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Introduction: Daratumumab, lenalidomide, and dexamethasone (DRd) and bortezomib, lenalidomide, and dexamethasone (VRd) are the only preferred treatment regimens for patients with transplant-ineligible (TIE) newly diagnosed multiple myeloma (NDMM). As there are no randomized head-to-head studies of DRd versus VRd, this analysis aimed to compare real-world time-to-next-treatment (TTNT) or death in this population., Methods: Patients with NDMM who received front-line (FL) DRd or VRd were identified from the Acentrus database (January 1, 2018 to May 31, 2023). Those with a record of a stem cell transplant or aged < 65 years were excluded to limit analysis to the TIE population. Inverse probability of treatment weighting was used to balance baseline patient characteristics. A doubly robust Cox proportional hazards model was used to compare TTNT or death between cohorts., Results: A total of 149 and 494 patients who initiated DRd and VRd, respectively, were identified. After weighting (weighted N DRd  = 302, weighted N VRd  = 341), cohorts had similar baseline characteristics. Of these, 98 (32.4%) DRd and 175 (51.2%) VRd patients either received a subsequent line of therapy or died, with a median TTNT or death of 37.8 months in the DRd cohort and 18.7 months in the VRd cohort (hazard ratio: 0.58, 95% confidence interval: 0.35, 0.81; p < 0.001)., Conclusion: Treatment of TIE NDMM patients with DRd led to a significantly longer TTNT or death compared to VRd, evidenced by a 42% risk reduction, supporting the effectiveness of DRd over VRd as FL treatment in this patient population., (© 2024 Janssen Scientific Affairs, LLC. Cancer Medicine published by John Wiley & Sons Ltd.)

Introduction: Daratumumab plus lenalidomide and dexamethasone (D-Rd) and bortezomib plus lenalidomide and dexamethasone (VRd) are commonly used treatment combinations for transplant-ineligible (TIE) patients with newly diagnosed multiple myeloma (NDMM). D-Rd and VRd demonstrated superior efficacy relative to lenalidomide and dexamethasone (Rd) in the MAIA and SWOG S0777 trials, respectively, but have not been compared directly in a head-to-head trial. Naïve comparisons of efficacy across the two trials may be biased because MAIA enrolled only TIE patients (median age 73 years), whereas SWOG S0777 enrolled both TIE patients and transplant-eligible patients who chose to defer/refuse frontline stem cell transplantation (median age 63 years). The present study compared progression-free survival (PFS) in TIE patients with NDMM treated with D-Rd versus VRd based on an adjusted indirect treatment comparison (ITC) that leveraged individual patient-level data from MAIA and SWOG S0777., Methods: Harmonized inclusion/exclusion criteria (including age ≥ 65 years as a proxy for transplant ineligibility) and propensity-score weighting were used to balance the trial populations on measured baseline characteristics. After differences in trial populations were adjusted for, an anchored ITC was performed wherein within-trial PFS hazard ratios (HRs) for D-Rd versus Rd and VRd versus Rd were estimated and used to make indirect inference about PFS for D-Rd versus VRd., Results: PFS HRs were 0.52 (95% confidence interval [CI] 0.41-0.67) for D-Rd versus Rd based on MAIA data, 0.88 (95% CI 0.63-1.23) for VRd versus Rd based on SWOG S0777 data, and 0.59 (95% CI 0.39-0.90) for the Rd-anchored ITC of D-Rd versus VRd. Sensitivity and subgroup analyses produced results consistent with the primary results., Conclusion: This anchored ITC demonstrated a greater PFS benefit for D-Rd versus VRd in TIE patients with NDMM. In the absence of head-to-head trials comparing D-Rd and VRd, the present trial may help inform treatment selection in this patient population., (© 2024. The Author(s).)

Aim: Obtain clinical consensus on factors impacting first-line prescribing for transplant-ineligible (TIE) patients with newly diagnosed multiple myeloma (NDMM). Materials & methods: A double-blinded, modified Delphi panel was employed. USA-based hematologists/oncologists who treat TIE patients with NDMM were selected as expert panelists. Results: Consensus was reached that patient frailty, performance status, comorbidities, treatment efficacy, and adverse event profile affect first-line prescribing. All panelists agreed it is important to use the most efficacious treatment first; 88% of panelists considered daratumumab-containing regimens the most efficacious. Panelists agreed treatment should be continued until progression while benefits outweigh risk. Conclusion: Findings reinforce the importance of using the most efficacious regimen upfront for TIE NDMM, and nearly all panelists considered daratumumab-containing regimens the most efficacious treatment.

