The detection and monitoring of lithium in environmental and clinical settings is becoming increasingly important. In this review, sensors incorporating conductive polymers and lithium bronzes are discussed, together with electrochemical and spectroscopic approaches. Ionophore-based methods have been employed extensively, with varying degrees of selectivity and sensitivity, and these are discussed in depth. Lithium is present in the Earth’s crustal rocks at a concentration of 20–60 ppm, and in 0.2 ppm in seawater, from which it is extracted for a variety of uses, including lightweight metal alloys, batteries, ceramics and glasses, and medication . Consequ+ concentrations in biofluids such as blood and urine accurately and precisely is consequently a vital target for analysis. Uncontrolled high concentrations of Li+ can permanently affect the nervous system and kidneys phenol substituent (22) to produce a chromogenic Li+-specific ligand styryl]acridine and KTpClPB in a PVC membrane to measure Li+ in pharmaceuticals and saliva, over a 1 × 10−4 to 1 × 10−2 mol dm−3 concentration range . La. La25, wphore 26 , showed  at best . Selectiphore 27  40]. Mo. Mo25, ws tested .28  sensor 28  42]. Fl. Fl28  as the Li+-binding element of an electrode with a self-assembled monolayer modified surface  + complexes which would be more facile in solution than when the macrocycles are tethered to a surface.Wanichacheva et al. employed a cryptand attached to an alkane thiol ( surface  44]. Le. Le30) ahan 2.0) . The res+ fluoroionophores that incorporated a coumarin [32 and 33  dimethacrylate (n = 16) as the crosslinker, gave a response time of 60 s. It was reversible, however, its Li+ binding strength was almost an order of magnitude larger than its non-polymerized analogue. The group’s Li+-selective sensor, 34 (34 was though urethane bond formation between its terminal hydroxyl function and an isocyanate-modified glass surface. In an improvement over 33, the material’s response was found to be reversible between 1 × 10−4 mol dm−3 to above 0.1 mol dm−3, and no interference was reported for Na+, K+, Mg2+ or Ca2+ up to 1 × 10−1 mol dm−3.Citterio and Suzuki developed 14-crown-4 Licoumarin  or boroncoumarin  group. I2 and 33 , were prnsor, 34 , was inc+ in preference to other metal cations [35 (+ generated increases in fluorescence which could be reversed with white light. Reaction with N-hydroxysuccinamide gave derivative 36 (+ could be washed out during the ‘off’ cycle. Furthermore, multiple ‘on’ and ‘off’ cycles could be achieved with very little attenuation of output intensity. Abell combined the nitrophenol chromogenic moiety and crown ether receptor first reported by Pacey  and intr cations ,49. Inveions [35  showed tative 36 , which c37 (−2 mol dm−3 alkali metal salt solutions [38 (+ [37 which is able to adopt a conformation that preferentially bound Li+ over other alkali metals (39 (Kothur et al. attached pillar  arene 3737  to gold olutions . In prevolutions  arene 38ions [38  was incos [38 (+ . This hii metals . No seletals (39 , an indi40 (4 was added. Dissociation constants, calculated from fluorescence quantum yields of the zinc azaphthalocyanines and their complexes, gave 6.4 × 10−2 mol dm−3 for Zn·40·Li+ and 1.5 × 10−2 mol dm−3 for Zn·40·Na+, with all other cations having values below 1 × 10−2 mol dm−3.Lochman et al. prepared a series of crown ether-appended rigid, planar fluorescent azaphthalocyanines and demonstrated the expected selectivity based on the fit of the cation to the crown . The zin40  gave a sThese developments in synthetic chemistry and electrode composition have had an impact on the selectivity for lithium; to illustrate this, the properties of a range of systems are given in +-ionophore affinity possible in numerous ways, from in-line clinical monitoring to environmental sampling. As the need to accurately determine Li+ concentrations grows, so will the ingenuity of those methods designed for the detection of Li+. The need to detect and monitor lithium both in the environment and in clinical settings has led to several approaches in sensing this element. Its emission spectrum can be used to determine its concentration, as can its X-ray fluorescence spectrum, albeit indirectly. Conductive polymers and lithium bronzes have been investigated, but ionophore-based sensors, whether colorimetric, fluorescent or electrochemical, clearly have the greatest potential for specificity and selectivity. Examples described include solution phase and surface-bound ionophores that make detection based on Li


Despite best evidence-based treatment including statins, residual cardiovascular risk poses a major challenge for clinicians in the twenty first century. Atherogenic dyslipidaemia, in particular elevated triglycerides, a marker for increased triglyceride-rich lipoproteins and their remnants, is an important contributor to lipid-related residual risk, especially in insulin resistant conditions such as type 2 diabetes mellitus. Current therapeutic options include peroxisome proliferator-activated receptor alpha (PPARα) agonists, (fibrates), but these have low potency and limited selectivity for PPARα. Modulating the unique receptor–cofactor binding profile to identify the most potent molecules that induce PPARα-mediated beneficial effects, while at the same time avoiding unwanted side effects, offers a new therapeutic approach and provides the rationale for development of pemafibrate , a novel selective PPARα modulator (SPPARMα). In clinical trials, pemafibrate either as monotherapy or as add-on to statin therapy was effective in managing atherogenic dyslipidaemia, with marked reduction of triglycerides, remnant cholesterol and apolipoprotein CIII. Pemafibrate also increased serum fibroblast growth factor 21, implicated in metabolic homeostasis. There were no clinically meaningful adverse effects on hepatic or renal function, including no relevant serum creatinine elevation. A major outcomes study, PROMINENT, will provide definitive evaluation of the role of pemafibrate for management of residual cardiovascular risk in type 2 diabetes patients with atherogenic dyslipidaemia despite statin therapy. Preventing cardiovascular disease (CVD) is a key challenge facing clinicians world-wide this century. Despite improvement in CVD mortality in developed regions, low to middle-income countries have a growing burden of CVD mortality and disability, driven by increasing rates of obesity, diabetes and dyslipidaemia –3. IndeeLow-density lipoprotein cholesterol (LDL-C) is undoubtedly the primary priority for lipid management to prevent CVD , 7. StatInhibition of cholesteryl ester transfer protein (CETP), which mediates the heteroexchange of triglycerides from VLDL or LDL and cholesteryl esters from HDL, was considered a possible therapeutic approach. However, the first three CETP inhibitors had either safety issues (torcetrapib) or no effect on clinical cardiovascular outcomes in high-risk patients  –16. The With growing recognition of the importance of targeting atherogenic dyslipidaemia comes a re-evaluation of available therapeutic options. Fibrates, peroxisome proliferator-activated receptor (PPAR) α agonists, are the most logical option as an add-on to statin therapy for management of elevated TGs and atherogenic dyslipidaemia, typically associated with insulin resistant conditions , 23. WhiWith escalating rates of diabetes, obesity and CVD, especially in developing regions, management of atherogenic dyslipidaemia, as well as metabolic abnormalities and chronic inflammation typically associated with these conditions, is now a priority for reducing residual cardiovascular risk. This scenario highlights the need for new therapeutic options. Selective PPARα modulators–SPPARMα agents—could well provide one approach to resolving this challenge, and offer a more accessible therapeutic approach to managing this dyslipidaemia .This review describes the profile of a novel SPPARMα, determined using a search methodology as follows. The literature was searched using Medline, Current Contents, PubMed, and relevant references with the terms ‘residual cardiovascular risk’, ‘peroxisome proliferator-activated receptor’, ‘pemafibrate’, ‘K-877’, ‘atherogenic dyslipidaemia’, ‘triglycerides’, ‘type 2 diabetes’, ‘clinical trials.’ Main articles published in English between 2000 and 2017 were reviewed and included. The author also requested information regarding clinical trials and preclinical studies with pemafibrate from Kowa Company, Ltd., Japan.PPARs are nuclear hormone receptors which bind to DNA as a heterodimer with the Retinoid X Receptor (RXR), and together they recognise specific DNA sequences in and around target genes referred to as PPAR response elements (PPREs). There are three different types of PPARs, ɑ, β and γ . PPARα iWhen activated by the binding of either an endogenous ligand or synthetic PPAR agonist (such as a fibrate for PPARα), heterodimerisation with a ligand-activated RXR results in a conformational change, leading to the transrepression or transactivation of target genes. During transrepression, the activated PPAR binds to cytokine-activated transcription factors, such as nuclear factor kappa B or activator protein-1, and blocks the interaction between the activated transcription factors and the promoter region of the target gene, thereby preventing transcription and, in this example, reducing inflammation. In contrast, during transactivation, the activated PPAR binds to PPRE upstream of the target gene, and with the involvement of cofactors, renders the PPAR complex ‘transcriptionally active’ , 32–35. The focus of interest in this review, PPARɑ, plays a key role in metabolic homeostasis, regulating lipid metabolism, specifically HDL synthesis and metabolism, and VLDL turnover, by controlling the expression of key targets including apo A-I, A-II, A-V and C-III, lipoprotein lipase (LPL), scavenger receptor B1 (SR-B1), the ATP-binding cassette transporters ABCA1 and ABCG1, and acyl CoA synthetase , 35, 36.PPARs possess a large lipid-binding pocket capable of encompassing a range of endogenous ligands. On binding, each ligand triggers a unique conformational change,leading to differential pattern of coactivator recruitment, which in turn results in specific tissue- and gene-selective effects. It is important to note that while PPAR ligands may share cofactors leading to a shared biological response, there are also differences in cofactor selectivity, resulting in differing responses Fig. . Thus, mThe SPPARM concept has already been applied to PPARγ. The first such SPPARMγs, INT131 and MK0533, have been shown in preclinical studies to exhibit at least comparable antidiabetic effects to pioglitazone but with an improved adverse event profile , 38, 39.)) was identified as an agonist with very high potency and selectivity against human PPARα as a result of screening based on the SPPARMα concept. Pemafibrate differed from other PPARα agonists currently available, which were developed without definitive knowledge of their specific mechanism of action. Structurally, pemafibrate (K-877) has an acidic region as in other PPARα agonists, but with the addition of unique benzoxazole and phenoxyalkyl side-chains, resulting in greatly enhanced PPARα activity and selectivity . Th. ThData In summary, preclinical data show that pemafibrate modulated gene expression mediated by PPARα, which in turn led to improved beneficial effects on atherogenic dyslipidaemia, inflammation and atherosclerosis, when compared with current PPARα agonists. There was also evidence to suggest the possibility of novel targets, notably FGF21.Data on file, Kowa Company, Ltd.]. Key published trials are summarised in Table Pemafibrate is being investigated in a comprehensive clinical trial programme, initially in Japan and latterly in Europe/USA, as both monotherapy and add-on to statin therapy. To date, over 2300 patients, the majority with dyslipidaemia and over one-quarter with concomitant type 2 diabetes mellitus, have been studied . Similar findings have been reported in type 2 diabetes patients with dyslipidaemia [Interestingly, a clinical pharmacology study in patients with dyslipidaemia showed that pemafibrate 0.4 mg/day increased hepatic glucose uptake, as evaluated by the hyperinsulinaemic-euglycaemic clamp procedure . These fpidaemia .Furthermore, there were significant increases in plasma levels of FGF21 after treatment with pemafibrate 0.1–0.4 mg/day for 12 weeks compared with placebo, whereas no effect was observed with fenofibrate 100 mg/day , 58. TakAs add-on to statin therapy in 423 Japanese patients with hypertriglyceridaemia (> 200 mg/dL or 2.3 mmol/L), pemafibrate 0.2 mg/day reduced TGs by 47% (versus 0.8% with placebo) Table . UptitraPemafibrate (0.1–0.4 mg/day) was investigated in a large Phase II study in Europe in patients with atherogenic dyslipidaemia despite well controlled LDL-C levels on stable statin therapy . At the Taken together, these studies show that pemafibrate (0.2–0.4 mg/day) is effective in the management of hypertriglyceridaemia, with or without low plasma HDL-C concentration, either as monotherapy or against a background of stable statin therapy. There are encouraging data to suggest that pemafibrate improves glucose metabolism, which merits further study.Beyond efficacy, safety and tolerability are key considerations for any novel therapy. As already discussed, the SPPARM concept was extended to PPARα in the search for an agent with improved potency and selectivity so as to overcome clinical concerns with the use of fibrates, specifically relating to adverse effects on liver and renal function. Evidence from the phase II/III study  showed tImportantly, the available safety data show that treatment with pemafibrate does not appear to affect renal function parameters. In the integrated analysis of phase II/III Japanese studies, there was no change in estimated glomerular filtration rate (eGFR), whereas a significant decline in eGFR was observed with fenofibrate treatment over 12 weeks (p < 0.001) , consistIn conclusion, the available safety experience with pemafibrate is encouraging, with no clinically meaningful deleterious effects on renal or hepatic function. However, long-term data in real world practice are needed to fully evaluate the safety profile of this novel agent.The available data show the benefits of applying the SPPARM concept to PPARα, showing effective lowering of TGs, as well as other atherogenic measures, notably VLDL- and remnant cholesterol, in statin-treated patients with atherogenic dyslipidaemia. The safety profile of pemafibrate appears promising, with no evidence of clinically meaningful adverse effects on renal or hepatic function during treatment for up to 24 weeks, thus reinforcing the value of improved selectivity with this SPPARMα.The key question, however, is whether this novel SPPARMα impacts the high residual risk of cardiovascular events that persists in statin-treated patients with atherogenic dyslipidaemia, especially those with concomitant type 2 diabetes mellitus. Both the FIELD and ACCORD Lipid trials have failed to provide definitive answers with the use of fenofibrate, largely due to methodological reasons. The FIELD trial was initiated before statin use was considered first-line for prevention of CVD; indeed, the study criteria excluded patients with an indication for lipid-lowering therapy. The study population was at low to moderate global CVD risk, 75% had no prior CVD and only about one-third of patients had the characteristic atherogenic dyslipidaemic profile associated with type 2 diabetes  . While iTo address these remaining uncertainties regarding PPARα-targeted agents, PROMINENT (Pemafibrate to Reduce cardiovascular OutcoMes by reducing triglycerides IN diabetic patiENTs) has been initiated. This study aims to recruit 10,000 patients with type 2 diabetes mellitus and elevated TGs (≥ 200 mg/dL or 2.3 mmol/L and < 500 mg/dL (5.7 mmol/L) and low HDL-C (≤ 40 mg/dL or 1.0 mmol/L), with and without established CVD . PatientAnother area of potential therapeutic interest is in the management of non-alcoholic fatty liver disease (NAFLD). Already there are promising results from a phase II b trial with elafibranor (GFT-505), a PPAR-α/PPAR-δ agonist, that improving atherogenic dyslipidaemia and insulin resistance may have benefit in patients with non-alcoholic steatohepatitis (NASH) . With a db/db mice also showed that pemafibrate inhibited the diacylglycerol-protein kinase C-NAD(P)H oxidase pathway, resulting in suppression of lipid accumulation and oxidative stress in the kidney [Finally, potential benefits of pemafibrate on microvascular complications of type 2 diabetes, notably diabetic retinopathy, previously reported for fenofibrate , 73, mere kidney . Thus, gAtherogenic dyslipidaemia, prevalent among patients with type 2 diabetes and obesity, is a contributor to lipid-related residual cardiovascular risk. Accumulating evidence has led to renewed thinking about the importance of targeting this dyslipidaemia, in particular elevated TGs, to reduce this risk. Among current therapeutic options, fibrates are the logical option, but have relatively low potency and selectivity for PPARα. Extension of the SPPARM concept to PPARα has led to the development of pemafibrate, which in preclinical studies, has shown improved potency and selectivity compared with currently available fibrates. In clinical trials, pemafibrate was effective in the management of atherogenic dyslipidaemia, either as monotherapy or add-on to statin treatment. There were no clinically meaningful adverse renal effects, including no elevation in serum creatinine, and no significant effects on hepatic function. Long-term data are needed, however, for definitive evaluation of the benefit versus risk of this novel SPPARMα for reduction of residual cardiovascular risk. Finally, investigation of a possible role for pemafibrate in the management of microvascular complications of diabetes, notably diabetic retinopathy, as well as in NAFLD, may be merited. A novel approach to addressing residual cardiovascular risk may be well within our grasp.


