Association of telomerase activity with radio- and chemosensitivity of neuroblastomas

<p>Abstract</p> <p>Background</p> <p>Telomerase activity compensates shortening of telomeres during cell division and enables cancer cells to escape senescent processes. It is also supposed, that telomerase is associated with radio- and chemoresistance. In the here described study we systematically investigated the influence of telomerase activity (TA) and telomere length on the outcome of radio- and chemotherapy in neuroblastoma.</p> <p>Methods</p> <p>We studied the effects on dominant negative (DN) mutant, wild type (WT) of the telomerase catalytic unit (hTERT) using neuroblastoma cell lines. The cells were irradiated with ⁶⁰Co and treated with doxorubicin, etoposide, cisplatin and ifosfamide, respectively. Viability was determined by MTS/MTT-test and the GI₅₀was calculated. Telomere length was measured by southernblot analysis and TA by Trap-Assay.</p> <p>Results</p> <p>Compared to the hTERT expressing cells the dominant negative cells showed increased radiosensitivity with decreased telomere length. Independent of telomere length, telomerase negative cells are significantly more sensitive to irradiation. The effect of TA knock-down or overexpression on chemosensitivity were dependent on TA, the anticancer drug, and the chemosensitivity of the maternal cell line.</p> <p>Conclusions</p> <p>Our results supported the concept of telomerase inhibition as an antiproliferative treatment approach in neuroblastomas. Telomerase inhibition increases the outcome of radiotherapy while in combination with chemotherapy the outcome depends on drug- and cell line and can be additive/synergistic or antagonistic. High telomerase activity is one distinct cancer stem cell feature and the here described cellular constructs in combination with stem cell markers like CD133, Aldehyddehydrogenase-1 (ALDH-1) or Side population (SP) may help to investigate the impact of telomerase activity on cancer stem cell survival under therapy.</p

Implementation of the asparaginase activity assessment technique for clinical use : experience of a Brazilian Center

Acute lymphoid leukemia is a childhood cancer that in high‑income countries has event‑free survival rates of 80% and global survival rates of 90%. In Brazil these rates are under 70%. This difference may be due to the implementation of supportive care, including the assessment of asparaginase (ASNase) activity. ASNase may cause hypersensitivity reactions and silent drug inactivation. For this reason, ASNase activity monitoring is an essential tool to ensure an effective treatment. Our aim was to implement an ASNase activity measurement technique at a hospital setting. samples from children who were given Escherichia coli‑derived ASNase were collected. The results of the analyses conducted in our laboratory Hospital de Clínicas de Porto Alegre were compared to those of two institutions: Centro Infantil Boldrini and University of Munster. 262 samples were assessed. The results of the first analyses were compared with those obtained at Centro Infantil Boldrini and showed an ICC of 0.954. Thirty samples were sent to the University of Munster and presented an ICC was 0.960. Our results, when compared to those of national and international centers, showed an excellent agreement. The study was able to implement an ASNase activity test to monitor the treatment

Pre-existing antibodies against polyethylene glycol reduce asparaginase activities on first administration of pegylated <i>E. coli</i> asparaginase in children with acute lymphocytic leukemia

Antibodies against polyethylene glycol (PEG) in healthy subjects raise concerns about the efficacy of pegylated drugs. We evaluated the prevalence of antibodies against PEG among patients with acute lymphoblastic leukemia (ALL) prior to and/or immediately after their first dose of pegylated E.coli asparaginase (PEG-ASNase). Serum samples of 701 children, 673 with primary ALL, 28 with relapsed ALL, and 188 adults with primary ALL were analyzed for anti-PEG IgG and IgM. Measurements in 58 healthy infants served as reference to define cut-points for antibody-positive and -negative samples. Anti-PEG antibodies were detected in ALL patients prior the first PEG-ASNase with a prevalence of 13.9% (anti-PEG IgG) and 29.1% (anti-PEG IgM). After administration of PEG-ASNase the prevalence of anti-PEG antibodies decreased to 4.2% for anti-PEG IgG and to 4.5% for anti-PEG IgM. Pre-existing anti-PEG antibodies did not inhibit PEG-ASNase activity but significantly reduced PEGASNase activity levels in a concentration dependent manner. Although pre-existing anti-PEG antibodies did not boost, pre-existing anti-PEG IgG were significantly associated with firstexposure hypersensitivity reactions (CTCAE grade 2) (p<0.01; Fisher’s exact test). Two of 4 patients with pre-existing anti-PEG IgG and first-exposure hypersensitivity reactions were not switched to Erwinia ASNase and continued on PEG-ASNase with sufficient activities (≥100U/L). Pre-existing anti-PEG antibodies were detected in a considerable proportion of patients with ALL, did not inhibit PEG-ASNase activity but were associated with lower serum PEGASNase activity levels. Patients with pre-existing antibodies may show mild to moderate signs of hypersensitivity reaction after their first PEG-ASNase, which may be successfully addressed by re-challenge

Asparagine levels in the cerebrospinal fluid of children with acute lymphoblastic leukemia treated with pegylated-asparaginase in the induction phase of the AIEOP-BFM ALL 2009 study

