Accelerated partial breast irradiation using sole interstitial multicatheter brachytherapy compared with whole-breast irradiation with boost for early breast cancer: 10-year results of a GEC-ESTRO randomised, phase 3, non-inferiority trial.

BACKGROUND: Several randomised, phase 3 trials have investigated the value of different techniques of accelerated partial breast irradiation (APBI) for patients with early breast cancer after breast-conserving surgery compared with whole-breast irradiation. In a phase 3 randomised trial, we evaluated whether APBI using multicatheter brachytherapy is non-inferior compared with whole-breast irradiation. Here, we present the 10-year follow-up results. METHODS: We did a randomised, phase 3, non-inferiority trial at 16 hospitals and medical centres in Austria, Czech Republic, Germany, Hungary, Poland, Spain, and Switzerland. Patients aged 40 years or older with early invasive breast cancer or ductal carcinoma in situ treated with breast-conserving surgery were centrally randomly assigned (1:1) to receive either whole-breast irradiation or APBI using multicatheter brachytherapy. Whole-breast irradiation was delivered in 25 daily fractions of 50 Gy over 5 weeks, with a supplemental boost of 10 Gy to the tumour bed, and APBI was delivered as 30·1 Gy (seven fractions) and 32·0 Gy (eight fractions) of high-dose-rate brachytherapy in 5 days or as 50 Gy of pulsed-dose-rate brachytherapy over 5 treatment days. Neither patients nor investigators were masked to treatment allocation. The primary endpoint was ipsilateral local recurrence, analysed in the as-treated population; the non-inferiority margin for the recurrence rate difference (defined for 5-year results) was 3 percentage points. The trial is registered with ClinicalTrials.gov, NCT00402519; the trial is complete. FINDINGS: Between April 20, 2004, and July 30, 2009, 1328 female patients were randomly assigned to whole breast irradiation (n=673) or APBI (n=655), of whom 551 in the whole-breast irradiation group and 633 in the APBI group were eligible for analysis. At a median follow-up of 10·36 years (IQR 9·12-11·28), the 10-year local recurrence rates were 1·58% (95% CI 0·37 to 2·8) in the whole-breast irradiation group and 3·51% (1·99 to 5·03) in the APBI group. The difference in 10-year rates between the groups was 1·93% (95% CI -0·018 to 3·87; p=0·074). Adverse events were mostly grade 1 and 2, in 234 (60%) of 393 participants in the whole-breast irradiation group and 314 (67%) of 470 participants in the APBI group, at 7·5-year or 10-year follow-up, or both. Patients in the APBI group had a significantly lower incidence of treatment-related grade 3 late side-effects than those in the whole-breast irradiation group (17 [4%] of 393 for whole-breast irradiation vs seven [1%] of 470 for APBI; p=0·021; at 7·5-year or 10-year follow-up, or both). At 10 years, the most common type of grade 3 adverse event in both treatment groups was fibrosis (six [2%] of 313 patients for whole-breast irradiation and three [1%] of 375 patients for APBI, p=0·56). No grade 4 adverse events or treatment-related deaths have been observed. INTERPRETATION: Postoperative APBI using multicatheter brachytherapy after breast-conserving surgery in patients with early breast cancer is a valuable alternative to whole-breast irradiation in terms of treatment efficacy and is associated with fewer late side-effects. FUNDING: German Cancer Aid, Germany.

Image-guided high-dose-rate brachytherapy for prostate cancer patients with previous rectal resection and pelvic irradiation: feasibility study.

PURPOSE: The aim of this study was to describe treatment procedure and early clinical outcomes of high-dose-rate (HDR) interstitial brachytherapy (BT) in clinically localized prostate cancer patients previously treated for rectal cancer with abdominoperineal rectal resection and external beam radiation therapy (EBRT). MATERIAL AND METHODS: Between February and July 2015, two patients with clinically localized prostate cancer without rectal access were treated in our brachytherapy department. HDR interstitial brachytherapy was conducted with the guidance of fluoroscopy and computed tomography (CT) imaging. Brachytherapy was combined with hormonal therapy. RESULTS: Follow-up lasted for 34 and 39 months for patient 1 and 2, respectively. Both patients remained free from biochemical recurrence according to the Phoenix definition. No severe G3/G4 late toxicity was observed, and neither patient experienced any gastrointestinal morbidity. Acute and late urinary toxicities were at an acceptance level, and were scored G1 and G2 for patient 1, and G3 and G1 for patient 2, respectively. CONCLUSIONS: Fluoroscopy and 3D CT image-guided interstitial brachytherapy is feasible and appears to be a suitable treatment technique for patients with clinically localized prostate cancer after previous rectal resection and external beam radiation therapy.