The objective of the study was to reveal through pragmatic MCDA (EVIDEM) the contribution of a broad range of criteria to the value of the orphan drug lenvatinib for radioiodine refractory differentiated thyroid cancer (RR-DTC) in country-specific contexts.The study was designed to enable comprehensive appraisal  in the current disease context  of France, Italy and Spain. Data on the value of lenvatinib was collected from diverse stakeholders during country-specific panels and included: criteria weights ; performance scores (judgments on evidence—collected through MCDA systematic review); qualitative impacts of contextual criteria; and verbal and written insights structured by criteria. The value contribution of each criterion was calculated and uncertainty explored.Comparative effectiveness, Quality of evidence  and Disease severity (France) received the greatest weights. Four criteria contributed most to the value of lenvatinib, reflecting its superior Comparative effectiveness , the severity of RR-DTC (16–22%), significant unmet needs (14–21%) and robust evidence (14–20%). Contributions varied by comparator, country and individuals, highlighting the importance of context and consultation. Results were reproducible at the group level. Impacts of contextual criteria varied across countries reflecting different health systems and cultural backgrounds. The MCDA process promoted sharing stakeholders’ knowledge on lenvatinib and insights on context.The value of lenvatinib was consistently positive across diverse therapeutic contexts. MCDA identified the aspects contributing most to value, revealed rich contextual insights, and helped participants express and explicitly tackle ethical trade-offs inherent to balanced appraisal and decisionmaking.The online version of this article (doi:10.1186/s12885-017-3258-9) contains supplementary material, which is available to authorized users. N = 392) placebo-controlled, phase III clinical trial. Lenvatinib prolonged progression-free survival (PFS) by 14.7 months  and significantly reduced the risk of death after adjustment for placebo patients’ cross-over . [Lenvatinib is a tyrosine kinase inhibitor (TKI), indicated for the treatment of patients with progressive, locally advanced or metastatic, differentiated thyroid carcinoma, refractory to radioactive iodine (RR-DTC).  The effiSorafenib, another TKI, is the only other medicine for RR-DTC approved in Europe.  In the aLenvatinib carries orphan drug designations for papillary and follicular thyroid cancers based on their rarity and debilitating and life threatening nature, and the significant benefit it provides. , 15.Appraisal of new products for reimbursement, particularly orphan products, , 17 is cPragmatic multi-criteria decision analysis (MCDA) can enable holistic appraisals and helps reveal and tackle the ethical trade-offs between conflicting demands to facilitate accountable decisionmaking. –23 EVIDEThe objectives of this study were to assess the contribution of a broad range of decision criteria to the value of lenvatinib for RR-DTC from the perspective of three country-specific panels representing a diversity of stakeholders using pragmatic MCDA.The study was designed based on analysis of the context in which lenvatinib will be appraised Fig. . ComparaMCDA Evidence Matrices were created using a systematic review protocol (included in the EVIDEM framework) for identification, analysis, synthesis and reporting of evidence following good HTA practice  adapted Panels included policy decisionmakers, specialists, patient representatives, and methodologists with decisionmaking expertise, who were identified using predefined selection criteria  of each quantitative criterion was calculated as the product of its normalized weight  and standardized score (Sx = score/5). The overall MCDA value estimate (VE) is the sum of all criteria value contributions:Criteria weights, scores and impacts were analyzed as reported previously.  The valuValue estimates obtained using different weighting methods were compared using t-tests, paired at the level of the individual panelist. Consistency of retest data was assessed by calculating intra-rater correlation coefficients   for normIn each country, the top-3 highest ranking criteria accounted for 30 to 31% of the total weight: “Comparative effectiveness” , “Disease severity” , “Quality of evidence” , “Comparative safety” (France) and “Type of therapeutic benefit” (Spain) Fig. . The higAlthough the study was not powered to measure variations across categories of stakeholders, exploratory analysis indicated that patient representatives tended to assign higher weights to “Comparative patient-perceived health/PROs” than other panelists, and lower to “Comparative cost” , reflecting panelists’ perception of its impact on mortality  andLenvatinib’s “Comparative effectiveness” versus watchful waiting was considered high , based on a 14.7-month improvement in PFS and reduced risk of death after adjustment for cross-over. , 3 One pLenvatinib’s “Comparative safety/tolerability” versus watchful waiting received a moderately negative score , reflecting greater frequency of treatment-related serious adverse events (AEs) compared to placebo (30.3% vs 6.0%).  “ComparaAssessment of lenvatinib’s “Comparative PROs” also showed wide variations: although mean scores were slightly positive , SDs were very large . Panelists had difficulty assessing this criterion using the evidence available, which included PRO data from the sorafenib RCT  and utiFor “Quality of evidence”, panelists assigned a mean score of 3.19 based on an analysis of quality of the lenvatinib clinical program performed by the investigators. Comments included that the program featured a robust phase III study with a well-defined patient population, that evidence on PFS was strong and convincing, and that the ITC data seemed robust. However, there was concern about the lack of patient PRO data for lenvatinib and the effect of crossover on the validity of the OS data. One panelist commented that to receive the maximum score of 5 “there must be outcomes data reported by the patient and something definitive in terms of OS.”With respect to the type of benefit that lenvatinib provides, panelists’ scores reflected that extending life but not curing the disease, it represents a treatment that provides a moderate type of therapeutic benefit (mean 2.44).Lenvatinib’s “Comparative cost” was assessed based on manufacturer’s budget impact (BI) models. For Italy, these indicated an incremental cost versus watchful waiting of €19–24 million per year over five years, reflecting that lenvatinib would displace lower-cost off-label therapies  and allow treating more patients. Taking the availability of sorafenib into account, the incremental impact was estimated at €13–17 million per year. Panelists commented that the low incidence of RR-DTC limited its BI, which they judged, on average, as moderate to low, with mean scores of −1.81 versus watchful waiting and −0.50 versus sorafenib, with large variations . Lenvatinib’s “Comparative other costs”, which included potential savings due to fewer hospitalizations and physician visits, was judged, on average, as positive but small.There was general agreement across the three countries on assessment of “Disease severity”, “Size of population”, “Unmet needs”, “Comparative effectiveness”, “Type of preventive”, “Type of therapeutic benefit” and “Quality of evidence”, with less than 1 unit difference in mean scores across the three panels Fig. . HoweverFigure With watchful waiting as comparator, over 50% of positive value contributions were made by three criteria: “Comparative effectiveness” (21–22%) and “Unmet needs” (18–21%) in all three countries, together with “Disease severity” (19–20%) in France and Italy and “Quality of evidence” (18%) in Spain. “Comparative cost” and “Comparative safety/tolerability” contributed the most to reducing the value of lenvatinib, particularly in France. The MCDA value estimate for lenvatinib was 0.33 (SD 0.13) in both Italy and Spain and 0.22 (SD 0.11) in France.With sorafenib as comparator, the top-3 value contributors were “Disease severity” , “Comparative effectiveness” , “Quality of evidence” , “Type of therapeutic benefit”  and “Unmet needs” . “Comparative cost” negatively contributed to value