Development of a rapid detection assay for acetolactate synthase inhibitors resistance in three Amaranthus weed species through loop-mediated isothermal amplification.

BACKGROUND: The early detection of herbicide resistance in weeds is a key factor to avoid herbicide waste and improve agriculture sustainability. The present study aimed to develop and validate an allele-specific loop-mediated isothermal amplification (AS-LAMP) assay for the quick on-site detection of the resistance-endowing point mutation Trp-574-Leu in the acetolactate synthase (ALS) gene in three widely diffused Amaranthus weed species: Amaranthus retroflexus, Amaranthus hybridus and Amaranthus tuberculatus. RESULTS: The AS-LAMP protocol was developed on wild-type and ALS-mutant plants of the three species and revealed that the amplification approach with only the primer set specific for the mutant allele (574-Leu) was the most promising. The validation and estimation of the AS-LAMP performance evaluated by comparing the results with those of the molecular marker (cleaved amplified polymorphic sequences) indicated that, although the sensitivity and specificity were relatively high in all species (overall 100 and > 65%, respectively), precision was high for A. hybridus L. and A. retroflexus L. (75 and 79%, respectively), but quite low for A. tuberculatus (Moq.) J. D. Sauer (59%). The LAMP assay was also effective on crude genomic DNA extraction, allowing the quick detection of mutant plants in field situation (on site resistance detection). CONCLUSION: The proposed AS-LAMP method has proven to be a promising technique for rapid detection of resistance as a result of Trp-574-Leu on the two monoecious weedy Amaranthus species but resulted less effective in the genetically variable dioecious species A. tuberculatus. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Trastuzumab-related cardiotoxicity in patients with nonlimiting cardiac comorbidity.

BACKGROUND: Significant and symptomatic cardiac comorbidity is a contraindication to adjuvant trastuzumab in breast cancer patients. However, some patients with asymptomatic, nonlimiting cardiac comorbidity and normal baseline left ventricular ejection fraction (LVEF) receive adjuvant trastuzumab in the clinical practice. We sought to describe the tolerability of trastuzumab in these patients. PATIENTS AND METHODS: Retrospective analysis of patients with baseline asymptomatic, nonlimiting cardiac comorbidity receiving adjuvant trastuzumab at six Institutions between July 2007 and January 2016. RESULTS: Thirty-seven patients with HER2-positive, surgery treated breast cancer at high risk of relapse were studied. Median age was 64 years (range 36-82), median baseline LVEF 61% (range 50%-85%). Thirteen patients (35%) received trastuzumab with adjuvant anthracycline and taxane-based regimens, 19 (51%) with taxane-based, three (8%) with off-label vinorelbine and two (5%) with off-label endocrine therapy. Most frequent cardiac comorbidities were ischemic heart disease (35%), valvular disease (30%), atrial fibrillation (19%), and conduction disorders (14%). Nine patients (24.3%) experienced a cardiac event: congestive heart failure (one patient, 3%), asymptomatic LVEF reduction (six patients, 16%), and rhythm disturbances (two patients, 5%). Trastuzumab had to be discontinued either permanently (five patients, 14%) or temporarily (two patients, 5%). At the time of last follow-up visit, all patients showed LVEF within normal limits, except one who had experienced a symptomatic cardiac event (LVEF value at last follow-up 46%). CONCLUSIONS: Caution is needed in patients with significant ongoing cardiovascular risk factors, but when adjuvant trastuzumab is deemed beneficial on breast cancer outcomes, nonlimiting cardiac comorbidity should not preclude treatment.

A retrospective analysis of the activity and safety of oral Etoposide in heavily pretreated metastatic breast cancer patients.

Metastatic breast cancer (MBC) patients derive benefit from chemotherapy, but options become limited after several prior chemotherapeutic regimens. Oral etoposide (VP-16) has previously been found to be clinically active in MBC patients in phase II trials. However, with increasing availability of other drugs, etoposide use has declined in spite of its unfavorable toxicity profile probably being overestimated. We therefore evaluated the clinical benefit and safety of oral etoposide in a population of MBC patients who had failed multiple regimens of currently used therapies. Sixty-six patients with MBC previously treated with a median of eight (range 2-13) regimens of therapy were eligible for the study. Patients received 50 mg/day oral etoposide in 20-day cycles with 1-week of rest. All patients were evaluated for clinical benefit (clinical benefit rate [CBR], complete response, partial response, and disease stabilization >24 weeks), progression-free survival (PFS), overall survival (OS), and toxicities. Median PFS was 4 months, CBR was 18% (overall response rate 4%), and median OS from the start of treatment was 11 months. Little clinically significant or high-grade toxicity were observed. No patients withdrew from treatment due to etoposide-induced toxicity. The favorable clinical response, low toxicity, and low cost of the drug suggest that etoposide is a viable option for patients with heavily pretreated MBC.