Heterogeneously Catalysed Oxidative Dehydrogenation of Menthol in a Fixed-Bed Reactor in the Gas Phase.

For the first time, the oxidative dehydrogenation of (-)-menthol to (-)-menthone and (+)-isomenthone in a marketable quality was carried out in a continuous gas phase reactor as a sustainable process using molecular oxygen as green oxidant and solid catalysts which do not contaminate the product mixture and which are easily to remove. The diastereomeric purity remained largely unchanged. Three types of catalysts were found to be very active and selective in the formation of menthone and isomenthone: AgSr/SiO2, CuO distributed on a basic support and RuMnCe/CeO2, where Ru, Mn and Ce exist in an oxidized state. The best overall yield of menthon/isomenthone obtained with an Ag-based catalyst was 58 % at 64 % selectivity, with a Cu-based catalyst 41 % at 51 % selectivity and with a Ru-based catalyst 68 % at 73 % selectivity. Reaction conditions were widely optimized.

Quality-of-life results for accelerated partial breast irradiation with interstitial brachytherapy versus whole-breast irradiation in early breast cancer after breast-conserving surgery (GEC-ESTRO): 5-year results of a randomised, phase 3 trial.

BACKGROUND: Previous results from the GEC-ESTRO trial showed that accelerated partial breast irradiation (APBI) using multicatheter brachytherapy in the treatment of early breast cancer after breast-conserving surgery was non-inferior to whole-breast irradiation in terms of local control and overall survival. Here, we present 5-year results of patient-reported quality of life. METHODS: We did this randomised controlled phase 3 trial at 16 hospitals and medical centres in seven European countries. Patients aged 40 years or older with 0-IIA breast cancer were randomly assigned (1:1) after breast-conserving surgery (resection margins ≥2 mm) to receive either whole-breast irradiation of 50 Gy with a boost of 10 Gy or APBI using multicatheter brachytherapy. Randomisation was stratified by study centre, tumour type, and menopausal status, with a block size of ten and an automated dynamic algorithm. There was no masking of patients or investigators. The primary endpoint of the trial was ipsilateral local recurrence. Here, we present 5-year results of quality of life (a prespecified secondary endpoint). Quality-of-life questionnaires (European Organisation for Research and Treatment of Cancer QLQ-C30, breast cancer module QLQ-BR23) were completed before radiotherapy (baseline 1), immediately after radiotherapy (baseline 2), and during follow-up. We analysed the data according to treatment received (as-treated population). Recruitment was completed in 2009, and long-term follow-up is continuing. The trial is registered at ClinicalTrials.gov, number NCT00402519. FINDINGS: Between April 20, 2004, and July 30, 2009, 633 patients had accelerated partial breast irradiation and 551 patients had whole-breast irradiation. Quality-of-life questionnaires at baseline 1 were available for 334 (53%) of 663 patients in the APBI group and 314 (57%) of 551 patients in the whole-breast irradiation group; the response rate was similar during follow-up. Global health status (range 0-100) was stable in both groups: at baseline 1, APBI group mean score 65·5 (SD 20·6) versus whole-breast irradiation group 64·6 (19·6), p=0·37; at 5 years, APBI group 66·2 (22·2) versus whole-breast irradiation group 66·0 (21·8), p=0·94. The only moderate, significant difference (difference of 10-20 points) between the groups was found in the breast symptoms scale. Breast symptom scores were significantly higher (ie, worse) after whole-breast irradiation than after APBI at baseline 2 (difference of means 13·6, 95% CI 9·7-17·5; p<0·0001) and at 3-month follow-up (difference of means 12·7, 95% CI 9·8-15·6; p<0·0001). INTERPRETATION: APBI with multicatheter brachytherapy was not associated with worse quality of life compared with whole-breast irradiation. This finding supports APBI as an alternative treatment option after breast-conserving surgery for patients with early breast cancer. FUNDING: German Cancer Aid.

Late side-effects and cosmetic results of accelerated partial breast irradiation with interstitial brachytherapy versus whole-breast irradiation after breast-conserving surgery for low-risk invasive and in-situ carcinoma of the female breast: 5-year results of a randomised, controlled, phase 3 trial.