Background: Daratumumab, lenalidomide and dexamethasone (DRd) and bortezomib, lenalidomide and dexamethasone (VRd) are preferred regimens for transplant ineligible (TIE) patients with newly diagnosed multiple myeloma (NDMM). Both DRd and VRd demonstrated superior efficacy versus Rd in the MAIA and SWOG S0777 trials, respectively, but there is no head-to-head (H2H) clinical trial comparing their efficacy. Differing populations in the MAIA and S0777 trials make an unadjusted comparison of outcomes challenging and biased. The current TAURUS study is the first real-world H2H study comparing progression-free survival (PFS) among TIE NDMM patients treated with DRd or VRd as first-line (1L) in similar clinical settings., Materials and Methods: A multicenter chart review study was conducted at nine sites across the United States. All TIE patients treated with DRd and a randomly selected population of VRd patients were included. TIE NDMM patients aged ≥65 were included if they initiated 1L DRd/VRd between January 2019 and September 2021. PFS was defined as the time from DRd/VRd initiation until disease progression or death. A doubly-robust multivariable Cox regression model combined with inverse probability of treatment weighting (IPTW) methodology was used to compare PFS between cohorts., Results: Weighted cohorts comprised 91 DRd and 87 VRd patients. Thirteen DRd and 24 VRd patients experienced progression/death. Patients treated with DRd had a lower risk of progression/death versus VRd (adjusted hazard ratio: 0.35, 95% confidence interval: [0.17; 0.73])., Conclusion: DRd is associated with a significantly lower risk of disease progression or death compared to VRd as 1L treatment for TIE NDMM patients., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Daratumumab (DARA) is an anti-CD38 monoclonal antibody approved as a combination therapy for newly diagnosed multiple myeloma (MM) and as monotherapy and combination therapy for relapsed or refractory MM cases. We assessed the length of DARA use across lines of therapy and the probabilities of treatment discontinuation in patients with MM in the real-world. We used the deidentified Clinformatics Data Mart database from Optum to identify patients with MM (n=2124) who received DARA-containing treatment between November 1, 2015 and March 31, 2021 in the United States. Patients were excluded if they had received a stem cell transplant. The duration of DARA use was defined as the time interval between the first initiation and discontinuation of DARA as a time-to-event outcome using the Kaplan-Meier method. A gap of more than 60 days between 2 consequent DARA claim dates was defined as DARA discontinuation. The median duration of continuous DARA use was 16.6 months. By 24 months, 33.1% of patients remained on DARA treatment. In a subgroup analysis of patients with 12 months or more continuous insurance coverage (n=1246), the median length of DARA use was 24.7 months; by 24 months, 51.8% remained on DARA treatment. The dose adherence ratios (observed DARA doses relative to the label) were close to 1.0, particularly among patients with longer follow-up, indicating that real-world DARA dosing frequency was similar to that on the approved label. In summary, this real-world analysis reported that the median duration of continuous DARA use is 16.6 months, with high dosing adherence in patients who have MM., Competing Interests: R.F. served as a consultant for AbbVie, Aduro, Amgen, Bayer, Bristol Myers Squibb, Celgene, GSK, Janssen, Juno, Kite Pharma, Merck, Novartis, ONCOtracker, Pharmacyclics, Sanofi, and Takeda; participated in scientific advisory boards for Adaptive Biotechnologies and ONCOtracker, Board of Directors Antegene. E.E.C., N.G-W., A.Z.F., & S.K. are employees of the Janssen Pharmaceutical Companies of Johnson & Johnson. All other authors declare no competing interests., (© 2023 The Authors.)

Purpose: Median duration of daratumumab (DARA) administration for treatment of multiple myeloma is 3-7 hours for the intravenous formulation (DARA IV) and 3-5 minutes for the subcutaneous formulation (DARA SC). Here, we describe clinical administration characteristics of DARA using a novel method for data extraction from electronic health records., Methods: Time-based measurements were extracted using a scheduling/pharmacy software program that tracked patient movement through appointments for patients initiating DARA in Mayo Clinic infusion centers from April 5, 2017, to October 14, 2021. Cohorts included patients who received DARA IV or DARA SC, or converted from DARA IV to DARA SC. The DARA SC cohort was further analyzed before (DARA SC initial) and after (DARA SC shortened) a reduction in the postadministration observation time mandated by the treatment plan. Events associated with administration-related reactions (ARRs) were also identified., Results: Median total clinic times were 2.7-3.0 hours shorter for DARA SC versus DARA IV. Median clinic times were highest at dose 1 and decreased with subsequent doses. Median total chair times were 2.7-2.8 hours shorter for DARA SC versus DARA IV. Incidences of ARR-related events with DARA SC were low across doses., Conclusion: Reduced clinic times were observed with DARA SC, indicating that use of DARA SC as a treatment option results in time savings that may free clinic resources. Furthermore, novel methods of electronic health record data extraction can provide insights that may help inform clinic resource optimization.

Aim: To evaluate among multiple myeloma (MM) patients, the proportions with first-line bortezomib/lenalidomide/dexamethasone (VRd) dose modifications and the associated baseline patient characteristics. Patients & methods: Adult MM patients treated with first-line VRd were selected from the Optum claims database. VRd dose modifications were defined based on lenalidomide dose. Results: Among 1497 MM patients, 33% received VRd lite and 22% VRd reduced. Compared with VRd regular, VRd lite usage was more likely to be associated with patients aged ≥75 years and female sex; VRd reduced usage was more likely to be associated with female sex and frailty. Conclusion: A large proportion of MM patients received VRd dose modifications in the real-world, which could potentially result in reduced effectiveness of VRd.

Background: There is limited real-world evidence that describes patients with newly diagnosed multiple myeloma (NDMM) treated with the bortezomib, lenalidomide, and dexamethasone (VRd) triplet regimen. We evaluated patient characteristics and treatment outcomes among nontransplanted NDMM patients who received VRd as their first line of therapy (LOT) in US oncology practice settings., Methods: This retrospective observational cohort study evaluated patients from the Flatiron MM Core Registry who received VRd as first LOT between November 1, 2015, and February 28, 2021. Progression-free survival (PFS) was analyzed using the Kaplan-Meier method. Associations between patient demographic and clinical characteristics and PFS were evaluated using a multivariable Cox proportional hazards model., Results: A total of 2342 eligible patients with VRd as first LOT were identified (mean age, 67.0 years). Among all identified patients, 64.3% were ≥ 65 years of age, 25.5% were elderly (≥75 years), and 47.9% were frail. Among patients with available data, 21.2% had high-risk cytogenetics, and the majority had International Staging System (ISS) stage I/II disease (71.8%), and Eastern Cooperative Oncology Group performance status (ECOG PS) score 0/1 (81.2%). Median duration of therapy was 5.5 months. With median follow-up of 21.0 months, median PFS and time-to-next-treatment were 26.5 and 16.1 months, respectively. Higher risk of disease progression or death was seen in patients categorized as elderly (hazard ratio [HR] = 1.37; 95% confidence interval [CI]: 1.13-1.66 vs patients < 65 years), having high-risk cytogenetics (HR = 1.44; 95% CI: 1.19-1.75 vs standard risk), having ISS disease stages II and III (HR = 1.31; 95% CI: 1.06-1.63 and HR = 1.37; 95% CI: 1.10-1.70 versus stage I, respectively), and having worse ECOG PS score (≥2) (HR = 1.49; 95% CI: 1.22-1.81 versus functionally active patients)., Conclusions: The majority of patients treated with VRd in this study were ≥ 65 years of age, were ISS stage I/II, had an ECOG PS score of 0/1, and had standard cytogenetic risk. Median PFS observed in real-world practice was notably shorter than that observed in the SWOG S0777 clinical trial. In nontransplanted patients treated with VRd as first LOT, a higher risk of disease progression or death was associated with older age, having high-risk cytogenetics, worse disease stage, and worse ECOG PS score., (© 2022. The Author(s).)