Positive signature-tagged mutagenesis in Pseudomonas aeruginosa: tracking patho-adaptive mutations promoting airways chronic infection.

The opportunistic pathogen Pseudomonas aeruginosa can establish life-long chronic infections in the airways of cystic fibrosis (CF) patients. Persistent lifestyle is established with P. aeruginosa patho-adaptive variants, which are clonal with the initially-acquired strains. Several reports indicated that P. aeruginosa adapts by loss-of-function mutations which enhance fitness in CF airways and sustain its clonal expansion during chronic infection. To validate this model of P. aeruginosa adaptation to CF airways and to identify novel genes involved in this microevolution, we designed a novel approach of positive-selection screening by PCR-based signature-tagged mutagenesis (Pos-STM) in a murine model of chronic airways infection. A systematic positive-selection scheme using sequential rounds of in vivo screenings for bacterial maintenance, as opposed to elimination, generated a list of genes whose inactivation increased the colonization and persistence in chronic airways infection. The phenotypes associated to these Pos-STM mutations reflect alterations in diverse aspects of P. aeruginosa biology which include lack of swimming and twitching motility, lack of production of the virulence factors such as pyocyanin, biofilm formation, and metabolic functions. In addition, Pos-STM mutants showed altered invasion and stimulation of immune response when tested in human respiratory epithelial cells, indicating that P. aeruginosa is prone to revise the interaction with its host during persistent lifestyle. Finally, sequence analysis of Pos-STM genes in longitudinally P. aeruginosa isolates from CF patients identified signs of patho-adaptive mutations within the genome. This novel Pos-STM approach identified bacterial functions that can have important clinical implications for the persistent lifestyle and disease progression of the airway chronic infection.

Diversity and Spread of Acetolactate Synthase Allelic Variants at Position 574 Endowing Resistance in Amaranthus hybridus in Italy.

Poor control of Amaranthus spp. with herbicides inhibiting acetolactate synthase (ALS) has been observed for several years in soybean fields in north-eastern Italy, but to date only a few ALS-resistant populations have been confirmed. An extensive sampling of putatively resistant Amaranthus accessions was completed in the Friuli Venezia Giulia region, across an arable land area of about 3000 km2. In total, 58 accessions were tested to confirm their resistance status, recognize the Amaranthus species, identify the mutant ALS alleles endowing the resistance and determine the efficacy of 3 pre-emergence herbicides. Most accessions resulted in cross-resistance to thifensulfuron-methyl and imazamox. Genomic DNA were extracted from single seeds with a newly developed protocol; an allele-specific PCR assay revealed the presence of the 574-leucine in 20 accessions, of the 574-methionine in 22, and of both alleles in 9 accessions. The two variants showed a different spatial distribution. All resistant populations were ascribed to A. hybridus. A. hybridus resistant to ALS herbicides is well-established in this Italian region and its resistance is due to two ALS mutant alleles. Metribuzin, clomazone and metobromuron can be used as alternative herbicides to be applied in pre-emergence and they should be integrated into the management strategies to limit the spread of resistance.

Genetic basis and origin of resistance to acetolactate synthase inhibitors in Amaranthus palmeri from Spain and Italy.

