patients with newly diagnosed DLBCL treated at a single center with R-CHOP as part of a multi-center study undertaken to assess the safety and efficacy of this drug. R-CHOP (rituximab-375mg/m2; cyclophopsphamide-750mg/m2; adriamycin-50mg/m2; vincristine-1.4mg/m2 on day 1 and prednisolone-60mg/m2 on days 1 to 5) was given every 3 weeks for a total of 6 cycles. Blood samples for measurement of rituximab were collected just prior to start of infusion and 10min, 24, 72, 192 and 360 hours post-infusion for all patients during cycle 1 and in 6 patients during cycle 6 also. Additional samples were collected pre- and 10 minutes post-infusion after cycles 2, 3, 4, 5 and 6. Plasma rituximab levels were quantified using an immunoassay (sensitivity: 1ug/ml). B-lymphocyte counts were measured in peripheral blood samples taken from all patients at the beginning of each cycle. All patients were evaluated for clinical and radiological response after the 2nd, 4th and 6th cycles. Patients, mean age: 52 years (range:31–71) had disease in the following stages: stage II: 5, stage III: 7, stage IV: 5. Twelve out of 17 patients achieved complete remission while 5 had partial response (NCI criteria). At a mean follow-up of 5 months (range: 3–8), 3 patients had relapse of the disease. The arithmetic mean ±SD of PK parameters of Rituximab during cycle 1 were as follows: T½(hrs): 167±63; Cmax(ug/ml): 186±49; Cmin(ug/ml): 22.4±12.84; AUC0-∞ (ug.hrs/ml): 28162±11227) and Cl/F (ml/kg/hr): 23.8±10.8. These data are comparable with values previously reported for rituximab in other conditions. Though a 2–7 fold inter-individual variation was noted among these patients, there was no significant difference in these parameters between those in whom the disease relapsed as opposed to those who maintained remission. Among the 6 patients in whom data was available for the 1st and 6th cycles (table), there was significant reduction in Cl/F with associated increase in Cmax and AUC in the 6th cycle as compared to the 1st cycle. In 16 patients for whom the data was available, pre-treatment mean B lymphocyte count which was 121/ul (range:1.5–410.5) dropped to a mean of 9.9/ul (range:0.3–62.3) after the first cycle and remained in that range for the rest of treatment period. These data show that even with a 3-weekly regimen, therapeutic trough levels (25 μg/ml) of rituximab was observed across all cycles. In fact, the changing PK parameters of the drug with progressive cycles of R-CHOP suggest that fixed-dose regimens may not be the optimal way to administer this drug.ParameterCycle1 (n=6)Cycle 6 (n=6)p valueT ½ (hours)2003860.0481Cmax (μg/ml)2032790.0556Cmin (360Hr)24.7182.160.0028AUC0- ∞ (μg.hrs/ml)31167922400.0049Cl/F(ml/kg/hr)20.57.60.0030

Role of artrofoon in complex treatment of rheumatoid arthritis

Objective. To study influence of artrofoon on clinicolaboratory measures in pts with rheumatoid arthritis (RA). Material and methods. 119 RA pts (97 female, 22 male) were included. 104 of them were seropositive. Mean age was 52,3± 11,2 years, mean disease duration — 6,87+5,89 years. Disease activity level varied from 1 to 3 stage. Most pts had II or III radiological stage of RA. Usual clinical and laboratory examination of pts was performed at baseline and then after 1, 3, 6 and 12 months of treatment. Quantitative examination of serum cytokines was done using biochip technology method of Randox Laboratories Ltd (Great Britain). During the study all pts received standard treatment with disease modifying antirheumatic drugs (DMARDs) and nonsteroidal anti-inflammatory drugs (NSAIDs). The following NSAIDs were used: diclofenac in 54 pts (not more than 100 mg/day), nimesulid 200 mg/day in 37 pts, other NSAIDs - in 28 pts. The pts were divided into 2 groups. 91 pts of group 1 received artrofoon besides standard treatment. 28 pts of group 2 received only standard treatment. Results. Administration of artrofoon was accompanied by significant decrease of tumor necrosis factor a and interleukin (IL)ip levels after 3, 6 and 12 months of treatment, increase of IL4, IL10 and vascular endothelial growth factor to 6th month, decrease of epidermal growth factor to 12th month of treatment in comparison with control. These changes were accompanied by decrease of laboratory (ESR, CRP) and clinical measures of disease activity (VAS, Ritchie index, morning stiffness). Adverse events were absent in the main group of pts. Conclusion. Artrofoon exerted positive influence on the main clinicolaboratory and immunologic measures in pts with RA. Maximal effect was reached after 6 months of treatment. A part of pts decreased NSAID dose during treatment with artrofoon. The drug was well tolerated and safe during 12 months of treatment. Artrofoon can be administered with other drugs used in the treatment of RA.

Evaluation of a rapid screening test for rifampicin resistance in re-treatment tuberculosis patients in the Eastern Cape.

Patients with multidrug-resistant (MDR) tuberculosis (TB) are at high risk of treatment failure. It is anticipated that early identification of MDR-TB and appropriate treatment will improve patient outcome and disease control. We evaluated the rapid detection of rifampicin resistance in previously treated TB patients, directly from acidfast bacilli (AFB)-positive sputum using a phage-based test, FASTPlaque-Response (Biotec Laboratories Ltd, Ipswich, UK). The ability of rifampicin resistance to predict MDR-TB was also determined. A prospective study was done comparing performance of the rapid phage test with conventional culture and drug susceptibility testing (DST) in AFB-positive TB patients. Five primary health clinics and one TB referral centre in the Port Elizabeth Metropolitan area, Eastern Cape. Sensitivity, specificity and overall accuracy of the phage test were determined compared with gold standard culture and DST. Discrepant results were resolved by molecular detection of mutations conferring rifampicin resistance. The proportion of rifampicin-resistant strains that were MDR was also determined. Previously treated patients were at a high risk of MDRTB (35.7%). Sensitivity, specificity and overall accuracy of FASTPlaque-Response for rifampicin resistance determination were 95.4% (95% confidence interval (CI): 91.0 - 99.8%), 97.2% (95% CI: 94.5 - 99.9%) and 96.5% (95% CI: 94.1 - 98.9%) respectively compared with conventional DST (unresolved), calculated for specimens that had both FASTPlaque-Response and conventional DST results available. FASTPlaque-Response results were available in 2 days instead of 28 - 85 days with conventional DST. However, only 70.6% of FASTPlaque-Response results were interpretable compared with 86.3% of conventional DST results. The majority (95.5%) of rifampicinresistant strains were MDR-TB. Rapid detection of rifampicin resistance using FASTPlaque-Response could contribute to improved management of patients at risk of MDR-TB, such as previously treated patients. However, improvement in control of specimen-related contamination is needed to ensure that a higher proportion of FASTPlaque-Response results are interpretable. Where indicated, early modification of therapy could improve patient prognosis and reduce disease transmission.