Background: Daratumumab was approved for multiple myeloma (MM) in 2015. While its safety and efficacy are well documented, there is limited real-world information on its use and outcomes in patients of different races., Methods: We conducted a retrospective chart review of adult patients with MM initiating daratumumab in any line of therapy (LOT) between November 2015 and May 2020. De-identified data were retrieved from 2 US clinical sites; patient characteristics, treatment patterns, and response rate were described for black and white patients, stratified by LOT. Overall response rate (ORR), progression-free survival (PFS), and time to next LOT (TTNT) were compared between black and white patients initiating daratumumab in second line (2L) or later, adjusting for age and number of prior lines., Results: Two hundred and fifty-two patient charts (89 black, 163 white) were extracted. Black patients were younger at diagnosis (61.7 vs. 67.0 years) and had a similar proportion of females (black: 44.9%, white: 46.6%). Black patients had longer time between MM diagnosis and daratumumab initiation (43.2 vs. 34.1 months) and received more prior LOTs (median 3.0 vs. 2.0). ORR for black and white patients initiating daratumumab in 1L was 100.0%, with very good partial response or better in 75.0% and 66.7%, respectively. Similar trends were observed in 2L and 3L+. There were no significant differences in ORR, PFS, or TTNT between groups., Conclusion: Daratumumab had similar clinical outcomes (ORR, PFS, and TTNT) in black and white patients. Black patients initiated daratumumab later in their treatment, suggesting potential discrepancies in access to new MM treatments., (Copyright © 2022. Published by Elsevier Inc.)

Background: Patients with high-risk, newly diagnosed multiple myeloma (HR-NDMM) who are ineligible for autologous stem cell transplant (ASCT) have limited first-line treatment options. Recent meta-analyses evaluating the impact of incorporating daratumumab in the backbone regimen on progression-free survival (PFS) have found mixed results in these patients., Materials and Methods: A pooled analysis of patient-level data for ASCT-ineligible patients with HR-NDMM [ie, del(17p), t(4;14), t(14;16)] from the MAIA and ALCYONE trials; stratified by study identifier and adjusting for cytogenetic abnormality subtype, baseline performance status, International Staging System stage, myeloma type, and renal impairment; was conducted. Impact of daratumumab on PFS and rates of complete response or better (≥CR), minimal residual disease (MRD)-negative CR, very good partial response or better (≥VGPR), and overall response (ORR) was compared to control., Results: Among 101 patients in the daratumumab and 89 patients in the control cohort, median follow-up was 43.7 months. Daratumumab reduced the risk of progression or death by 41% (adjusted hazard ratio for PFS [95% confidence interval (CI)] = 0.59 [0.41-0.85]) versus control. At 36 months, the estimated proportion of patients who did not progress and were still alive was 41.3% in the daratumumab and 19.9% in the control cohort. Rates of ≥CR (41.6% vs. 22.5%), MRD-negative CR (24.8% vs. 5.6%), ≥VGPR (75.2% vs. 46.1%), and ORR (92.1% vs. 74.2%) were higher for daratumumab versus control., Conclusion: These findings demonstrate that incorporation of daratumumab in frontline treatment regimens reduced the risk of progression or death and improved response rates among ASCT-ineligible HR-NDMM patients., (© The Author(s) 2022. Published by Oxford University Press.)

Kuhns was the first to suggest that theories in science do not develop in small increments but rather in major leaps to paradigms that examine the same question through very different perspectives. Theories on the mechanism responsible for control of human food intake fall into Kuhn's description. This article describes how the two major theories of the control of food intake in humans, the Glucostatic Theory, and the Lipostatic Theory, showed initial promise as explanations, but later deteriorated with the slow accumulation experimental data. The locus of theories considered eating behavior as a part of physiological system that regulates the storage of energy on the body. We challenge this fundamental belief with data which suggests that we must be ready to accept a major change in the way we think about eating behavior if we are ever to decrease the prevalence of obesity., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Levitsky, Barre, Michael, Zhong, He, Mizia and Kaila.)

Background: Daratumumab, a CD38 monoclonal antibody, has demonstrated efficacy as monotherapy and combination therapy across several indications, both among newly-diagnosed and refractory patients with multiple myeloma (MM). However, there is limited evidence on treatment patterns and effectiveness of daratumumab in the real-world setting, particularly in first line (1 L). This study aimed to describe real-world treatment patterns and clinical outcomes among patients initiating daratumumab across different lines of therapy., Methods: A retrospective chart review of adult patients with MM initiating daratumumab between November 2015 and March 2021 was conducted at two clinical sites in the United States. De-identified patient-level data were abstracted in an electronic case report form. Patient characteristics and treatment patterns were described. Clinical outcomes including overall response rate (ORR), progression-free survival, and time to next line of therapy were reported using descriptive statistics and stratified by line of therapy (1 L, second line [2 L] or third line or later [3 L+]). A sub-group analysis evaluated treatment patterns and ORR among patients re-treated with daratumumab., Results: A total of 299 patients were included in the study (mean age: 68 years; 55% male). Among them, 26 were 1 L patients, 66 were 2 L patients, and 207 were 3 L+ patients; 110 patients (36.8%) received a stem cell transplant prior to daratumumab initiation. The mean duration of follow-up was 10 months among 1 L patients and 19 months among 2 L and 3 L+ patients. Patients who initiated daratumumab in 1 L had a 100% ORR, while those initiating in 2 L and 3 L+ had an ORR of 78.8 and 65.2%, respectively. Among re-treated patients, ORR was 66.7% during the first treatment segment, and 52.9% during the second treatment segment. Kaplan-Meier rates of progression-free survival at 12 months were 89.9, 75.2, and 53.1% among patients who initiated daratumumab in 1 L, 2 L, and 3 L+, respectively. Kaplan-Meier rates of time to next line of therapy at 12 months were 94.1, 73.4, and 50.0% among patients who initiated daratumumab in 1 L, 2 L, and 3 L+, respectively., Conclusions: These findings suggest that daratumumab-based regimens are an effective treatment option across all lines of therapy, with highest response rate in 1 L., (© 2021. The Author(s).)