BACKGROUND: Amaranthus palmeri is an aggressive annual weed native to the United States, which has become invasive in some European countries. Populations resistant to acetolactate synthase (ALS) inhibitors have been recorded in Spain and Italy, but the evolutionary origin of the resistance traits remains unknown. Bioassays were conducted to identify cross-resistance to ALS inhibitors and a haplotype-based genetic approach was used to elucidate the origin and distribution of resistance in both countries. RESULTS: Amaranthus palmeri populations were resistant to thifensulfuron-methyl and imazamox, and the 574-Leu mutant ALS allele was found to be the main cause of resistance among them. In two Spanish populations, 376-Glu and 197-Thr mutant ALS alleles were also found. The haplotype analyses revealed the presence of two and four distinct 574-Leu mutant haplotypes in the Italian and Spanish populations, respectively. None was common to both countries, but some mutant haplotypes were shared between geographically close populations or between populations more than 100 km apart. Wide genetic diversity was found in two very close Spanish populations. CONCLUSION: ALS-resistant A. palmeri populations were introduced to Italy and Spain from outside Europe. Populations from both countries have different evolutionary histories and originate from independent introduction events. ALS resistance then spread over short and long distances by seed dispersal. The higher number and genetic diversity among mutant haplotypes from the Spanish populations indicated recurrent invasions. The implementation of control tactics to limit seed dispersal and the establishment of A. palmeri is recommended in both countries. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Prescribing pattern of anticoagulants in patients with cancer associated thrombosis: Results of a survey among MITO group and AIOM society.

INTRODUCTION: Low molecular weight heparin (LMWH) has been the backbone of the treatment of cancer associated thrombosis (CAT). Direct-acting oral anticoagulants (DOACs) have shown efficacy and safety not inferior to LMWH and guidelines included DOACs as an option for CAT treatment. Nevertheless, DOACs are still poorly prescribed in patients with cancer. The aim of this survey was to better understand prescription patterns of anticoagulants, in particular of DOACs, especially in gynecological cancers (GCs). METHODS: Our survey was made up of 21 questions, the last four questions addressed to medical doctors (MDs) involved in GCs. An invitation to complete the survey was sent by e-mail to 691 MITO (Multicentre Italian Trials in Ovarian cancer and gynaecologic malignancies) and 2093 AIOM (Associazione Italiana di Oncologia Medica) members. RESULTS: Overall, 113 MDs completed the questionnaire, 69 involved in GCs. Most respondents (46, 41%) were aged 30-40 years old, worked in public hospitals (59, 52.2%), were medical oncologists (86, 76.1%). LMWH was the preferred choice for the treatment of CAT (104, 92%). However, 89 respondents (78.8%) prescribed or asked to prescribe a DOAC for CAT. The major concern about DOACs was the difficulty in verifying the therapeutic effect and the absence of antidotes in case of bleeding (37.9%). In patients with GCs, DOACs were used with niraparib, olaparib, rucaparib and immune checkpoint inhibitors (ICIs) in less than 10 patients by 23%, 20%, 9% and 10.2% of respondents, respectively. CONCLUSION: The responders are aware of the Direct-acting oral anticoagulants option and would like to use them.

HER2-positive breast cancer cells resistant to trastuzumab and lapatinib lose reliance upon HER2 and are sensitive to the multitargeted kinase inhibitor sorafenib.

Trastuzumab has changed the prognosis of HER2 positive breast cancers. Despite this progress, resistance to trastuzumab occurs in most patients. Newer anti-HER2 therapies, like the dual tyrosine-kinase inhibitor (TKI) lapatinib, show significant antitumor activity, indicating that HER2 can be still exploited as a target after trastuzumab failure. However, since a high proportion of patients fail to respond to these alternative strategies, it is possible that cell escape from HER2 targeting may rely on HER2 independent pathways. The knowledge of these pathways deserve to be exploited to develop new therapies. We characterized two human HER2 overexpressing breast cancer cell lines resistant to trastuzumab and lapatinib (T100 and JIMT-1) from a molecular and biological point of view. Indeed, we assessed both in vitro and in vivo the activity of the multitarget inhibitor sorafenib. In both cell lines, the previously proposed mechanisms did not explain resistance to HER2 inhibitors. Notably, silencing HER2 by shRNA did not affect the growth of our cells, suggesting loss of reliance upon HER2. Moreover, we identified alterations in two antiapoptotic proteins Mcl-1 and Survivin which are known to be targets of the multikinase inhibitor sorafenib. Moreover, sorafenib, strongly inhibited the in vitro growth of T100 and JIMT-1 cells, through the downregulation of both Mcl-1 and Survivin. Similar results were obtained in JIMT-1 xenografts subcutaneously injected in NOD SCID mice. We provide preclinical evidence that tumor cells resistant to trastuzumab and lapatinib may rely on HER2 independent pathways that can be efficiently inhibited by sorafenib.

Tutorial for the Characterization of Fatty Acid Methyl Esters by Gas Chromatography with Highly Polar Capillary Columns.