BACKGROUND: We previously confirmed the non-inferiority of accelerated partial breast irradiation (APBI) with interstitial brachytherapy in terms of local control and overall survival compared with whole-breast irradiation for patients with early-stage breast cancer who underwent breast-conserving surgery in a phase 3 randomised trial. Here, we present the 5-year late side-effects and cosmetic results of the trial. METHODS: We did this randomised, controlled, phase 3 trial at 16 centres in seven European countries. Women aged 40 years or older with stage 0-IIA breast cancer who underwent breast-conserving surgery with microscopically clear resection margins of at least 2 mm were randomly assigned 1:1, via an online interface, to receive either whole-breast irradiation of 50 Gy with a tumour-bed boost of 10 Gy or APBI with interstitial brachytherapy. Randomisation was stratified by study centre, menopausal status, and tumour type (invasive carcinoma vs ductal carcinoma in situ), with a block size of ten, according to an automated dynamic algorithm. Patients and investigators were not masked to treatment allocation. The primary endpoint of our initial analysis was ipsilateral local recurrence; here, we report the secondary endpoints of late side-effects and cosmesis. We analysed physician-scored late toxicities and patient-scored and physician-scored cosmetic results from the date of breast-conserving surgery to the date of onset of event. Analysis was done according to treatment received (as-treated population). This trial is registered with ClinicalTrials.gov, number NCT00402519. FINDINGS: Between April 20, 2004, and July 30, 2009, we randomly assigned 1328 women to receive either whole-breast irradiation (n=673) or APBI with interstitial brachytherapy (n=655); 1184 patients comprised the as-treated population (551 in the whole-breast irradiation group and 633 in the APBI group). At a median follow-up of 6·6 years (IQR 5·8-7·6), no patients had any grade 4 toxities, and three (<1%) of 484 patients in the APBI group and seven (2%) of 393 in the whole-breast irradiation group had grade 3 late skin toxicity (p=0·16). No patients in the APBI group and two (<1%) in the whole-breast irradiation group developed grade 3 late subcutaneous tissue toxicity (p=0·10). The cumulative incidence of any late side-effect of grade 2 or worse at 5 years was 27·0% (95% CI 23·0-30·9) in the whole-breast irradiation group versus 23·3% (19·9-26·8) in the APBI group (p=0·12). The cumulative incidence of grade 2-3 late skin toxicity at 5 years was 10·7% (95% CI 8·0-13·4) in the whole-breast irradiation group versus 6·9% (4·8-9·0) in the APBI group (difference -3·8%, 95% CI -7·2 to 0·4; p=0·020). The cumulative risk of grade 2-3 late subcutaneous tissue side-effects at 5 years was 9·7% (95% CI 7·1-12·3) in the whole-breast irradiation group versus 12·0% (9·4-14·7) in the APBI group (difference 2·4%; 95% CI -1·4 to 6·1; p=0·28). The cumulative incidence of grade 2-3 breast pain was 11·9% (95% CI 9·0-14·7) after whole-breast irradiation versus 8·4% (6·1-10·6) after APBI (difference -3·5%; 95% CI -7·1 to 0·1; p=0·074). At 5 years' follow-up, according to the patients' view, 413 (91%) of 454 patients had excellent to good cosmetic results in the whole-breast irradiation group versus 498 (92%) of 541 patients in the APBI group (p=0·62); when judged by the physicians, 408 (90%) of 454 patients and 503 (93%) of 542 patients, respectively, had excellent to good cosmetic results (p=0·12). No treatment-related deaths occurred, but six (15%) of 41 patients (three in each group) died from breast cancer, and 35 (85%) deaths (21 in the whole-breast irradiation group and 14 in the APBI group) were unrelated. INTERPRETATION: 5-year toxicity profiles and cosmetic results were similar in patients treated with breast-conserving surgery followed by either APBI with interstitial brachytherapy or conventional whole-breast irradiation, with significantly fewer grade 2-3 late skin side-effects after APBI with interstitial brachytherapy. These findings provide further clinical evidence for the routine use of interstitial multicatheter brachytherapy-based APBI in the treatment of patients with low-risk breast cancer who opt for breast conservation. FUNDING: German Cancer Aid.

GEC-ESTRO multicenter phase 3-trial: Accelerated partial breast irradiation with interstitial multicatheter brachytherapy versus external beam whole breast irradiation: Early toxicity and patient compliance.