Introduction: In 2015, Boehringer Ingelheim (BI) created a support program for patients with idiopathic pulmonary fibrosis (IPF) treated with nintedanib, to help patients obtain their prescription, learn about their disease and medication, and provide support in the management of their IPF. The purpose of this study was to measure the impact of the program on nintedanib persistence among patients with IPF newly treated with the medication., Methods: A retrospective cohort analysis of BI Pharmaceuticals, Inc.'s Specialty Pharmacy (SP) database was conducted. Patients at least 18 years of age, newly treated with nintedanib from April 1, 2015 to January 31, 2018, and with at least one diagnosis of IPF were included in the study; earliest nintedanib prescription was the index date. Patients were classified into two mutually exclusive cohorts: enrolled in the patient support program within 60 days of index or not enrolled in the program at any time. The cohorts were compared in terms of patient characteristics, time to nintedanib discontinuation (a gap of more than 60 days between refills), and proportion of persistent patients at 6, 12, 18, and 24 months after index. Time to discontinuation was compared between the cohorts using Kaplan-Meier analysis. A multivariable Cox proportional hazards model assessed the impact of program participation on time to discontinuation within the first 12 months., Results: A total of 3114 enrolled and 9388 non-enrolled patients were identified. The proportion of patients persistent on nintedanib was higher among enrolled patients throughout the post-index period (57.8% vs. 49.7% at 6 months, 34.7% vs. 28.9% at 12 months; p < 0.05). In adjusted analyses, being enrolled in the program was associated with a 21% decreased hazard of discontinuing nintedanib over the first-year post-index [hazard ratio (HR) = 0.79, 95% CI 0.75-0.83, p < 0.05)., Conclusion: Real-world evidence suggests a persistence benefit for patients with IPF treated with nintedanib who are enrolled in the patient support program., (© 2021. The Author(s).)

Climate change during the last 40 years has had a serious impact on agriculture and threatens global food and nutritional security. From over half a million plant species, cereals and legumes are the most important for food and nutritional security. Although systematic plant breeding has a relatively short history, conventional breeding coupled with advances in technology and crop management strategies has increased crop yields by 56 % globally between 1965-85, referred to as the Green Revolution. Nevertheless, increased demand for food, feed, fiber, and fuel necessitates the need to break existing yield barriers in many crop plants. In the first decade of the 21st century we witnessed rapid discovery, transformative technological development and declining costs of genomics technologies. In the second decade, the field turned towards making sense of the vast amount of genomic information and subsequently moved towards accurately predicting gene-to-phenotype associations and tailoring plants for climate resilience and global food security. In this review we focus on genomic resources, genome and germplasm sequencing, sequencing-based trait mapping, and genomics-assisted breeding approaches aimed at developing biotic stress resistant, abiotic stress tolerant and high nutrition varieties in six major cereals (rice, maize, wheat, barley, sorghum and pearl millet), and six major legumes (soybean, groundnut, cowpea, common bean, chickpea and pigeonpea). We further provide a perspective and way forward to use genomic breeding approaches including marker-assisted selection, marker-assisted backcrossing, haplotype based breeding and genomic prediction approaches coupled with machine learning and artificial intelligence, to speed breeding approaches. The overall goal is to accelerate genetic gains and deliver climate resilient and high nutrition crop varieties for sustainable agriculture., (Copyright © 2020 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Ferroportin (Fpn/IREG1/MTP1) is the only known transporter mediating iron efflux from epithelial cells and macrophages, and thus regulates how much iron is released into the circulation. Consequently, Fpn mutations are associated with haemochromatosis. Fpn itself is post-translationally regulated by hepcidin (Hepc) which induces its redistribution and degradation in a ubiquitin-dependent process. Together, the two proteins appear to be the nexus for iron homeostasis. Here we show that a rare gain-of-function mutation (K240E) that is associated with iron overload, impedes Fpn binding and subcellular trafficking by the small ubiquitin-like modifier (SUMO). Whereas wild-type Fpn is ensconced within vesicular bodies, the FpnK240E mutant appeared diffused within the cell when co-expressed with SUMO. Furthermore, compared with wild type Fpn, the sumoylation-defective mutant was constitutively-active, resulting in a lower intracellular labile iron pool than the former. These findings suggest that SUMO may regulate iron homeostasis by controlling Fpn trafficking., Competing Interests: The authors have no competing interests., (© 2020 Published by Elsevier B.V.)

Objectives: Triple therapy is indicated for patients with very severe chronic obstructive pulmonary disease (COPD). Use of this treatment in the appropriate patient population is important to ensure optimal outcomes. This study quantified the use of triple therapy and assessed concordance with 2013-2016 Global Initiative for Chronic Obstructive Lung Disease (GOLD) recommendations within a national health plan., Study Design: Retrospective cohort study using data from a large national health plan., Methods: To estimate the prevalence of triple therapy using claims data, patients in the first of 2 cohorts were indexed on their first diagnosis of COPD between January 1, 2012, and December 31, 2014, and required to have 24 months postindex continuous enrollment. To assess concordance with GOLD recommendations, a second cohort was created and indexed on the date of triple therapy initiation between January 1, 2013, and November 30, 2016, and required to have 12 months preindex and 1 month postindex continuous enrollment. For both cohorts, patients were aged 40 years or older, with no International Classification of Diseases code for asthma, cystic fibrosis, or lung cancer during the study period., Results: In the first cohort of 92,248 patients with COPD receiving any COPD maintenance medication, 17% were prescribed triple therapy. In the second cohort (n = 19,645), the majority (60%) of patients on triple therapy were classified as GOLD group A or B (ie, no evidence of any exacerbation or only 1 exacerbation not resulting in hospitalization at baseline)., Conclusions: Results showed that triple therapy was often prescribed among patients classified as GOLD group A or B. Additional research is required, however, to further assess whether these patients may have had an exacerbation that was not evident in claims data. Treatment of COPD should be individualized to optimize outcomes and reduce adverse events.

Aim: To compare health plan-paid costs, exacerbations and pneumonia outcomes for patients with chronic obstructive pulmonary disease (COPD) initiating combination tiotropium olodaterol (TIO + OLO) versus triple therapy (TT: long-acting muscarinic antagonist + long-acting β 2 agonists + inhaled corticosteroid). Patients & methods: COPD patients initiating TIO + OLO or TT between 1 January 2014 and 30 June 2016 were identified from a managed care Medicare database and balanced for baseline characteristics using inverse probability of treatment weighting before assessment of outcomes. Results: Annual COPD-related and all-cause costs were US$4118 (35%) and US$5384 (23%) lower for TIO + OLO versus TT (both p ≤ 0.001). TIO + OLO patients had nearly half the severe exacerbations (8.3 vs 15.5%; p = 0.014) and pneumonia was also less common (18.9 vs 30.9%; p < 0.001). Conclusion: TIO + OLO was associated with improved economic and COPD health outcomes versus TT.