The fatty acid composition of fats and oils is commonly determined by gas chromatography after preparing fatty acid methyl esters (FAME). Capillary columns coated with polyethylene glycol emerged as the preferred separation tool for the quantification of the polyunsaturated fatty acids contained primarily in marine oils. However, their selectivity is inadequate for measuring the trans fatty acids (TFA) contained in refined vegetable oils, dairy fats, and marine oils. Highly polar 100% poly(biscyanopropyl siloxane) capillary columns provide the necessary selectivity, but small differences in the phase polarity caused by column age, conditioning, or manufacturing variations affect the reproducibility of their separations of these complex samples. In this study, a simple procedure is described to compensate for small variations in column selectivity by adjusting the elution temperature. The balance between the dipole-induced dipole interactions and dispersive interactions was determined by measuring selectivity factors [SF(i)] corresponding to the elution of an unsaturated FAME such as 18:3n-3 relative to two saturated FAME such as 20:0 and 22:0. Knowing the SF(i) provided by the installed capillary column at a given elution temperature, and the SF(i) of the target separation, we propose a simple calculation to determine the necessary elution temperature adjustment to achieve (or restore) the desired separation. After determining the SF(i) which provides the optimal separation of TFA, the novel methodology was applied to the separation of refined vegetable oils, butter fats, and marine oils.

Population structure and evolution of resistance to acetolactate synthase (ALS)-inhibitors in Amaranthus tuberculatus in Italy.

BACKGROUND: Before 2010, Amaranthus tuberculatus (Moq.) J. D. Sauer was barely known to farmers and stakeholders in Italy. Since then, several populations resistant to acetolactate synthase (ALS)-inhibiting herbicides have been collected. In most populations, a known target site resistance-endowing mutation was found, a Trp to Leu substitution at position 574 of the ALS gene, but it was unclear whether they had evolved resistance independently or not. The aims of the work were (i) to elucidate the population structure of Italian ALS-resistant A. tuberculatus populations, and (ii) to analyze the ALS haplotypes of the various populations to determine whether resistance arose multiple times independently. RESULTS: In order to determine the population structure of eight A. tuberculatus populations, eight previously described microsatellite loci were used. Two ancestors were found: three populations derived from one, and five from the other. In the 4-kb ALS region of the genome, including the 2-kb coding region, 389 single nucleotide polymorphisms were found. In silico haplotype estimation was used to reconstruct the sequence of three distinct haplotypes carrying the Trp574Leu mutation. In addition, no mutation was found in 83% of plants of a single population. CONCLUSIONS: (i) Resistance must have arisen independently at least three times; (ii) at least one population was already resistant to ALS inhibitors when introduced in Italy; (iii) a single haplotype with a Trp574Leu mutation was shared among six populations, probably because of broad seed dispersal; and (iv) one population likely evolved nontarget site ALS inhibitors resistance. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

In-House Validation of an SPE-GC-FID Method for the Detection of Free and Esterified Hydroxylated Minor Compounds in Virgin Olive Oils.

Minor compounds in vegetable oils are distributed between free and esterified forms, and the ratio of these two fractions could represent an important parameter for assessment of oil authenticity. A simple method based on offline SPE-GC-FID for the analysis of free and esterified hydroxylated minor compounds in olive and sunflower oils has been developed and in-house validated. A satisfactory repeatability relative standard deviation (<7.5%) was obtained in all cases. The method, which requires simple instrumentation, allows for reliable quantification in a single chromatographic run with the advantages of minimizing sample manipulation, use of toxic solvents and reagents, and time consumption. The analytical procedure was applied to pure oil samples, including 15 authentic extra virgin olive oils collected from different European countries (Spain, Italy, Greece, and Portugal). Finally, the proposed SPE-GC-FID methodology could detect changes in the ratio between the free and esterified forms in pure extra virgin olive oil when mixed with refined sunflower oil at different percentages of 2, 5, 10, 15, and 20% (w/w) to simulate adulteration.

Target-Site Mutations and Expression of ALS Gene Copies Vary According to Echinochloa Species.