in all three countries. The MCDA value estimate was 0.36 (SD 0.15) in Italy, 0.38 (SD 0.11) in Spain and 0.28 (SD 0.14) in France.Italian panelists agreed that lenvatinib, being a treatment for cancer, was aligned with the “mandate and scope of their healthcare system”, with a positive impact on its value Fig. . The “PoWith respect to “System capacity and appropriate use”, most Italian panelists agreed that their health system was prepared to ensure appropriate use of lenvatinib. “Stakeholder pressures and barriers” were thought by the majority to have no impact on the value of lenvatinib. Half of panelists considered that “Political, historical & cultural context,” would have a negative impact, noting that in Italy the drug may be subject to a risk sharing agreement, such as payment by results. Overall, consideration of the contextual criteria had a positive impact on the value of lenvatinib in Italy.Spanish panelists agreed that consideration of “Population priorities and access” had a positive impact on lenvatinib’s value Fig. ; howeverFrench panelists expressed a diversity of views regarding prioritization of rare diseases, with the majority indicating no or negative impact Fig. . Most oft-tests P > .05).Use of the HPA weighting method increased the mean (group) value estimates by less than 5% in Italy (absolute increases <0.02) and, depending on comparator, by 15 to 24% in France and Spain (absolute increases 0.04–0.08). These differences were not statistically significant  contributed negatively to value. The overall value of lenvatinib was positive in all analyses, with variability across individuals and countries , pointing to the impact of individual perspectives and different cultural backgrounds. The EVIDEM value scale is rooted in the triple aim of healthcare,  i.e., doAppraisals derived from the EVIDEM methodology have been completed for a number of interventions, including drugs, devices and diagnostic tests, with value estimates ranging between 0.22 and 0.72 , 42–44. Weighting revealed that criteria, often not explicitly considered in appraisal processes such as disease severity, are important, confirming results from large surveys among healthcare decisionmakers and stakeholders. , 46 ConsThe scoring exercise showed a broad consensus in judgments on the severity of RR-DTC and the limitations of current treatments. There was also general agreement that lenvatinib provides major improvements in efficacy over watchful waiting as well as over sorafenib. Also, although their assessments varied in degree, all panelists agreed that the toxicity profile of lenvatinib was a limitation . In the SELECT trial, grade 3 or higher toxicities were seen in 75% of patients, resulting in dose reductions, dose interruptions and discontinuations in 67%, 82%, and 14% of patients, respectively. The most frequent grade 3 or higher treatment-related AEs were hypertension (42%), proteinuria (10%), fatigue (9%), diarrhea (8%), arterial and venous thromboembolic effects , acute renal failure (1.9%), and hepatic failure (0.4%).  Broadly Where evidence was lacking, limited or ambiguous, such as for PRO outcomes, clinical guidelines (which at the time of the study had not been updated), and some economic outcomes, performance scores typically differed widely (or score ranges were assigned), reflecting uncertainty in judging the evidence. Discussions with stakeholders allowed identifying which evidence is acceptable and most useful for a specific country, as related to cultural values. For example, French panelists noted that PFS data were very strong and relevant, while PROs derived from the general population are irrelevant in the French context.Contextual criteria were considered qualitatively in this study in terms of type of impact on overall value; however, some of these criteria could be quantitatively operationalized in specific contexts . ConsideThe study had some limitations. The 8-member panels were too small to be regarded as representative of their country. Clearly, individuals vary in their assessments, which may be influenced by personal and professional factors, such as experience, role in society and education. This study was not designed to investigate the impact of these factors on assessments; however, it included a diversity of stakeholders in an attempt to capture a broad variety of perspectives Additional file 2:Targeted systematic literature review methodology. (DOCX 41 kb)Additional file 3:MCDA evidence matrix for lenvatinib in the Italian context. (DOCX 49 kb)Additional file 4:Recruitment criteria for panelists. (DOCX 31 kb)Additional file 5:Exploratory analysis of weights by category of panelists. (DOCX 33 kb)


This study was performed to compare the efficacies and acute toxicities in weekly- and three weekly- cisplatin based concurrent chemoradiotherapy (CCRT) for advanced HNC patients.779 patients of 10 studies were eligible. No difference in the 2-, 3-year OS or 1-, 2-year LRFS was observed, whereas patients in three weekly CCRT arm tended to have a better 5-year OS . Weekly arm seemed to show less gastrointestinal toxicities , but similar hematologic toxicity compared to three weekly arm. Subgroup analysis displayed more grade ≥3 mucositis , and more chemotherapy related delay/interrupt  in weekly arm for non-nasopharynx carcinoma (non-NPC) HNC.We conducted the meta-analysis by searching PubMed, MEDLINE, ScienceDirect, Cochrane Library and China National Knowledge Infrastructure (CNKI) databases. The primary endpoint was overall survival (OS) with secondary endpoints locoregional recurrence-free survival (LRFS) and grade≥3 acute adverse events. RevMan 5.2 was used to perform statistical analyses.Three weekly cisplatin-based CCRT might achieve a higher long-term OS with no significant difference in a shorter OS and LRFS. Weekly arm was associated with less gastrointestinal toxicities but more grade≥3 mucositis and chemotherapy related delay/interrupt. Large randomized trials were urgent to further define superiority of these two regimens. A 100Multiple studies compared the outcomes of the weekly and three-weekly cisplatin-based CCRT in advanced HNC. A randomized control trial was conducted for patients with advanced oral squamous cell carcinoma, showing higher compliance and lower acute toxicity in three-weekly arm . Though The preliminary literature screening yielded 3950 records from the five databases. Finally 10 studies –15 of 77Six studies , 13–15 rThree studies , 14–15 wGrade≥3 adverse effects were gathered from the enrolled studies, including hematologic toxicity  and non-hematologic toxicity, including gastrointestinal toxicity, dermatitis, mucositis.Six studies supplied data of grade≥3 neutropenia , 10–13 aNausea and/or vomiting were the most common gastrointestinal reactions for patients treated with ciplatin-based CRT. The data of grade≥3 nausea/vomiting were extracted from six eligible studies –13, 15 iSix eligible studies –11, 15 hEight articles , 12–15 o2= 59%, p=0.03). After looking all eligible studies through, in Ho KF's study [2= 35%, p=0.19). It is important to note that patients in weekly group suffered more chemotherapy delay/interrupt than three weekly patients   and three weekly (3cycles) from six studies -8cycles , 13, 15 's study , there w2 during the RT showed a better 5-year OS rate when compared to RT only  at the end of follow up [2 might lead to similar tumor control [2   , eventuaMany studies have confirmed that cisplatin acted as a radiosensitizer and cisplatin-based concurrent CRT had more toxicity than radiotherapy alone on severe (grade 3 or higher) adverse effects, whether weekly (p<0.001) or three weekly cisplatin (p=0.001) was administrated –26. It wOne of the most serious adverse reactions is mucositis, which might be attributed to wide RT portal (extending from the skull base to the root of the neck) and significant doses of RT delivered to the mucosal surfaces due to bulky primary or cervical nodes . In our In terms of gastrointestinal effects, patients in three weekly cisplatin group tended to occur more nausea and/or vomiting than those in weekly group . While a phase II trial fractionated CDDP into four daily doses during the concurrent CRT to reduce the emesis rate and improved tolerability . Same vi2 vs. 162 mg/m2, p<0.0001). Similar conclusion was supported by