BACKGROUND AND PURPOSE: To compare early side effects and patient compliance of accelerated partial breast irradiation (APBI) with multicatheter brachytherapy to external beam whole breast irradiation (WBI) in a low-risk group of patients with breast cancer. MATERIAL AND METHODS: Between April 2004 and July 2009, 1328 patients with UICC stage 0-IIA breast cancer were randomized to receive WBI with 50Gy and a boost of 10Gy or APBI with either 32.0Gy/8 fractions, or 30.1Gy/7 fractions (HDR-brachytherapy), or 50Gy/0.60-0.80Gy per pulse (PDR-brachytherapy). This report focuses on early side-effects and patient compliance observed in 1186 analyzable patients. ClinicalTrials.gov identifier: NCT00402519. RESULTS: Patient compliance was excellent in both arms. Both WBI and APBI were well tolerated with moderate early side-effects. No grade 4 toxicity had been observed. Grade 3 side effects were exclusively seen for early skin toxicity (radiation dermatitis) with 7% vs. 0.2% (p<0.0001), and breast infection with 0% vs. 0.2% (p=n.s.) for patients treated with WBI and APBI. The incidence of grades 1-2 early side effects for WBI and APBI was 86% vs. 21% (p<0.0001) for skin toxicity, 2% vs. 20% (p<0.0001) for mild hematoma, and 2% vs. 5% (p=0.01) for mild breast infection rates, respectively. No differences had been found regarding grades 1-2 early breast pain (26% vs. 29%, p=0.23). CONCLUSIONS: APBI with interstitial multicatheter brachytherapy was tolerated very well and dramatically reduced early skin toxicity in comparison to standard WBI.

5-year results of accelerated partial breast irradiation using sole interstitial multicatheter brachytherapy versus whole-breast irradiation with boost after breast-conserving surgery for low-risk invasive and in-situ carcinoma of the female breast: a randomised, phase 3, non-inferiority trial.

BACKGROUND: In a phase 3, randomised, non-inferiority trial, accelerated partial breast irradiation (APBI) for patients with stage 0, I, and IIA breast cancer who underwent breast-conserving treatment was compared with whole-breast irradiation. Here, we present 5-year follow-up results. METHODS: We did a phase 3, randomised, non-inferiority trial at 16 hospitals and medical centres in seven European countries. 1184 patients with low-risk invasive and ductal carcinoma in situ treated with breast-conserving surgery were centrally randomised to either whole-breast irradiation or APBI using multicatheter brachytherapy. The primary endpoint was local recurrence. Analysis was done according to treatment received. This trial is registered with ClinicalTrials.gov, number NCT00402519. FINDINGS: Between April 20, 2004, and July 30, 2009, 551 patients had whole-breast irradiation with tumour-bed boost and 633 patients received APBI using interstitial multicatheter brachytherapy. At 5-year follow-up, nine patients treated with APBI and five patients receiving whole-breast irradiation had a local recurrence; the cumulative incidence of local recurrence was 1.44% (95% CI 0.51-2.38) with APBI and 0.92% (0.12-1.73) with whole-breast irradiation (difference 0.52%, 95% CI -0.72 to 1.75; p=0.42). No grade 4 late side-effects were reported. The 5-year risk of grade 2-3 late side-effects to the skin was 3.2% with APBI versus 5.7% with whole-breast irradiation (p=0.08), and 5-year risk of grade 2-3 subcutaneous tissue late side-effects was 7.6% versus 6.3% (p=0.53). The risk of severe (grade 3) fibrosis at 5 years was 0.2% with whole-breast irradiation and 0% with APBI (p=0.46). INTERPRETATION: The difference between treatments was below the relevance margin of 3 percentage points. Therefore, adjuvant APBI using multicatheter brachytherapy after breast-conserving surgery in patients with early breast cancer is not inferior to adjuvant whole-breast irradiation with respect to 5-year local control, disease-free survival, and overall survival. FUNDING: German Cancer Aid.

Arabidopsis ABA-Activated Kinase MAPKKK18 is Regulated by Protein Phosphatase 2C ABI1 and the Ubiquitin-Proteasome Pathway.

Phosphorylation and dephosphorylation events play an important role in the transmission of the ABA signal. Although SnRK2 [sucrose non-fermenting1-related kinase2] protein kinases and group A protein phosphatase type 2C (PP2C)-type phosphatases constitute the core ABA pathway, mitogen-activated protein kinase (MAPK) pathways are also involved in plant response to ABA. However, little is known about the interplay between MAPKs and PP2Cs or SnRK2 in the regulation of ABA pathways. In this study, an effort was made to elucidate the role of MAP kinase kinase kinase18 (MKKK18) in relation to ABA signaling and response. The MKKK18 knockout lines showed more vigorous root growth, decreased abaxial stomatal index and increased stomatal aperture under normal growth conditions, compared with the control wild-type Columbia line. In addition to transcriptional regulation of the MKKK18 promoter by ABA, we demonstrated using in vitro and in vivo kinase assays that the kinase activity of MKKK18 was regulated by ABA. Analysis of the cellular localization of MKKK18 showed that the active kinase was targeted specifically to the nucleus. Notably, we identified abscisic acid insensitive 1 (ABI1) PP2C as a MKKK18-interacting protein, and demonstrated that ABI1 inhibited its activity. Using a cell-free degradation assay, we also established that MKKK18 was unstable and was degraded by the proteasome pathway. The rate of MKKK18 degradation was delayed in the ABI1 knockout line. Overall, we provide evidence that ABI1 regulates the activity and promotes proteasomal degradation of MKKK18.