The sustainability of rice cropping systems is jeopardized by the large number and variety of populations of polyploid Echinochloa spp. resistant to ALS inhibitors. Better knowledge of the Echinochloa species present in Italian rice fields and the study of ALS genes involved in target-site resistance could significantly contribute to a better understanding of resistance evolution and management. Using a CAPS-rbcL molecular marker, two species, E. crus-galli (L.) P. Beauv. and E. oryzicola (Vasinger) Vasing., were identified as the most common species in rice in Italy. Mutations involved in ALS inhibitor resistance in the different species were identified and associated with the ALS homoeologs. The relative expression of the ALS gene copies was evaluated. Molecular characterization led to the identification of three ALS genes in E. crus-galli and two in E. oryzicola. The two species also carried different point mutations conferring resistance: Ala122Asn in E. crus-galli and Trp574Leu in E. oryzicola. Mutations were carried in the same gene copy (ALS1), which was significantly more expressed than the other copies (ALS2 and ALS3) in both species. These results explain the high resistance level of these populations and why mutations in the other ALS copies are not involved in herbicide resistance.

Impact of Baseline and On-Treatment Glycemia on Everolimus-Exemestane Efficacy in Patients with Hormone Receptor-Positive Advanced Breast Cancer (EVERMET).

PURPOSE: The mTOR complex C1 (mTORC1) inhibitor everolimus in combination with the aromatase inhibitor exemestane is an effective treatment for patients with hormone receptor-positive (HR+), HER2-negative (HER2-), advanced breast cancer (HR+/HER2- aBC). However, everolimus can cause hyperglycemia and hyperinsulinemia, which could reactivate the PI3K/protein kinase B (AKT)/mTORC1 pathway and induce tumor resistance to everolimus. EXPERIMENTAL DESIGN: We conducted a multicenter, retrospective, Italian study to investigate the impact of baseline and on-treatment (i.e., during first 3 months of therapy) blood glucose levels on progression-free survival (PFS) in patients with HR+/HER2- aBC treated with everolimus-exemestane. RESULTS: We evaluated 809 patients with HR+/HER2- aBC treated with everolimus-exemestane as any line of therapy for advanced disease. When evaluated as dichotomous variables, baseline and on-treatment glycemia were not significantly associated with PFS. However, when blood glucose concentration was evaluated as a continuous variable, a multivariable model accounting for clinically relevant patient- and tumor-related variables revealed that both baseline and on-treatment glycemia are associated with PFS, and this association is largely attributable to their interaction. In particular, patients who are normoglycemic at baseline and experience on-treatment diabetes have lower PFS compared with patients who are already hyperglycemic at baseline and experience diabetes during everolimus-exemestane therapy (median PFS, 6.34 vs. 10.32 months; HR, 1.76; 95% confidence interval, 1.15-2.69; P = 0.008). CONCLUSIONS: The impact of on-treatment glycemia on the efficacy of everolimus-exemestane therapy in patients with HR+/HER2- aBC depends on baseline glycemia. This study lays the foundations for investigating novel therapeutic approaches to target the glucose/insulin axis in combination with PI3K/AKT/mTORC1 inhibitors in patients with HR+/HER2- aBC.

A family affair: resistance mechanism and alternative control of three Amaranthus species resistant to acetolactate synthase inhibitors in Italy.

BACKGROUND: Several soybean fields in Italy were found to be infested by multiple species of Amaranthus spp. not adequately controlled by acetolactate (ALS) inhibitor herbicides. The objectives of this research were (i) to create a simplified botanical key to identify weedy amaranths; (ii) to determine the number and type of sites of action the accession are resistant to, i.e. resistance pattern; and (iii) to determine the main resistance mechanisms involved d) to evaluate the efficacy of herbicides with different site of action. RESULTS: An easy-to-use botanical key was devised and successfully used in the infested sites and results were confirmed through a species-specific molecular marker. Amaranthus retroflexus L. (redrood pigweed) was found in three sites; plants with Asp376 Glu substitution at the ALS gene were resistant to thifensulfuron-methyl. Amaranthus tuberculatus (Moq.) J.D.Sauer (waterhemp) and Amaranthus hybridus L. (smooth pigweed) accessions were cross-resistant to thifensulfuron-methyl and imazamox; most ALS-resistant plants had a point mutation at position 574. One A. hybridus accession had the substitution Trp574 Met, new for Amaranthus genus. All ALS-resistant accessions were controlled by glyphosate and metribuzin. A. retroflexus accessions were controlled by bentazon, instead an A. hybridus and some A. tuberculatus accession were not. CONCLUSIONS: The simplified botanical key proposed herein could be a useful tool for farmers and weed scientists to reliably identify Amaranthus species in the field. The main resistance mechanism in the three Amaranthus species is target-site mediated. This is the first evidence of ALS-resistant A. tuberculatus outside its native North American range. © 2019 Society of Chemical Industry.