others [Kidney damage was also common adverse reaction during CRT . Study ry others –13. One y others , and onlChemotherapy administration costs and drug acquisition costs were the most significant components of treatment for inpatients and outpatients . Peter GNevertheless, we had to mention several limitations in this study. Firstly, there were only two RCTs eligible, while the other eights were retrospective. Secondly, not all articles reported the data of OS, progression-free survival (PFS), LRFS, disease-free survival (DFS), especially the long-term survival data. Thirdly, we only analyzed the acute toxicities, whereas, chronic toxicities were not obtainable, such as xerostomia, dysphagia, hearing loss, radiation-induced brain injury and so on.Taking together, weekly cisplatin CRT had no difference in overall survival but less comfort in treatment process compared with three weekly cisplatin regimen. It is important to proceed with a long-term follow-up of the chronic toxicities and survival events to explore which treatment could make patients benefit most. Kunieda F et al had launched a randomized phase II/III study in Japan , and we We performed a literature search in the PubMed, MEDLINE, ScineceDirect, Cochrane Library and CNKI databases published between 1982-2014. The key words were: “head and neck neoplasms” OR “head and neck cancer” OR “head and neck tumor” OR “head-neck tumors”, “cisplatin” OR “Cis-platinum” OR “Platinol” OR “CDDP”, “triweekly” OR “three weeks” OR “every three weeks”, “per week” OR “every week” OR “weekly” OR “once a week” and “randomized controlled trial” OR “randomized control trials” OR “randomized clinical trial” OR “RCT” OR “randomly allocation” OR “randomly” OR “random” OR “controlled” OR “trial” OR. We also searched for “lip” “oral cavity” “oropharyngeal” “hypopharyngeal” “nasopharyngeal” “laryngeal” “sinus” “salivary gland”, respectively. All of the eligible articles and their references were retrieved. We excluded reviews, case reports and animal experiments, moreover, meeting abstracts that had not been published full-text were also excluded. We would consider the studies as a single one if they had been published twice or more by the seam team and based on the same patient source.Following information were extracted from the included studies: first author, publication year, study design, treatment protocol, number of patients, staging information, acute adverse reactions and survival events. Two authors conducted the eligibility assessment and data verification independently. If agreement could not be reached between the two authors, a third author would participate in the discussion until reaching final agreement.http://sourceforge.net) and DerSimonian-Laird random effect analysis was used to estimate the difference. Subgroup analyses were conducted according to the range of follow-up times and lesion sites. Heterogeneity was assessed by forest plots, chi-squared (χ2) tests and I2 statistic percentages. P values below 0.05 were defined as significant outcomes. Fixed-effect model was applied when homogeneity was fine , otherwise, a random-effect model was used.Endpoints were determined as overall survival, locoregional recurrence-free survival and adverse events. RevMan 5.2 software (Cochrane Collaboration's Information Management System) was used to perform this meta-analysis. Risk ratios of adverse effect were calculated with the correspondent 95% confidence interval (CI), in addition, time-to-event data from individual trials were summarized by the log hazard ratio (HR) and its variance. If the trials did not report survival information directly, Kaplan-Meier curves were read by the Engauge Digitizer version 4.1 (free software downloaded from


Zea mays L.). The study was conducted over three consecutive years and consisted of four treatments (each replicated three times): i) RM with N application rate of 260 kg N ha−1 (RM-N260); ii) RM with 180 kg N ha−1 (RM-N180); iii) a traditional flat cultivation system without mulching (F) with 260 kg N ha−1 (F-N260); iv) F with 180 kg N ha−1 (F-N180). Mean soil water content during the maize growing season was increased by RM in 2013 only. However, RM increased the soil water storage significantly at the 3-leaf (V3) and 6-leaf stage (V6), and decreased evapotranspiration (ET) during pre-silking stage in all years. Compared to F, RM significantly improved maize grain yield by 79–123% in 2013, 23–25% in 2014, and 11–12% in 2015. Following three years of maize cultivation, soil inorganic N content increased substantially (two- to three-fold) in the RM system and 60% of the total inorganic N was accumulated in the top soil layers (0–60 cm) under the mulched ridge. Relative changes were much smaller in F, and most of inorganic N was stored in 0–20 cm and 100–160 cm soil layers. Generally, RM resulted in higher soil water storage during the pre-silking stage, which was the main reason for the improved maize grain yield. The nitrate leaching risk was reduced in RM-N180 compared with F, but nitrate leaching from the furrows between ridges was observed in RM-N260. However, the large increase in soil inorganic N content in RM-N180 after three years’ cultivation indicates an oversupply of N and a potential risk of N losses to the environment over the longer term. Our study indicates, therefore, that RM is a suitable system for maize cropping in the semi-arid region of the Loess Plateau, with benefits in water and N use efficiency, but recommendations for appropriate N application rates are required to ensure long term agricultural sustainability, accounting for grain yields and environmental impacts. The mechanisms for inorganic N accumulation under the RM system are not fully understood and warrant further investigation.The semi-arid region of the Loess Plateau is typical of rain-fed agricultural production in Northwestern China. In this area, the ridge mulched system (RM) is a widely-used measure to increase crop yield. The purpose of this study was to investigate the effect of RM on soil water and inorganic nitrogen (N) distribution, and grain yield of maize ( SeveralWater is the most limiting factor for crop production under rain-fed agriculture in arid and semiarid areas. However, studies have shown that the temporally irregular rainfall distribution and inefficient management of rainwater, rather than total rainfall amount, are the primary limits for crop production . Zhu et Fertilization can increase crop yield and enhance drought resistance of crops in arid and semiarid regions . HoweverMany studies have investigated the relationship between soil water content and crop yield to clarify the yield increase mechanism under plastic film mulching, however, very few have considered the impacts of the effects on water content and soil inorganic N distribution on both food security and sustainable agriculture. This study focused on a ridge mulched system in a maize crop on the Loess Plateau of China, taking crop production and environmental sustainability into consideration. The objectives were to 1) evaluate the effect of the ridge mulched system on soil water content, soil water storage, and water use efficiency; and 2) investigate soil inorganic N pools and distribution, considering the ridge and furrow as different units.22.1−3; pH, 8.4; soil organic carbon content, 9.5 g kg−1; total N content, 1.05 g kg−1; available phosphorus (Olsen-P) content, 20.7 mg kg−1; available potassium (NH4OAc-K) content, 133.1 mg kg−1; and mineral N content, 28.8 mg kg−1.The field experiments were conducted from 2013 to 2015 at the Changwu Agricultural and Ecological Experimental Station, located on the Loess Plateau of China . The annual mean air temperature is 9.2 °C and average annual rainfall is 582 mm, 73% of which occurs during the maize-growing season. The groundwater table is approximately 80 m below the surface. The cropping system in this area is one crop of maize or wheat per year. According to Chinese Soil Taxonomy, the soils are Heilutu, belonging to Cumuli-Ustic Isohumosols (light silt loam) . The soi2.2−1, as recommended for traditional cultivation; 260 kg N ha−1 as per local farmer practice, giving a total of four treatments . Plots were established a 5 × 10 m in a randomized block design with three replicate plots per treatment. Urea (N 46%) was