Brachytherapy for vaginal intraepithelial neoplasia.

OBJECTIVE: To retrospectively evaluate the efficacy of high-dose-rate brachytherapy of vaginal intraepithelial neoplasia with a special focus on analysis of toxicity. STUDY DESIGN: Twenty consecutive patients were irradiated with brachytherapy of vaginal intraepithelial neoplasia with component ca in situ (N=3). Late complications of the vagina graded using the CTCAE v.3.0. General assessment three-step scale was introduced for simplicity of analysis. RESULTS: The median age was 57 years (range: 28-80 years). The median follow-up time was 39 months (range: 14-115 months). Vaginal intraepithelial neoplasia recurrence was observed in 1 patient. The 3-year disease free survival rate was 90% (95% confidence interval [CI]: 71-100%). Observed late side effects: libido grades 1-2 in 15 (75%), vaginal discharge grade 2 (pad use indicated) in 2 (10%), dryness grade 2 (dyspareunia) in 7 (35%), mucositis grades 2-3 in 6 (30%), stenosis grades 2-3 in 7 (35%) and vaginitis grades 2-3 in 4 (20%) cases. General assessment was good in 9 (45%), average in 2 (10%), and bad in 9 (45%) patients. Treatment dose affected the toxicity (p=0.05). In groups of patients irradiated with biologically equivalent dose (assuming α/ß=3Gy) of 47.3-63Gy and ≥70Gy, the risk of poor or moderate toxicity amounted to 16.7% (95% CI: 0-47%) and 71.4% (95% CI: 48-95%), respectively. CONCLUSION: Brachytherapy revealed to be effective method of vaginal intraepithelial neoplasia treatment, but applying EQD2≥70Gy into vagina generates unacceptable toxicity.

Interplays between nitric oxide and reactive oxygen species in cryptogein signalling.

Nitric oxide (NO) has many functions in plants. Here, we investigated its interplays with reactive oxygen species (ROS) in the defence responses triggered by the elicitin cryptogein. The production of NO induced by cryptogein in tobacco cells was partly regulated through a ROS-dependent pathway involving the NADPH oxidase NtRBOHD. In turn, NO down-regulated the level of H2O2. Both NO and ROS synthesis appeared to be under the control of type-2 histone deacetylases acting as negative regulators of cell death. Occurrence of an interplay between NO and ROS was further supported by the finding that cryptogein triggered a production of peroxynitrite (ONOO(-)). Next, we showed that ROS, but not NO, negatively regulate the intensity of activity of the cryptogein-induced protein kinase NtOSAK. Furthermore, using a DNA microarray approach, we identified 15 genes early induced by cryptogein via NO. A part of these genes was also modulated by ROS and encoded proteins showing sequence identity to ubiquitin ligases. Their expression appeared to be negatively regulated by ONOO(-), suggesting that ONOO(-) mitigates the effects of NO and ROS. Finally, we provided evidence that NO required NtRBOHD activity for inducing cell death, thus confirming previous assumption that ROS channel NO through cell death pathways.

ß-Aminobutyric acid (BABA)-induced resistance in Arabidopsis thaliana: link with iron homeostasis.

ß-Aminobutyric acid (BABA) is a nonprotein amino acid inducing resistance in many different plant species against a wide range of abiotic and biotic stresses. Nevertheless, how BABA primes plant natural defense reactions remains poorly understood. Based on its structure, we hypothesized and confirmed that BABA is able to chelate iron (Fe) in vitro. In vivo, we showed that it led to a transient Fe deficiency response in Arabidopsis thaliana plants exemplified by a reduction of ferritin accumulation and disturbances in the expression of genes related to Fe homeostasis. This response was not correlated to changes in Fe concentrations, suggesting that BABA affects the availability or the distribution of Fe rather than its assimilation. The phenotype of BABA-treated plants was similar to those of plants cultivated in Fe-deficient conditions. A metabolomic analysis indicated that both BABA and Fe deficiency induced the accumulation of common metabolites, including p-coumaroylagmatine, a metabolite previously shown to be synthesized in several plant species facing pathogen attack. Finally, we showed that the protective effect induced by BABA against Botrytis cinerea was mimicked by Fe deficiency. In conclusion, the Fe deficiency response caused by BABA could bring the plant to a defense-ready state, participating in the plant resistance against the pathogens.

Abnormal urinary excretion of NKCC2 and AQP2 in response to hypertonic saline in chronic kidney disease: an intervention study in patients with chronic kidney disease and healthy controls.