Molecularly Imprinted Polymer as Selective Sorbent for the Extraction of Zearalenone in Edible Vegetable Oils.

A method based on the selective extraction of zearalenone (ZON) from edible vegetable oils using molecularly imprinted polymer (MIP) has been developed and validated. Ultra-high-pressure liquid chromatography coupled with a fluorescence detection system was employed for the detection of zearalenone. The method was applied to the analysis of zearalenone in maize oil samples spiked at four concentration levels within the maximum permitted amount specified by the European Commission Regulation (EC) No. 1126/2007. As a result, the proposed methodology provided high recoveries (>72%) with good linearity (R2 > 0.999) in the range of 10-2000 µg/kg and a repeatability relative standard deviation below 1.8%. These findings meet the analytical performance criteria specified by the European Commission Regulation No. 401/2006 and reveal that the proposed methodology can be successfully applied for monitoring zearalenone at trace levels in different edible vegetable oils. A comparison of MIP behavior with the ones of QuEChERS and liquid-liquid extraction was also performed, showing higher extraction rates and precision of MIP. Finally, the evolution of ZON contamination during the maize oil refining process was also investigated, demonstrating how the process is unable to completely remove (60%) ZON from oil samples.

Temperature Dependence of Oxidation Kinetics of Extra Virgin Olive Oil (EVOO) and Shelf-Life Prediction.

Producers have to guarantee the extra virgin olive oil (EVOO) quality characteristics reported in the Regulation (CEE) 2568/91 throughout the product shelf-life (SL). Unfortunately, due to the development of oxidative reactions, some quality indices change during storage leading to a progressive deterioration of EVOO quality. To avoid the risk of product downgrading in the virgin oil category, the development of effective shelf-life prediction models is extremely important for the olive oil industry. In this research, the accelerated shelf-life testing (ASLT) protocol was applied to evaluate the temperature dependence of selected oxidation indexes as well as to develop a shelf-life predictive model. The evolution of conventional (peroxide value, K232, K270, polyphenols, tocopherols and hexanal) and unconventional parameters (conjugated trienes and pyropheophytin a) was monitored in bottled EVOO stored in the dark at increasing temperature (25, 40, 50 and 60 °C). Accordingly, for well-packed products with reduced oxygen in headspace, the best shelf-life index allowing the ability to predict EVOO SL turned out to be K270. In addition, pyropheophytin a (%) has been shown to be more sensitive to temperature changes than the secondary oxidation indices, thus suggesting its use as a freshness indicator for storage temperatures higher than 25 °C.

Transcriptional wiring of the TOL plasmid regulatory network to its host involves the submission of the sigma54-promoter Pu to the response regulator PprA.

Implantation of the regulatory circuit of the degradation pathway of TOL plasmid pWW0 in the native transcriptional network of the host Pseudomonas putida involves interplay between plasmid- and chromosome-encoded factors. We have employed a reverse genetics approach to investigate such a molecular wiring by identifying host proteins that form stable complexes with Pu, the sigma(54)-dependent promoter of the upper TOL operon of pWW0. This approach revealed that the Pu upstream activating sequences (UAS), the target sites of the cognate activator XylR, form a specific complex with a host protein which, following DNA affinity purification and mass spectrometry analysis, was identified as the LytTR-type two-component response regulator PprA. Directed inactivation of pprA resulted in the upregulation of the Pu promoter in vivo, while expression of the same gene from a plasmid vector strongly repressed Pu activity. Such a downregulation of Pu by PprA could be faithfully reproduced both in vitro with purified components and in an in vivo reporter system assembled in Escherichia coli. The overlap of the PprA and XylR binding sites suggested that the basis for the inhibitory effect on Pu was a mutual exclusion mechanism between the two proteins to bind the UAS. We argue that the binding of the response regulator PprA to Pu (a case without precedents in sigma(54)-dependent transcription) helps to anchor the TOL regulatory subnetwork to the wider context of the host transcriptome, thereby allowing the entry of physiological signals that modulate the outcome of promoter activity.