Purpose: There is general agreement that some dietary polyphenols block non-haem iron uptake, but the mechanisms by which they achieve this action are poorly understood. Since the polyphenol quercetin is ingested daily in significant amounts, we have investigated the effect of quercetin on duodenal non-haem iron absorption in vivo, as well as its effect on factors known to be involved in systemic iron metabolism., Methods: Rats were subject to gastric gavage and systemic quercetin administration. Treatments were followed with uptake studies using radiolabeled iron, serum iron and transferrin saturation measurements, LC-MS/MS analysis of quercetin metabolites in serum, determination of tissue non-haem iron content and analysis of gene expression of iron-related proteins., Results: Both oral and intraperitoneal (IP) quercetin caused serum and tissue iron depletion by two means, first by increasing mucosal iron uptake and inhibiting iron efflux from duodenal mucosa, and second by decreasing levels of duodenal DMT1, Dcytb and FPN. Additionally, IP quercetin induced highly significant increased liver expression of hepcidin, a hormone known to inhibit intestinal iron uptake., Conclusions: Oral quercetin significantly inhibited iron absorption, while IP quercetin significantly affected iron-related genes. These results could lead to development of new effective ways of preventing and treating iron deficiency anaemia, the most widespread nutritional disorder in the world.

Background: A claims-based model to predict patients likely to have undiagnosed COPD was developed by Moretz et al in 2015. This study aims to assess the performance of the aforementioned model using prospectively collected spirometry data., Methods: A study population aged 40-89 years enrolled in a Medicare Advantage plan with prescription drug coverage or commercial health plan and without a claim for COPD diagnosis was identified from April 1, 2012 to March 31, 2016 in the Humana claims database. This population was stratified into subjects likely or unlikely to have undiagnosed COPD using the claims-based predictive model. Subjects were randomly selected for spirometry evaluation of FEV 1 and FVC. The predictive model was validated using airflow limitation ratio (FEV 1 /FVC <0.70)., Results: A total of 218 subjects classified by the predictive model as likely and 331 not likely to have undiagnosed COPD completed spirometry evaluation. Those predicted to have undiagnosed COPD had a higher mean age (70.2 vs 67.9 years, P =0.0012) and a lower mean FEV 1 /FVC ratio (0.724 vs 0.753, P =0.0002) compared to those predicted not to have undiagnosed COPD. Performance metrics for the predictive model were: area under the curve =0.61, sensitivity =52.5%, specificity =64.6%, positive predictive value =33.5%, and negative predictive value =80.1%., Conclusion: The claims-based predictive model identifies those not at risk of having COPD eight out of ten times, and those who are likely to have COPD one out of three times., Competing Interests: Disclosure Rakesh Luthra, Asif Shaikh, and Shuchita Kaila are employees of Boehringer Ingelheim. Chad Moretz, Srinivas Annavarapu, and Seth Goldfarb are employees of Comprehensive Health Insights, Inc., which conducted the study. Andrew Renda is an employee of Humana Inc., and provided project consultation. The authors report no other conflicts of interest in this work.

Background: Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality and is associated with substantial economic burden. There is a lack of data regarding COPD outcomes and costs in a real-world setting, particularly by Global Initiative for Chronic Obstructive Lung Disease (GOLD) severity., Objectives: To (a) characterize a commercially insured U.S. population with COPD and (b) assess prevalence of exacerbations, health care resource utilization (HCRU), costs, and treatment patterns in a cohort of patients with confirmed COPD, overall and stratified by GOLD stage., Methods: This retrospective observational cohort study used administrative claims data from the HealthCore Integrated Research Database to identify patients with ≥ 1 inpatient, emergency room (ER), or office visit claim for COPD between January 1, 2012, and November 30, 2013, and continuous enrollment for 1 year before and 2 years after the first COPD diagnosis date. Patients with a spirometry claim within 12 months were eligible for medical record abstraction to confirm COPD diagnosis (forced expiratory volume in 1 second [FEV 1 ]/forced vital capacity ratio < 0.7) and GOLD 1-4 classification (based on postbronchodilator FEV 1 percent predicted). HCRU, costs, treatment patterns, and rate of moderate/severe exacerbation were identified from diagnosis up to 24 months. Outcomes were analyzed by univariate analysis stratified by GOLD classification. Multivariable analysis was conducted to assess associations between GOLD classification and outcomes of interest., Results: 53,484 patients newly diagnosed with COPD were identified who met initial inclusion criteria: 14,293 (27%) had a qualifying spirometry claim, and 1,505 had confirmed COPD (GOLD 1, 333 [22%]; GOLD 2, 823 [55%]; GOLD 3, 317 [21%]; GOLD 4, 32 [2%]). Patients with greater disease severity had higher rates of moderate/severe COPD exacerbations (GOLD 1 and 2, 40.4 and 48.9 per 100 person-years, respectively; GOLD 3 and 4, 83.6 and 89.1 per 100 person-years, respectively). All-cause and COPD-related inpatient admissions, COPD-related office visits, and COPD-related ER visits were more prevalent with more severe GOLD classification. Mean annual COPD-related medical costs increased with GOLD classification ($5,945 for GOLD 1 patients, $18,070 for GOLD 4). COPD maintenance medication was filled by 42%, 56%, 73%, and 75% of patients in GOLD 1-4 (57% overall), respectively; combination corticosteroid/long-acting beta2-agonist inhalers were the most commonly used medication, regardless of GOLD classification. Patients with more severe disease had greater adherence (range 44%-68% of days covered for GOLD 1-4) and persistence (range 107-209 days for GOLD 1-4)., Conclusions: Trends toward increases in exacerbations, HCRU, and costs were observed as airflow limitation worsened. Adherence and persistence with COPD maintenance therapy was suboptimal even with severe disease., Disclosures: This study was supported by Boehringer Ingelheim Pharmaceuticals (Ridgefield, CT), which was given the opportunity to review the manuscript for medical and scientific accuracy, as well as intellectual property considerations. Willey and Singer are employees of HealthCore (parent company Anthem), which received funding from Boehringer Ingelheim to complete this study. Wallace and Shinde were employed by HealthCore at the time of this study. Wallace and Singer report stock ownership in Anthem. Napier is an employee of Anthem. Kaila, Bayer, and Shaikh are employees of Boehringer Ingelheim Pharmaceuticsls. Portions of this research were presented at the following conferences: (a) A. Wallace, S. Kaila, V. Zubek, A. Shaikh, M. Shinde, V. Willey, M. Napier, and J. Singer, Healthcare resource utilization, costs, and exacerbation rates in patients with COPD stratified by GOLD airflow limitation classification in a US commercially insured population, presented at AMCP Nexus 2017; October 16-19, 2017; Dallas, TX; and (b) A.E. Wallace, V. Zubek, S. Kaila, A. Shaikh, M. Shinde, V. Willey, M.B. Napier, and J.R. Singer, Real-world treatment patterns among newly diagnosed COPD patients according to GOLD airflow limitation severity classification in a U.S. commercially insured/Medicare Advantage population, presented at CHEST 2017 Annual Meeting; October 28-November 1, 2017; Toronto, Ontario, Canada.