BACKGROUND: Renal handling of sodium and water is abnormal in chronic kidney disease (CKD). The aim of this study was to test the hypothesis that abnormal activity of the aquaporin-2 water channels (AQP2), the sodium-potassium-2chloride transporter (NKCC2) and/or the epithelial sodium channels (ENaC) contribute to this phenomenon. METHODS: 23 patients with CKD and 24 healthy controls at baseline and after 3% saline infusion were compared. The following measurements were performed: urinary concentrations of AQP2 (u-AQP2), NKCC2 (u-NKCC2), ENaC (u-ENaCγ), glomerular filtration rate (GFR) estimated by 51Cr-EDTA clearance, free water clearance (CH2O), urinary output (UO), fractional excretion of sodium (FENa), plasma concentrations of AVP, renin (PRC), Angiotensin II (ANG II), Aldosterone (Aldo) and body fluid volumes. RESULTS: At baseline, GFR was 34 ml/min in CKD patients and 89 ml/ml in controls. There were no significant differences in u-AQP2, u-NKCC2 or u-ENaCγ, but FENa, p-Aldo and p-AVP were higher in CKD patients than controls. In response to hypertonic saline, patients with CKD had an attenuated decrease in CH2O and UO. A greater increase in U-AQP2 was observed in CKD patients compared to controls. Furthermore, u-NKCC2 increased in CKD patients, whereas u-NKCC2 decreased in controls. Body fluid volumes did not significantly differ. CONCLUSIONS: In response to hypertonic saline, u-NKCC2 increased, suggesting an increased sodium reabsorption via NKCC2 in patients with CKD. U-AQP2 increased more in CKD patients, despite an attenuated decrease in CH2O. Thus, though high levels of p-AVP and p-Aldo, the kidneys can only partly compensate and counteract acute volume expansion due to a defective tubular response. TRIAL REGISTRATION: Clinical trial no: NCT01623661. Date of trial registration: 18.06.2012.

SNF1-related protein kinases type 2 are involved in plant responses to cadmium stress.

Cadmium ions are notorious environmental pollutants. To adapt to cadmium-induced deleterious effects plants have developed sophisticated defense mechanisms. However, the signaling pathways underlying the plant response to cadmium are still elusive. Our data demonstrate that SnRK2s (for SNF1-related protein kinase2) are transiently activated during cadmium exposure and are involved in the regulation of plant response to this stress. Analysis of tobacco (Nicotiana tabacum) Osmotic Stress-Activated Protein Kinase activity in tobacco Bright Yellow 2 cells indicates that reactive oxygen species (ROS) and nitric oxide, produced mainly via an l-arginine-dependent process, contribute to the kinase activation in response to cadmium. SnRK2.4 is the closest homolog of tobacco Osmotic Stress-Activated Protein Kinase in Arabidopsis (Arabidopsis thaliana). Comparative analysis of seedling growth of snrk2.4 knockout mutants versus wild-type Arabidopsis suggests that SnRK2.4 is involved in the inhibition of root growth triggered by cadmium; the mutants were more tolerant to the stress. Measurements of the level of three major species of phytochelatins (PCs) in roots of plants exposed to Cd(2+) showed a similar (PC2, PC4) or lower (PC3) concentration in snrk2.4 mutants in comparison to wild-type plants. These results indicate that the enhanced tolerance of the mutants does not result from a difference in the PCs level. Additionally, we have analyzed ROS accumulation in roots subjected to Cd(2+) treatment. Our data show significantly lower Cd(2+)-induced ROS accumulation in the mutants' roots. Concluding, the obtained results indicate that SnRK2s play a role in the regulation of plant tolerance to cadmium, most probably by controlling ROS accumulation triggered by cadmium ions.

[Improved prognosis of cystinosis achieved by treatment with cysteamine and by kidney transplantation]. / Bedre prognose af cystinose ved behandling med cysteamin og nyretransplantation.

Cystinosis is a rare, autosomal recessive disease with cystine deposits in different tissues. First signs come from kidneys and eyes, but during progression of the disease other organs can also be affected. Previously, patients with cystinosis had a very poor prognosis, but it is now considerably improved due to new methods of treatment. The purpose of this paper is to give a brief review of the disease and discuss the significant improvement of the prognosis, which has been achieved by specific medical treatment with cystine-depleting agents, and, if needed, by kidney transplantation.

Determination of active substances in binary mixture antiparasitic veterinary formulations by HPLC.