used as N fertilizer in all fertilized treatments. In 2014 and 2015, all N fertilizer was applied manually to the soil surface prior to sowing and plowed in as a basal dressing. In 2013, 70% of the N fertilizer was applied as basal dressing and the remaining 30% was applied during the silking stage using a hole-sowing machine following precipitation. A total of 60 kg P ha−1 as calcium superphosphate (P2O5 12%) and 75 kg K ha−1 as potassium sulfate (K2O 45%) were applied simultaneously with the basal N fertilizer for each plot. Ridges and furrows were made following fertilizer application.This study included a ridge mulched system (RM) and a traditional cultivation system (i.e. flat cultivation without mulching) (F) over three maize-growing seasons . There w−1, typical of local practice. The plastic film in all treatments was 0.008-mm-thick transparent polyethylene. A high-yielding maize hybrid (Pioneer 335) was selected for this study. The maize was planted at the end of April and harvested at the end of September. There was no irrigation during the maize-growing season.The RM system consisted of alternating ridges (70 cm) and furrows (30 cm), with only the ridges being mulched with plastic film . Two row2.32O System, Decagon, USA) at 1 h intervals during 2013 with 1 h interval. Six ECH2O systems were installed and each consisted of five ECH2O-TE sensors, which were installed at soil depths of 10, 30, 60, 100, and 160 cm, to derive soil water contents at the 0–20, 20–40, 40–80, 80–120, and 120–200 cm layers. Sensors were calibrated before installation and the soil moisture at each depth in the four soil profiles was assumed as homogenous before the experiment  and F treatments . The soi3−-N) and ammonium-N (NH4+-N) concentrations were measured using a continuous flow analyzer  after KCl extraction. The nitrite-N (NO2−-N) was ignored in this study, because most samples were lower than detection limits. We used the NO3−-N and NH4+-N concentrations in dry weight equivalent soil and bulk density to calculate the inorganic N pools in kg ha−1 for the different depths. For RM, the soil water storage (SWS) and inorganic N pools were calculated by using the respective data for RM-R and RM-F and multiplying by the ridge to furrow ratios .Soil samples  were collected at 15 day interval during maize growing season with a soil corer (diameter: 5 cm). Soil samples were refrigerated at 4 °C prior to subsequent analysis. The soil nitrate-N  and the least significant difference (LSD) was used for multiple comparisons. Differences were considered statistically significant at P < 0.05.33.1Compared with the long-term average (1957–2009), the annual precipitation was 68.6, 170.0, and 63.8 mm lower in 2013, 2014, and 2015, respectively . The preSignificant differences between treatments in mean soil water content across all soil layers were only observed in 2013, although some soil layers were also shown the differences in 2015 . In 2013The RM treatments increased SWS during the early stage of the maize growing season. Compared with the F treatments, the average SWS in the RM treatments was 13, 78, and 23 mm higher at the 3-leaf stage (V3) and 23, 42, and 26 mm higher at the 6-leaf stage (V6) in 2013, 2014, and 2015, respectively . From thThe changes in SWS across the 0–200 cm soil depth between harvest and sowing (△SWS) was significantly higher in RM than that in F in 2013; while the evapotranspiration (ET) was lower in RM . No signThese finding suggest that maize grain yield was not limited by the amount of rainfall during the maize growing season or the cumulative annual rainfall but by the irregular distribution of rainfall over time and the inefficient management of rainwater in semiarid areas . Many st3.2The dynamics of soil inorganic N content in the 0–160 cm soil layer during the maize growing season for 2013–2015 in RM and F treatments are shown in On average, the inorganic N contents in the 0–10 cm soil layer for RM-R were 1.3–23.4 times and 1.5–13.1 times higher than that in other soil layers for RM-N180 and RM-N260, respectively, across the three years. High levels of inorganic N were mainly distributed in the 0–10 cm layer in the ridge with mulching, as observed in other studies . There aGenerally, two peaks of inorganic N in RM-R were observed in the 0–10 cm soil layer during the maize growing season. One occurred after fertilizer application, and the other during crop maturity and harvest stages. The high soil inorganic N during the later stage of the maize growing season, was probably explained by the low nutrient requirement for crop growth at that stage and by enhanced soil mineralization caused by the higher temperature . There w−1 to less than 20 mg kg−1) occurred in the inorganic N content of the top soil layers following several rainfall events in RM-F and peaks occured in the deeper soil layers (60–100 cm) after storms , especially in the treatment with the higher N application rate. This is most likely because the plastic film intercepted rainfall, which then flowed from the ridges into the furrows, enhanced the leaching of inorganic N, especially nitrate, from the top to deeper soil layers  for RM-F. The first was caused by fertilizer application, but the peaks in June were probably caused by the lateral movement of N from the ridge due to lateral water movement . A large decrease ; however, an increase was observed during the later stage of the maize growing season in deep soil layers (60–100 cm) at the higher N application rate, due to N leaching during rainfall.3.32O emission in RM  for RM-N180 and RM-N260, respectively. About 40% of the total inorganic N was stored in the 60–100 cm soil layers in F treatments . Howeveron in RM .Fig. 8In−1), but under RM increased significantly, by two and three times for RM-N180 and RM-N260, respectively, after three years of maize cultivation  for RM-N180, ranging from 75% to 91%. The treatments at the higher N application rate of 260 kg N ha−1 had lower PFP values, even under RM. Across the three years, with the increase in the inorganic N pool, the PFP values in RM increased in 2014 but decreased in 2015. −1 for high yield maize production. This indicated that although maize grain yield has a positive relationship with N input, high N input or high residual soil N could reduce N use efficiency and limit the grain yield  cultivation significantly changed the soil water and inorganic N distribution over the 3-year study period. Compared with F, RM improved the soil water storage significantly during the pre-silking stage by decreasing the soil evaporation and increasing the plant transpiration, which improved the water use efficiency and contributed to enhanced grain yield. Higher SWS at sowing played an important role in maize production in rain-fed agriculture on the Loess Plateau, thus treatments such as plastic film mulching or straw residue left after harvest may be helpful in harvesting and storing rainfall in fallow season for subsequent use by the following crop. The soil inorganic N in RM accumulated in the top soil layer (0–10 cm) under the mulched ridge, probably because of reduced N leaching, increased soil mineralization, and lateral N movement from the furrow to the ridge. This was associated with a decrease in nitrate leaching to deeper soil layers. However, the nitrate leaching from the furrow was observed in the RM system at the higher N application rate. Inorganic N accumulation under the RM system increased by two to three times after three years of maize cultivation, which may be explained by enhanced soil mineralization. However, the mechanism for soil inorganic N accumulation under plastic film mulching systems is not clear, with potential interactions between a number of factors and soil processes, and further study is required. Overall, our result showed that RM-N180 could obtain high grain yield with low risk of nitrate leaching after one year of cultivation, but after three years of cultivation was associated with increased inorganic N accumulation in the soil profile. Thus, the recommended N application rate for traditional maize cultivation in this region may not be suitable under RM. Development of new recommendations for N fertilizer application rates under RM are needed to ensure the sustainability of rain-fed agriculture on the Chinese Loess Plateau.