Asparagine levels in cerebrospinal fluid and serum asparaginase activity were monitored in children with acute lymphoblastic leukemia treated with pegylated-asparaginase. The drug was given intravenously at a dose of 2,500 IU/m2 on days 12 and 26. Serum and cerebrospinal fluid samples obtained on days 33 and 45 were analyzed centrally. Since physiological levels of asparagine in the cerebrospinal fluid of children and adolescents are 4-10 μmol/L, in this study asparagine depletion was considered complete when the concentration of asparagine was ≤0.2 μmol/L, i.e. below the lower limit of quantification of the assay used. Over 24 months 736 patients (AIEOP n=245, BFM n=491) and 903 cerebrospinal fluid samples (n=686 on day 33 and n=217 on day 45) were available for analysis. Data were analyzed separately for the AIEOP and BFM cohorts and yielded superimposable results. Independently of serum asparaginase activity levels, cerebrospinal fluid asparagine levels were significantly reduced during the investigated study phase but only 28% of analyzed samples showed complete asparagine depletion while relevant levels, ≥1 μmol/L, were still detectable in around 23% of them. Complete cerebrospinal fluid asparagine depletion was found in around 5-6% and 33-37% of samples at serum asparaginase activity level

On the Early Digging of Peanut Fruits

1. The flowering period of peanut is about 100 days or more, which differentiates very much the respective maturity of each fruit of peanut, so it is difficult that growers catch its exact yielding time. 2. By the factors of climate, especially temperature, the flowering or maturing period is controlled, and the yielding period is from the middle of September to the end of October. 3. The effective flowering period was until the end of August, and the seeded plant at the beginning of July had its effective flowering period of only about one month, and so the July-seeded plant had only half of the product of the optimum seeded plant at the beginning of May. The seeded plant after July had the common pods, but brought no grain. 4. Immature grains decreased rapidly after about 90 days from the first flowering of each plant, or about 40 days from the maximum flowering time. 5. The plants harvested after the middle of October, which passed over the 110 days from the first flowering of each plant, produced many over-mature grains and germinating grains. 6. The early harvesting time is better than the customary time, and when the immature pods are many, the mature fruits should be harvested and the plants with immature pods should be gathered at the corner of the field and planted temporarily. After they were laid on till the frost time, the Secondly harvest should be done. 7. The optimum harvesting time was about after one month from the end of effective flowering period, or after one month and a half from the maximum of fertile percentage

Influence of cimetidine and its metabolites on Cisplatin-Investigation of adduct formation by means of electrochemistry/liquid chromatography/electrospray mass spectrometry

<p>Cimetidine has been studied as an additive in cancer chemotherapy. It is claimed to reduce the side effects of Cisplatin. This study focuses on possible interactions between Cisplatin and cimetidine on the molecular level. Due to the fact that cimetidine is metabolized in the liver, interactions between its metabolites and Cisplatin are also investigated. By means of LC/ESI-MS, Cisplatin-cimetidine adducts were detected. In a second step, the metabolism of cimetidine was simulated by electrochemical oxidation. These results were compared with microsomal incubations of cimetidine using rat and human liver cell microsomes. Because the two methods showed a correlation, the electrochemical approach was further used to investigate Cisplatin's interactions with metabolites of cimetidine. However, notable interactions that might take place in the human body could neither be observed for pure cimetidine nor for its metabolites. Finally, the impact of cimetidine on Cisplatin-protein interactions were studied using the model protein p-lactoglobulin A. In the presence of cimetidine, the affinity of Cisplatin towards the model protein appears to be increased. (C) 2013 Elsevier B.V. All rights reserved.</p>

Bioanalysis of doxorubicin aglycone metabolites in human plasma samples–implications for doxorubicin drug monitoring

The widespread clinical use of the cytostatic doxorubicin together with the induction of chronic cardiomyopathy necessitates the conduct of further pharmacokinetic trials. Novel analytical technologies suitable for point-of-care applications can facilitate drug level analyses but might be prone to interferences from structurally similar compounds. Besides the alcohol metabolite doxorubicinol, aglycone metabolites of doxorubicin might affect its determination in plasma. To evaluate their analytical relevance, a validated HPLC method for the quantification of doxorubicin, doxorubicinol and four aglycones was used. The degradation pattern of doxorubicin in plasma under long-term storage was analysed with respect to the formation of aglycone products. In addition, overall 50 clinical samples obtained within the EPOC-MS-001-Doxo trial were analysed. Substantial degradation of doxorubicin in plasma occurred within a storage period of one year, but this did not lead to the formation of aglycones. In clinical samples, 7-deoxydoxorubicinolone was the major aglycone detectable in 35/50 samples and a concentration range of 1.0–12.7 μg L^−1. If at all, the other aglycones were only determined in very low concentrations. Therefore, analytical interferences from aglycones seem to be unlikely with the exception of 7-deoxydoxorubicinolone whose concentration accounted for up to 65% of the doxorubicin concentration in the clinical samples analysed