Background: Patients with COPD often experience severe exacerbations involving hospitalization, which accelerate lung function decline and reduce quality of life. This study aimed to develop and validate a predictive model to identify patients at risk of developing severe COPD exacerbations using administrative claims data, to facilitate appropriate disease management programs., Methods: A predictive model was developed using a retrospective cohort of COPD patients aged 55-89 years identified between July 1, 2010 and June 30, 2013 using Humana's claims data. The baseline period was 12 months postdiagnosis, and the prediction period covered months 12-24. Patients with and without severe exacerbations in the prediction period were compared to identify characteristics associated with severe COPD exacerbations. Models were developed using stepwise logistic regression, and a final model was chosen to optimize sensitivity, specificity, positive predictive value (PPV), and negative PV (NPV)., Results: Of 45,722 patients, 5,317 had severe exacerbations in the prediction period. Patients with severe exacerbations had significantly higher comorbidity burden, use of respiratory medications, and tobacco-cessation counseling compared to those without severe exacerbations in the baseline period. The predictive model included 29 variables that were significantly associated with severe exacerbations. The strongest predictors were prior severe exacerbations and higher Deyo-Charlson comorbidity score (OR 1.50 and 1.47, respectively). The best-performing predictive model had an area under the curve of 0.77. A receiver operating characteristic cutoff of 0.4 was chosen to optimize PPV, and the model had sensitivity of 17%, specificity of 98%, PPV of 48%, and NPV of 90%., Conclusion: This study found that of every two patients identified by the predictive model to be at risk of severe exacerbation, one patient may have a severe exacerbation. Once at-risk patients are identified, appropriate maintenance medication, implementation of disease-management programs, and education may prevent future exacerbations., Competing Interests: Disclosure MG and SK are employees of Boehringer Ingelheim. SA and SG are employees of Comprehensive Health Insights, which conducted the study. CM was an employee of Comprehensive Health Insights at the time of this study and is now employed by GlaxoSmithKline. AR is an employee of Humana and provided project consultation. The authors report no other conflicts of interest in this work.

Background: Patients with chronic obstructive pulmonary disease (COPD) often have multiple underlying comorbidities, which may lead to increased health care resource utilization (HCRU) and costs., Objective: To describe the comorbidity profiles of COPD patients and examine the associations between the presence of comorbidities and HCRU or health care costs., Methods: A retrospective cohort study utilizing data from a large US national health plan with a predominantly Medicare population was conducted. COPD patients aged 40-89 years and continuously enrolled for 12 months prior to and 24 months after the first COPD diagnosis during the period of January 01, 2009, through December 31, 2010, were selected. Eleven comorbidities of interest were identified 12 months prior through 12 months after COPD diagnosis. All-cause and COPD-related hospitalizations and costs were assessed 24 months after diagnosis, and the associations with comorbidities were determined using multivariate statistical models., Results: Ninety-two percent of 52,643 COPD patients identified had at least one of the 11 comorbidities. Congestive heart failure (CHF), coronary artery disease, and cerebrovascular disease (CVA) had the strongest associations with all-cause hospitalizations (mean ratio: 1.56, 1.32, and 1.30, respectively; P <0.0001); other comorbidities examined had moderate associations. CHF, anxiety, and sleep apnea had the strongest associations with COPD-related hospitalizations (mean ratio: 2.01, 1.32, and 1.21, respectively; P <0.0001); other comorbidities examined (except chronic kidney disease [CKD], obesity, and osteoarthritis) had moderate associations. All comorbidities assessed (except obesity and CKD) were associated with higher all-cause costs (mean ratio range: 1.07-1.54, P <0.0001). CHF, sleep apnea, anxiety, and osteoporosis were associated with higher COPD-related costs (mean ratio range: 1.08-1.67, P <0.0001), while CVA, CKD, obesity, osteoarthritis, and type 2 diabetes were associated with lower COPD-related costs., Conclusion: This study confirms that specific comorbidities among COPD patients add significant burden with higher HCRU and costs compared to patients without these comorbidities. Payers may use this information to develop tailored therapeutic interventions for improved management of patients with specific comorbidities., Competing Interests: Disclosure This study was funded by Humana Inc. and Boehringer Ingelheim Pharmaceuticals Inc. Andrew Renda is an employee and shareholder of Humana Inc.; Amol D Dhamane, Kate Burslem, and Shuchita Kaila are employees of Boehringer Ingelheim Pharmaceuticals Inc.; Phil Schwab, Sari D Hopson, Chad Moretz, and Srinivas Annavarapu are employees of Comprehensive Health Insights, Inc., a wholly-owned subsidiary of Humana Inc., who were paid consultants to Boehringer Ingelheim Pharmaceuticals Inc. in connection with the analysis described in this study and with manuscript development. The authors report no other conflicts of interest in this work.