Agmatoploidy is a type of chromosome rearrangement that involves the fragmentation ofan entire chromosome complement, generating a diploid with double its originalchromosome number. Agmatoploidy and other related karyotype changes, such assymploidy , partial agmatoploidy,polyagmatoploidy, etc., are restricted to species with holokinetic chromosomes andare assumed to play an important role in their karyotype evolution. However, acritical review of the literature shows that examples of chromosome number doublingby agmatoploidy are rare and not clearly demonstrated, while partial agmatoploidy andpartial symploidy seem to be the same as ascending and descending disploidy,respectively. It is therefore proposed here that these terms should be avoided oreven abolished. Chromosome number variations found in different species of a genus are usually due todisploidy  or topolyploidy (a cell division error that produces an entire genome duplication) . In specLuzula were a little less than half of those of the diploid form andthose of the octoploid species approximately half those of the tetraploids". AsLuzula species have holokinetic chromosomes and X-ray-fragmentedholokinetic chromosomes are regularly inherited (Carexspecies (Cyperaceae) by Carex, chromosome numbersvary from n =6 to n = 66, including an almostcontinuous series of n =6 to n = 47 and more than 100species presenting different cytotypes  was originally proposed by Luzula could have long (AL type), medium (BL type) or short (CLtype) sized chromosomes, as in Carex. Species with 2n= 24BL that were closely related to species with 2n = 12AL appear tohave arisen by agmatoploidy, whereas species with 2n = 24 AL wereassumed to be polyploids. She also found karyotypes with "fragmentation" restricted toonly few chromosomes, later denominated partial agmatoploidy (Luzulaspicata the most common chromosome race had 2n = 24BL butthere were a few samples with 2n = 14 (10AL + 4BL) and2n = 12AL. Agmatoploid plants may also experience polyploidization,generating polyagmatoploids (toploids .The term symploidy (from Greek συν = together with), including complete simploidy andpartial symploidy, was proposed by Luzula species. The occurrence ofagmatoploidy in other genera of Juncaceae has been criticized because there is noevidence that they have holokinetic chromosomes .Similar variations in chromosome numbers and sizes or in DNA amount were later reportedfor several other species and genera, suggesting that agmatoploidy is an importantmechanism of chromosome evolution in plants with holokinetic chromosomes . It is iomosomes . In all Luzula spicatasubsp. spicata had 2n = 24BL whereas L.spicata subsp. conglomerata had 2n = 12AL,2n = 16 (8AL + 8BL), and 2n = 24BLwould be considered symploids and partial symploids, respectively, whereas if the lowestnumber is the basal one than 2n =16 and 2n = 24BLshould be, respectively, partial agmatoploid and complete agmatoploid.The interpretation of a karyotype change as agmatoploidy or symploidy depends on whichkaryotype/species is considered basal. For example, n = 24BL. AssuminLuzula species  and Samiacynthia (2n = 25-28), a moth species from a closely relatedfamily, showed that even a twofold variation in chromosome numbers most probablyreflected repeated disploidy events , with 2n = 6/7 to 2n = 46 (Akodon (rodents), with 2n = 10 to2n = 46 (Large chromosome number variation, however, is not limited to genera having holokineticchromosomes and is also found in genera of different families .LikewisAlthough the concept of agmatoploidy has been widely accepted by plant cytogeneticistssince it was first proposed over 65 years ago, there is still no compelling evidencethat supports the occurrence of chromosome number duplication by the simultaneousfragmentation of all of the chromosomes. Complete agmatoploidy, complete symploidy, andpolyagmatoploidy seem to be a misinterpretation of polyploidy. Partial symploidy andpartial agmatoploidy are indistinguishable disploidy. All these terms should, therefore,be abolished or at least avoided until more convincing evidence has been presented.


Elite parental lines are of great significance to crop breeding. To discover unique genomic loci associated with excellent economic traits in the elite cabbage inbred-line ‘01–20’, we performed comparisons of phenotypes as well as whole-genome insertion-deletion/simple sequence repeat loci between ‘01–20’ and each of its five sister lines. ‘01–20’ has a range of excellent agronomic traits, including early-maturing, and improvements in plant type and leaf colour. Eight unique loci were discovered for ‘01–20’ and ‘01-07-258’, another elite line similar to ‘01–20’ at the whole-genome level. In addition, two excellent double-haploid lines derived from a cross of ‘01–20’ also inherited these loci. Based on the quantitative trait locus association results, five of these loci were found to be associated with important agronomic traits, which could explain why the elite parent ‘01–20’ possesses greener outer leaves, a more compact and upright plant-type, rounder head, shorter core length, and better taste. Additionally, some of these loci have clustering effects for quantitative trait loci associated with different traits; therefore, important genes in these regions were analysed. The obtained results should enable marker-assisted multi-trait selection at the whole-genome level in cabbage breeding and provide insights into significant genome loci and their breeding effects. Elite parental lines, which usually possess a range of excellent properties and high combining ability, are important materials for hybrid breedinget al. reported that 80% of the U.S. soybean germplasm could be traced back to just 12 parental lineset al. reported that 19 elite parental barley lines contained 72% of the genetic variation of modern varietieset al. also estimated the parental contribution and coefficient of coancestry of 13 inbred maize lines with known pedigrees using restriction fragment length polymorphism and simple sequence repeat (SSR) markerset al. studied high yield and good performance using 35 elite chickpea lineset al. reported that alleles from the elite Spanish barley cultivar ‘Orria’ contribute to grain yield, reduce plant height and increase 1000-kernel weight, while alleles of chromosome 5H from the landrace ‘SBCC073’ contribute to early vigour, higher grain yield and earlier floweringWith the development of genetics, genomics and molecular biology, elucidating the genetic composition, and its associations with the characteristics, of elite parental lines at the molecular level has become a main approach for improving the accuracy and efficiency of parental selection in plant breeding. At present, studies of elite parents are mainly focused on field crops, such as soybean, barley and maize. Lorenzen 1 hybrids resulting from their crosses were 10, 6, 0, 0, 2 and 3, respectively, and the numbers of derived lines (excellent lines selected from their hybrids) were 9, 5, 0, 0, 3 and 0, respectively. To date, some of the hybrids still dominate the market of spring cabbage cultivars in China. The above data show that ‘01–20’ has made a major contribution to cabbage production. Why did ‘01–20’ become an elite parental line, while its sister lines did not? To answer this question, we analysed and compared ‘01–20’ and its five sister lines, as well as two excellent double-haploid (DH) lines derived from ‘01–20’, at the phenotypic and genomic levels, with the objective of identifying the ‘01–20’s distinctive genomic loci associated with excellent agronomic traits.However, few studies on elite parental lines have been reported in vegetable crops. For cabbage, we reported, and performed a pedigree analysis, on the elite inbred line ‘01–20’‘Early Vikings’, a conventional, early-maturing, spring cultivar introduced from Canada in 1966 by the Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences (IVF-CAAS), was used in this study as a population with genetic heterogeneity. Genetic heterogeneity was maintained in this population through open self-pollination every year in a greenhouse with honeybees isolated from the outside using a gauze covering.‘01–20’, an elite parental line, together with its five sister lines: ‘01-07-258’, ‘01-07-251’, ‘01-1-4’, ‘01–88’ and ‘01-16-5’, were all bred through a method of systemic selection from ‘Early Vikings’. ‘01–20’ is an elite, early-maturing, spring cabbage inbred line having upright and green outer leaves, little wax powder, a green and round head, high general and specific combining abilities, and a tender and crisp taste.As determined in our previous study, the numbers of inbred lines derived from ‘01–20’ and cultivars generated from it had reached 9 and 10, respectivelyIn addition to the six sister lines, ‘D77’ and ‘D83’, two excellent DH lines, selected from over 200 DH lines derived from the cross between ‘01–20’ and ‘96–100’ , two DH lines (‘D77’ and ‘D83’) and the parental population. In total, 25 traits from three categories were assessed : plant-tThese traits were assessed following standards described in ‘Descriptors and data standards for cabbage’Average values for each trait of each line were calculated from three randomly selected plants in each plot at the rosette or harvesting stage. Adjusted means for the traits were obtained and used for further analysis. Microsoft Excel 2007  and SPSS 12.0  software were used for statistical analyses and multiple comparisons.Whole-genome scanning for insertion/deletion (indel) and SSR loci was performed for all cabbage materials, using 406 pairs of SSR and indel primers, and the corresponding genetic maps. These markers and maps were developed in