The purpose of the study was to develop a simple, versatile HPLC method for the identification and quantification of praziquantel and ivermectin (in Equimax) or praziquantel and abamectin (in Abamitel Plus). A satisfactory separation was obtained using the Supelcosil LC-ABZ+ column in gradient system with a mobile phase A: acetonitrile / water in 40:60 ratio and phase B: acetonitrile. The UV detection was set at 245 nm. The correlation coefficient values (> or = 0,998) for all active substances confirmed that the calibration curves (peak area vs. concentration) are linear. The results of the quantification and the statistical evaluation confirmed that the method is accurate and precise. It can also be applied to confirm the identity of benzyl alcohol, methyl p-hydroxybenzoate and propyl p-hydroxybenzoate in Abamitel Plus formulation.

SNF1-related protein kinases 2 are negatively regulated by a plant-specific calcium sensor.

SNF1-related protein kinases 2 (SnRK2s) are plant-specific enzymes involved in environmental stress signaling and abscisic acid-regulated plant development. Here, we report that SnRK2s interact with and are regulated by a plant-specific calcium-binding protein. We screened a Nicotiana plumbaginifolia Matchmaker cDNA library for proteins interacting with Nicotiana tabacum osmotic stress-activated protein kinase (NtOSAK), a member of the SnRK2 family. A putative EF-hand calcium-binding protein was identified as a molecular partner of NtOSAK. To determine whether the identified protein interacts only with NtOSAK or with other SnRK2s as well, we studied the interaction of an Arabidopsis thaliana orthologue of the calcium-binding protein with selected Arabidopsis SnRK2s using a two-hybrid system. All kinases studied interacted with the protein. The interactions were confirmed by bimolecular fluorescence complementation assay, indicating that the binding occurs in planta, exclusively in the cytoplasm. Calcium binding properties of the protein were analyzed by fluorescence spectroscopy using Tb(3+) as a spectroscopic probe. The calcium binding constant, determined by the protein fluorescence titration, was 2.5 ± 0.9 × 10(5) M(-1). The CD spectrum indicated that the secondary structure of the protein changes significantly in the presence of calcium, suggesting its possible function as a calcium sensor in plant cells. In vitro studies revealed that the activity of SnRK2 kinases analyzed is inhibited in a calcium-dependent manner by the identified calcium sensor, which we named SCS (SnRK2-interacting calcium sensor). Our results suggest that SCS is involved in response to abscisic acid during seed germination most probably by negative regulation of SnRK2s activity.

Regulation of Nicotiana tabacum osmotic stress-activated protein kinase and its cellular partner GAPDH by nitric oxide in response to salinity.

Several studies focusing on elucidating the mechanism of NO (nitric oxide) signalling in plant cells have highlighted that its biological effects are partly mediated by protein kinases. The identity of these kinases and details of how NO modulates their activities, however, remain poorly investigated. In the present study, we have attempted to clarify the mechanisms underlying NO action in the regulation of NtOSAK (Nicotiana tabacum osmotic stress-activated protein kinase), a member of the SNF1 (sucrose non-fermenting 1)-related protein kinase 2 family. We found that in tobacco BY-2 (bright-yellow 2) cells exposed to salt stress, NtOSAK is rapidly activated, partly through a NO-dependent process. This activation, as well as the one observed following treatment of BY-2 cells with the NO donor DEA/NO (diethylamine-NONOate), involved the phosphorylation of two residues located in the kinase activation loop, one being identified as Ser158. Our results indicate that NtOSAK does not undergo the direct chemical modifications of its cysteine residues by S-nitrosylation. Using a co-immunoprecipitation-based strategy, we identified several proteins present in immunocomplex with NtOSAK in salt-treated cells including the glycolytic enzyme GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Our results indicate that NtOSAK directly interacts with GAPDH in planta. Furthermore, in response to salt, GAPDH showed a transient increase in its S-nitrosylation level which was correlated with the time course of NtOSAK activation. However, GADPH S-nitrosylation did not influence its interaction with NtOSAK and did not have an impact on the activity of the protein kinase. Taken together, the results support the hypothesis that NtOSAK and GAPDH form a cellular complex and that both proteins are regulated directly or indirectly by NO.

Surgery combined with brachytherapy in patients with retroperitoneal sarcomas.