Background: Patients with COPD often have multiple comorbidities requiring use of multiple medications, and adherence rates for maintenance COPD (mCOPD) medications are already known to be suboptimal. Presence of comorbidities in COPD patients, and use of medications used to treat those comorbidities (non-COPD medications), may have an adverse impact on adherence to mCOPD medications., Objective: The objective of the study was to evaluate the association between non-adherence to mCOPD medications and non-COPD medications in COPD patients., Methods: COPD patients were identified using a large administrative claims database. Selected patients were 40-89 years old and continuously enrolled for 12 months prior to and 24 months after the first identified COPD diagnosis (index date) during January 1, 2009 to December 31, 2010. Patients were required to have ≥1 prescription for a mCOPD medication within 365 days of the index date and ≥1 prescription for one of 12 non-COPD medication classes within ±30 days of the first COPD prescription. Adherence (proportion of days covered [PDC]) was measured during 365 days following the first COPD prescription. The association between non-adherence (PDC <0.8) to mCOPD and non-adherence to non-COPD medications was determined using logistic regression, controlling for baseline patient characteristics., Results: A total of 14,117 patients, with a mean age of 69.9 years, met study criteria. Of these, 40.9% were males and 79.2% were non-adherent to mCOPD medications with a mean PDC of 0.47. Non-adherence to mCOPD medications was associated with non-adherence to 10 of 12 non-COPD medication classes (odds ratio 1.38-1.78, all P <0.01)., Conclusion: Adherence to mCOPD medications is low. Non-adherence (or adherence) to mCOPD medications is positively related to non-adherence (or adherence) to non-COPD medications, implying that the need to take medications prescribed for comorbid conditions does not adversely impact adherence to mCOPD medications., Competing Interests: The authors report no conflicts of interest in this work.

Background: The Global Initiative for Chronic Obstructive Lung Disease (GOLD) recommends triple therapy (long-acting muscarinic receptor antagonists, long-acting beta-2 agonists, and inhaled corticosteroids) for patients with only the most severe COPD. Data on the proportion of COPD patients on triple therapy and their characteristics are sparse and dated. Objective 1 of this study was to estimate the proportion of all, and all treated, COPD patients receiving triple therapy. Objective 2 was to characterize those on triple therapy and assess the concordance of triple therapy use with GOLD guidelines., Patients and Methods: This retrospective study used claims from the IMS PharMetrics Plus database from 2009 to 2013. Cohort 1 was selected to assess Objective 1 only; descriptive analyses were conducted in Cohort 2 to answer Objective 2. A validated claims-based algorithm and severity and frequency of exacerbations were used as proxies for COPD severity., Results: Of all 199,678 patients with COPD in Cohort 1, 7.5% received triple therapy after diagnosis, and 25.5% of all treated patients received triple therapy. In Cohort 2, 30,493 COPD patients (mean age =64.7 years) who initiated triple therapy were identified. Using the claims-based algorithm, 34.5% of Cohort 2 patients were classified as having mild disease (GOLD 1), 40.8% moderate (GOLD 2), 22.5% severe (GOLD 3), and 2.3% very severe (GOLD 4). Using exacerbation severity and frequency, 60.6% of patients were classified as GOLD 1/2 and 39.4% as GOLD 3/4., Conclusion: In this large US claims database study, one-quarter of all treated COPD patients received triple therapy. Although triple therapy is recommended for the most severe COPD patients, spirometry is infrequently assessed, and a majority of the patients who receive triple therapy may have only mild/moderate disease. Any potential overprescribing of triple therapy may lead to unnecessary costs to the patient and health care system., Competing Interests: Boehringer Ingelheim, Inc. provided the funding for this study. RL and SK are salaried employees of Boehringer Ingelheim. JCS, TDB, JL, and TKW are currently employees of Evidera, which provides consulting and other research services to pharmaceutical, device, and other organizations. In their salaried positions, they work with a variety of companies and organizations and are precluded from receiving payment or honoraria directly from these organizations for services rendered. Evidera received funding from Boehringer Ingelheim for work on the project and the manuscript. XP was an employee of Evidera during the conduct of this study and the writing of this manuscript; she is currently employed by Sunovion Pharmaceuticals, Marlborough, MA. The authors report no other conflicts of interest in this work.

The liver expresses batteries of cytoprotective genes that confer cellular resistance to oxidative stress and xenobiotic toxins, and protection against cancer and other stress-related diseases. These genes are mainly regulated by Nrf2, making this transcription factor a target for small molecule discovery to treat such diseases. In this report, we identified dietary polyphenolic antioxidants that not only activated these genes but also relieved Nrf2 repression by Keap1, a Cul3-dependent ubiquitin ligase adaptor protein that mediates its degradation. Analysis of postprandial liver RNA revealed a marked activation of both genes by all test polyphenols compared with controls. Nrf2 inhibition by RNA interference reduced polyphenol effects on its target gene expression. Our data suggest that polyphenols may induce cellular defense genes by derepressing Nrf2 inhibition by Keap1. We posit that this ability to derepress Nrf2 and reactivate its target genes may underlie the protection conferred by polyphenols against oxidative stress-related diseases., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Chronic obstructive pulmonary disease (COPD) exacerbations account for a substantial proportion of COPD-related costs., Objective: To describe COPD exacerbation patterns and assess the association between exacerbation frequency and health care resource utilization (HCRU) and costs in patients with COPD in a Medicare population., Methods: A retrospective cohort study utilizing data from a large US national health plan was conducted including patients with a COPD diagnosis during January 1, 2007 to December 31, 2012, aged 40-89 years and continuously enrolled in a Medicare Advantage Prescription Drug plan. Exacerbation frequency, HCRU, and costs were assessed during a 24-month period following the first COPD diagnosis (follow-up period). Four cohorts were created based on exacerbation frequency (zero, one, two, and ≥three). HCRU and costs were compared among the four cohorts using chi-square tests and analysis of variance, respectively. A trend analysis was performed to assess the association between exacerbation frequency and costs using generalized linear models., Results: Of the included 52,459 patients, 44.3% had at least one exacerbation; 26.3%, 9.5%, and 8.5% had one, two, and ≥three exacerbations in the 24-month follow-up period, respectively. HCRU was significantly different among cohorts (all P<0.001). In patients with zero, one, two, and ≥three exacerbations, the percentages of patients experiencing all-cause hospitalizations were 49.7%, 66.4%, 69.7%, and 77.8%, respectively, and those experiencing COPD-related hospitalizations were 0%, 40.4%, 48.1%, and 60.5%, respectively. Mean all-cause total costs (medical and pharmacy) were more than twofold greater in patients with ≥three exacerbations compared to patients with zero exacerbations ($27,133 vs $56,033; P<0.001), whereas a greater than sevenfold difference was observed in mean COPD-related total costs ($1,605 vs $12,257; P<0.001)., Conclusion: COPD patients frequently experience exacerbations. Increasing exacerbation frequency is associated with a multiplicative increase in all-cause and COPD-related costs. This underscores the importance of identifying COPD patients at risk of having frequent exacerbations for appropriate disease management.