our previous study for gene mapping and the detection of quantitative trait loci (QTLs)82+ included), 1.6 μl of dNTP (2.5 mM each), 0.4 μl of Taq DNA polymerase (2.5 U/μl), 5 μl of DNA template (40 ng/μl), 0.6 μl of forward primer (10 μM), 0.6 μl of reverse primer (10 μM) and 9.8 μl of ddH2O, were used. The reaction mixture was incubated in a thermal cycler at 94 °C for 5 min, followed by 35 cycles of 94 °C for 30 s, 55 °C for 30 s and 72 °C for 45 s, and finally 72 °C for 7 min. The PCR products were separated on 8% polyacrylamide gels, which were then subjected to silver staining after running at 160 V for 1.5 hThe molecular marker assay protocol was as follows: polymerase chain reaction (PCR) mixture samples with a volume of 20 μl, which contained 2 μl of PCR buffer .The genes located in the regions associated with the distinctive loci were analysed and compared with those in Images in Similarity analyses among the sister lines were conducted using the software NTSYSpc2.11Phenotype measurements were carried out for all six sister lines , a genetically heterogeneous population compromising 28 individuals , and theGenerally, ‘01-07-258’ was the line most similar to ‘01–20’ in most traits, including Pt, leaf- and head-related traits ; Table 3Among the other sister lines, ‘01-07-251’ had the most upright Pt, the largest plant expansion (Pd and Ph), the highest Hw, the lowest Dmc and Cfc, and the largest Ss, but the longest maturing period (58.67 d) and the longest Cl (8.58 cm); ‘01-1-4’ had the most wrinkled leaf surface and the shortest Hm and Cl, but the smallest head (0.55 kg). ‘01–88’ had the most patulous Pt, a higher Hw and a shorter Hm, but the highest Dmc (6.42) and a low tolerance to splitting; and ‘01-16-5’ had a patulous plant type and greener outer leaves, but the highest Cl/Hvd (0.66) and Cfc (0.61), and the lowest tolerance to splitting. In addition, although ‘Early Vikings’ showed a relatively low uniformity, some individuals still showed outstanding performance in terms of Hm, Hw, leaf and head colour, Hsi and Hs .Of the two DH lines, ‘D77’ had the most upright Pt, the greyest Lc and Hc, the largest amount of Lw, the fewest leaves and the longest maturing period. ‘D83’ exhibited the largest plant expansion (Pd and Ph), the highest Hw, and the highest levels of both Dmc and Cfc. Both of these lines have excellent combining ability (data not shown), and thus their use in hybrid seed production is promising.Thus, ‘01–20’ had a range of excellent agronomic traits and no obvious defects, and it not only inherited the early-maturing trait from ‘Early Vikings’, but also exhibited improvements in Pt, Lc, and head- and quality-associated traits. The DH lines also inherited certain excellent traits from ‘01–20’, such as high Hw, low Cl/Hvd and compact Pt.For assays of indel/SSR loci, 406 pairs of SSR and indel primers were applied for the six sister lines, as well as the two DH lines. In total, 398 primer pairs amplified two alleles and eight amplified three alleles at each locus. Overall, 206 primer pairs were polymorphic, accounting for 50.74% of the total. Whole-genome scanning for indel/SSR loci for the six lines revealed that ‘01–20’ had one unique locus, ‘01-07-258’ had one unique locus, and ‘01–20’ and ‘01-07-258’ had six common loci, which were different from those of the other four sister lines . In totaIndel26 and Indel488 was shown to be 1.3 cM on C02, and the distance between scaffold29640 and Indel64 was 0.7 cM on C03. The distinctive loci were located on chromosomes C02, C03, C05, C08 and C09 . Some ofIndel139 on C05 , which explains why ‘01–20’ possesses greener leaves , which means that the Lc of ‘01–20’ tends to be yellowish rather than blue  that controls ammonium uptake capacity, indicating that strong selection on nitrogen uptake-related traits has occurred in rice23Fusarium wilt may be risk crop production as this disease is currently affecting cabbage-producing areas in northern China.Although elite lines can make major contributions to crop production and quality, excessively applying them may also create problems. During plant breeding, some important agronomic traits, such as disease resistance, high yield and excellent taste, are constantly under directional selection, resulting in significantly reduced genetic diversityElite parental lines are constantly subjected to the directional selection acting on genes that control desirable traits of agronomic importance during their domestication and improvement. Therefore, the genes or loci with a signature of selection from breeders should be identifiable by whole-genome nucleotide polymorphism scanning using DNA markers25et al. screened 501 maize genes with a signature of selection and identified 10 as agronomically important candidates because they showed evidence  of exposure to selective pressure. It was further confirmed that one of these, encoding a MADS box transcriptional regulator, experienced a selective sweep during maize domesticationet al. identified eight of the sequenced 1,095 maize genes through a selection test in diverse maize landraces and teosintes, and showed that their functions were consistent with agronomic selection for nutritional quality, maturity and productivity. Another way of identifying genes for corresponding traits is association analysis and QTL mapping using elite lineset al. detected several QTLs of important traits in rapeseed, including flowering time, Ph, protein content, oil content, glucosinolate content and grain yieldet al. identified marker–trait associations in 94 diverse elite wheat lines: marker XwPt-7187 was associated with kernel hardness, XwPt-1250 and XwPt-4628 with test weight, and marker Xgwm512 with PhWith the development of molecular biology and of the sequencing of numerous genomes, uncovering the genetic basis of important traits of elite parental lines has become the focus of theoretical and applied studies. One way to characterise genes responsible for phenotypic variation is based on the signature of selection in lines with particularly superior characteristics. Vigouroux B. oleracea reference genome acquired from BRAD  tool and set the score cutoff value to 400. The annotation of the functions of the genes included transmembrane transporter, transcription factor, ATP binding, kinase and cytochrome. Based on the alignments with Arabidopsis, some of the genes may be associated with related traits. For example, in Region 2 on chromosome C03, which is associated with Pd, Lw, Ll, Pl, Pw and Hs, the homologous gene ubiquitin ligases EOL1 can act with ETO1 and EOL2 collectively to regulate ethylene biosynthesis in Arabidopsis by controlling type-2 ACC synthase levelsSPI encodes a WD40/BEACH domain protein and shares a similar actin-regulating ARP2/3 pathway that affects plant growth in various organsYcf4 plays an essential role in the Photosystem I complexIn the current study, we compared elite line ‘01–20’ with five of its sister lines by scanning SSR/indel loci across the whole genome. We identified eight loci for which the elite lines were distinctive, which were further found to be associated with important agronomic traits, including Lc, Hc, Pt, leaf length, Pd and Hs. To discover interesting candidate genes at these loci, we performed a preliminary analysis for the genes located in the region for the five distinctive loci associated with important agronomic traits, according to the annotations for the The above results shed light on why ‘01–20’, rather than its sister lines, became an elite parental line. These loci could be useful for the development of whole-genome background markers for cabbage breeding and to promote our understanding of the genetic basis of selected traits.How to cite this article: Lv, H. et al. Genome-wide indel/SSR scanning reveals significant loci associated with excellent agronomic traits of a cabbage (Brassica oleracea) elite parental line 01-20. Sci. Rep.7, 41696; doi: 10.1038/srep41696 (2017).


Scientific Reports6: Article number: 22638; 10.1038/srep22638 published online: 03032016; updated: 10102016.This Article contains referencing errors. In the Introduction section,1−xCoxSi, Cu2OSeO33,4,5,6 with the bulk Dzyaloshinskii-Moriya interaction (DMI), has chiral crystal structure that supports vortex-like skyrmions. Another type of the systems are heavy metal/ultrathin ferromagnetic layers such as Fe/Ir, Ni/Co and Ta/CoFeB/TaOx10,11,28,29, that results in an interfacial DMI”.“One type of such systems such as MnSi, Feshould read:1−xCoxSi, Cu2OSeO33,4,5,6 with the bulk Dzyaloshinskii-Moriya interaction (DMI)28,29, has chiral crystal structure that supports vortex-like skyrmions. Another type of the systems are heavy metal/ultrathin ferromagnetic layers such as Fe/Ir, Ni/Co and Ta/CoFeB/TaOx10,11, that results in an interfacial DMI”.“One type of such systems such as MnSi, FeIn addition, in the Results and Discussion section,25–30”.“We would like to point out that sub-nanosecond current pulses can be routinely created in laboratories around the world since they have been used to manipulate the magnetic structures in both nanostrips and spin valvesshould read:25–27”.“We would like to point out that sub-nanosecond current pulses can be routinely created in laboratories around the world since they have been used to manipulate the magnetic structures in both nanostrips and spin valvesIn the Introduction section of the HTML version of the Article,11”.“It is noticed that the spin structure along the radial direction of a hedgehog skyrmion is very similar to that of a Neel wallshould read:30”.“It is noticed that the spin structure along the radial direction of a hedgehog skyrmion is very similar to that of a Neel wall