PURPOSE: The primary aim of this work was to analyze feasibility of combined treatment of retroperitoneal sarcomas (RS): surgery (S) and intraoperative brachytherapy (IOBRT). The secondary aim was to analyze results and complications after this treatment. MATERIAL AND METHODS: 84 patients with retroperitoneal sarcomas were qualified for combined treatment (S and IOBRT) between June 1998 and September 2006. 65 of the patients (77.4%) had local recurrences. Sarcomas with intermediate and high grade of histological malignancy (G2, G3 - 76.2%) were the most frequent within the all surgically treated patients. Resection ability (R0/R1) in analyzed group of patients was estimated as 85% (74 cases). After intraoperative evaluation, 57 (67.8%) patients were qualified for IOBRT. Since 2000, in 34 patients (60%) an adjuvant postoperative external beam radiation therapy (EBRT) in dose of 50 Gy was applied. Median follow-up of the surviving patients was 40 months. RESULTS: On the basis of the univariate analysis, relevant aspects negatively influencing overall survival rate within the RS group treated with IOBRT were as follows: surgery of sarcoma recurrence (p = 0.002), higher grade of histological malignancy (p = 0.05), histological type different than liposarcoma (p = 0.05) as well as no adjuvant EBRT (p = 0.05). On the basis of multivariate analysis one can ascertain that relevant factors negatively influencing LRFS in RS patients treated with IOBRT were: surgery due to recurrence of sarcoma (p = 0.008) and lack of EBRT (p = 0.01). CONCLUSIONS: Combined treatment (surgery and brachytherapy) was possible to be carried out on 68% of RS patients. The overall number of complications was quite high, however acceptable, taking into consideration the application of extensive, multi-organ treatments in case of sarcoma recurrences in this localization. The results suggest that the method of treatment will improve the final outcome when most of patients will be qualified for treatment of primary sarcomas in experienced centre.

Cosmetic effect in patients with early breast cancer treated with breast conserving therapy (BCT) and with HDR brachytherapy (HDR-BT) "boost".

PURPOSE: The estimation of cosmetic effect in 93 patients with early breast cancer treated with breast conserving surgery (BCS) followed by combined radiotherapy, including HDR brachytherapy (HDR-BT) boost. MATERIAL AND METHODS: After BCS (tumorectomy or quadrantectomy) external beam radiation therapy (EBRT) was used in total dose of 50 Gy for the whole breast. Tumor bed was localized basing on clinical and mammographic preoperative examinations and histopathology evaluation. 10 Gy in one fraction was applied to all patients using HDR-BT. Steel interstitial needles stabilized by plastic templates were used. 192-Ir with 10 Ci nominal activity and HDR-GammaMed 12i unit (Mick Radio-Nuclear Instruments, Inc., Mt. Vernon, NY) and ABACUS software were used. 31 patients received additional chemotherapy. Cosmetic effect was evaluated in 36 month after the end of brachytherapy treatment basing on modified EORTC scale. For statistical analysis the rang of correlation test, contingent test, linear regression test and ProbRough rulet induction test were used. RESULTS AND CONCLUSIONS: HDR-BT tolerance was good in most of the cases. Excellent and very good cosmetic effect was observed in 79 patients (85%). Statistically important correlations between following examined prognostic factors and cosmetics outcome were observed: clinical and mammographic tumor estimation, method of breast conserving surgery, type of skin incision, number of interstitial applicators, irradiated reference volume (PTV) and type of optimization method. No correlations with cosmetics effect were found in factors such as: age of patients, location of tumor or additional therapy.

HDR brachytherapy as a solution in recurrences of locally advanced prostate cancer.

PURPOSE: The object of this study was to analyze the outcome of salvage HDR brachytherapy treatment after local failure, for patients with prostate specific antigen (PSA) failure without distant metastasis, after external beam radiation and HDR brachytherapy treatment, or after radical prostatectomy, with or without hormonal therapy. MATERIAL AND METHODS: The group of 115 patients, without distant metastasis, after local failure and external beam radiation, followed by HDR brachytherapy treatment, or after radical prostatectomy, with hormonal therapy and without, have been enrolled to salvage HDR brachytherapy (SBR). All patients had minimum 3 months androgen deprivation therapy before salvage brachytherapy, which was continued until the next 9 months after SBR. Brachytherapy was administered in three 10 Gy fractions with 3 weeks gap between them. Each session of SBR was supported by trans-rectal USG real time pictures. The treatment planning was done on the base of Abacus system from Sauerwein® or with SWIFT system from Nucletron®. The following data were collected: Gleason score, clinical staging, the volume of the prostate, PSA before and after the initial treatment and periodically during the follow-up period. Also the time during which the PSA stays at the nadir level, patient's age and toxicity of treatments were taken into consideration. RESULTS: Doses from external radiotherapy or from HDR brachytherapy were recalculated to equivalent biological dose (EBD). The independence from biochemical progression in our group of patients after retreatment was 46% for patients with PSA ≤ 6 and 18% for patients with PSA > 6. Overall survival for patients with PSA ≤ 6 was 86% and 48% for patients with PSA > 6, respectively. CONCLUSIONS: Salvage prostate brachytherapy (SBR) can be safely performed with acceptable biochemical control and toxicity.