Treating advanced breast cancer with metronomic chemotherapy: what is known, what is new and what is the future?

The prognosis for patients with locally advanced or metastatic breast cancer (mBC) remains poor, with a median survival of 2-4 years. About 10% of newly diagnosed breast cancer patients present with metastatic disease, and 30%-50% of those diagnosed at earlier stages will subsequently progress to mBC. In terms of ongoing management for advanced/metastatic breast cancer after failure of hormonal therapy, there is a high medical need for new treatment options that prolong the interval to the start of intensive cytotoxic therapy, which is often associated with potentially serious side effects and reduced quality of life. Oral chemotherapeutic agents such as capecitabine and vinorelbine have demonstrated efficacy in patients with mBC, with prolonged disease control and good tolerability. Use of oral chemotherapy reduces the time and cost associated with treatment and is often more acceptable to patients than intravenous drug delivery. Metronomic administration of oral chemotherapy is therefore a promising treatment strategy for some patients with mBC and inhibits tumor progression via multiple mechanisms of action. Ongoing clinical trials are investigating metronomic chemotherapy regimens as a strategy to prolong disease control with favorable tolerability. This article provides an overview of metronomic chemotherapy treatment options in mBC, with perspectives on this therapy from a panel of experts.

Structural Determination and Occurrence in Ahiflower Oil of Stearidonic Acid Trans Fatty Acids.

Several marine oils and seed oils on the market contain relevant quantities of stearidonic acid (18:4n-3, SDA). The formation of 18:4n-3 trans fatty acids (tFA) during the refining of these oils necessitates the development of a method for their quantification. In this study, 18:4n-3 was isolated from Ahiflower and isomerized to obtain its 16 geometric isomers. The geometric isomers of 18:4n-3 were isolated by silver ion HPLC (Ag+ -HPLC) and characterized by partial reduction with hydrazine followed by gas chromatography analysis. The elution order of all 16 isomers was established using a 100 m × 0.25 mm 100% poly(biscyanopropyl siloxane) capillary column and at the elution temperature of 180 °C. The 4 mono-trans-18:4n-3 isomers produced during the refining of oils rich in 18:4n-3 were chromatographically resolved from each other, but c6,t9,c12,c15-18:4 coeluted with the tetra-cis isomer. These 2 fatty acids (FA) were resolved by reducing the separation temperature to 150 °C, but this change caused tetra-cis-18:4n-3 to coelute with t6,c9,c12,c15-18:4. Combining the results from 2 isothermal separations (180 and 150 °C) was necessary to quantify the 4 mono-trans 18:4n-3 FA in Ahiflower oil.

Self-evaluation of duration of adjuvant chemotherapy side effects in breast cancer patients: A prospective study.

BACKGROUND: We recently reported that self-evaluation of the incidence and severity of treatment-related side effects (TSEs) using a National Cancer Institute (NCI) Common Terminology Criteria for Adverse Events (CTCAE) v4.0-based questionnaire was feasible and more informative than doctor reports in patients undergoing standard adjuvant chemotherapy for operable breast cancer. Here, we compare self- and doctor-evaluated day of onset and duration of TSEs in the same population. PATIENTS AND METHODS: Six hundred and four patients were enrolled at 11 sites in Italy. CTCAE v4.0 definitions of grade of severity of nausea, vomiting, constipation, anorexia, dysgeusia, diarrhea, fatigue, pain, paresthesia, and dyspnea were translated into Italian and rephrased. Questionnaires were administered after the first and third chemotherapy cycles. At each time-point, information on TSEs was extracted from the medical charts and compared to patient questionnaires. RESULTS: A total of 594 and 573 paired patient and doctor questionnaires were collected after cycles one and three, respectively. TSE duration was significantly longer when reported by patients compared to doctors for six and seven of ten items after cycles one and three, respectively. Due to the combined effect of doctor underreporting of TSE incidence and duration, the mean percentages of cycle days with TSEs were significantly higher for all ten items when based on patient reports. Day of onset could not be evaluated because of insufficient numbers of complete records. CONCLUSIONS: Self-reporting TSE duration is feasible using a CTCAE-derived questionnaire. As doctors tend to underestimate TSE incidence and duration, patient-reported outcomes should be incorporated into clinical practice, perhaps using eHealth technologies, to harness their potential to better estimate total TSE burden.