Background: Despite the importance of early detection, delayed diagnosis of chronic obstructive pulmonary disease (COPD) is relatively common. Approximately 12 million people in the United States have undiagnosed COPD. Diagnosis of COPD is essential for the timely implementation of interventions, such as smoking cessation programs, drug therapies, and pulmonary rehabilitation, which are aimed at improving outcomes and slowing disease progression., Objective: To develop and validate a predictive model to identify patients likely to have undiagnosed COPD using administrative claims data., Methods: A predictive model was developed and validated utilizing a retro-spective cohort of patients with and without a COPD diagnosis (cases and controls), aged 40-89, with a minimum of 24 months of continuous health plan enrollment (Medicare Advantage Prescription Drug [MAPD] and commercial plans), and identified between January 1, 2009, and December 31, 2012, using Humana's claims database. Stratified random sampling based on plan type (commercial or MAPD) and index year was performed to ensure that cases and controls had a similar distribution of these variables. Cases and controls were compared to identify demographic, clinical, and health care resource utilization (HCRU) characteristics associated with a COPD diagnosis. Stepwise logistic regression (SLR), neural networking, and decision trees were used to develop a series of models. The models were trained, validated, and tested on randomly partitioned subsets of the sample (Training, Validation, and Test data subsets). Measures used to evaluate and compare the models included area under the curve (AUC); index of the receiver operating characteristics (ROC) curve; sensitivity, specificity, positive predictive value (PPV); and negative predictive value (NPV). The optimal model was selected based on AUC index on the Test data subset., Results: A total of 50,880 cases and 50,880 controls were included, with MAPD patients comprising 92% of the study population. Compared with controls, cases had a statistically significantly higher comorbidity burden and HCRU (including hospitalizations, emergency room visits, and medical procedures). The optimal predictive model was generated using SLR, which included 34 variables that were statistically significantly associated with a COPD diagnosis. After adjusting for covariates, anticholinergic bronchodilators (OR = 3.336) and tobacco cessation counseling (OR = 2.871) were found to have a large influence on the model. The final predictive model had an AUC of 0.754, sensitivity of 60%, specificity of 78%, PPV of 73%, and an NPV of 66%., Conclusions: This claims-based predictive model provides an acceptable level of accuracy in identifying patients likely to have undiagnosed COPD in a large national health plan. Identification of patients with undiagnosed COPD may enable timely management and lead to improved health outcomes and reduced COPD-related health care expenditures.

Objective: To assess factors associated with inpatient readmission among a US managed care population with chronic obstructive pulmonary disease (COPD)., Background: COPD is often accompanied by intermittent acute exacerbations, which may result in hospitalizations. These exacerbations are often associated with an increased frequency of subsequent exacerbations, which may lead to inpatient readmissions., Methods: We assessed US managed care claims data for enrollees≥40 years old with an inpatient admission with a primary diagnosis of COPD (ICD-9-CM codes 491.xx, 492.xx or 496.xx) between 1 January 2010 and 31 December 2013 (discharge date of first observed inpatient admission defined the "index date"). Patients were required to be continuously enrolled for ≥12 months before the index date. Two non-mutually exclusive cohorts were analyzed: (1) patients with ≥30 days of post-index date continuous enrollment (to evaluate 30-day readmission) and (2) patients with ≥90 days of post-index date continuous enrollment (to evaluate 90-day readmission). Logistic regression evaluated the association between patient characteristics and risk of 30- and 90-day COPD-related and all-cause readmission., Results: After applying selection criteria, 140,981 patients had ≥30 days of enrollment post-index date, and 123,545 patients had ≥90 days of enrollment post-index date. Within 30 days, nearly 20% of patients had an all-cause readmission and 7% had a COPD-related readmission. Within 90 days, 28% had an all-cause readmission and 12% had a COPD-related readmission. Logistic regression indicated that longer length of stay, older age, greater comorbidity burden, specific comorbidities and COPD complexity were associated with significantly greater odds of COPD-related 30- and 90-day readmission. Results for all-cause readmission were generally similar., Conclusions: Many of the factors associated with inpatient readmission documented here can be ascertained at discharge and may be used to inform discharge plans, with the end goal of improving patient outcomes, including reducing the risk of readmission.

This study assessed blood pressure (BP) goal maintenance in patients controlled with olmesartan monotherapy after switching to another angiotensin type II receptor blocker (ARB). Hypertensive patients prescribed olmesartan monotherapy were identified from GE Healthcare's Centricity electronic medical record between 2007 and 2011. After documentation of BP goal (<140/90 mm Hg) attainment, patients were placed into the continuation cohort if olmesartan monotherapy was maintained or into the switch cohort if they were changed to irbesartan, losartan, or valsartan. Follow-up assessments were the first BP measurement 28 to 390 days after attaining BP goal (continuation cohort) or after prescribing an alternative ARB (switch cohort). Of 3412 patients included (3027 continuation cohort, 385 switch cohort), 52% were women and mean age was 58.0 years. In the switch cohort, 310 (80.5%) were switched to losartan (n=236), irbesartan (n=58), or valsartan (n=16) monotherapy and 75 (19.5%) were switched to combination antihypertensive therapy. Mean baseline and follow-up BP were 122.5/75.8 mm Hg and 126.6/77.6 mm Hg, respectively, in the continuation cohort (P<.001) and 123.5/75.4 mm Hg and 129.6/78.5 mm Hg, respectively, in the switch cohort (P<.001). BP goal maintenance was 78.7% and 72.2% in the continuation and switch cohort, respectively (odds ratio, 0.707; 95% confidence interval, 0.555-0.899). Patients who continued on olmesartan monotherapy after attaining BP goal had a higher percentage of BP goal maintenance than patients who switched therapy., (©2013 Wiley Periodicals, Inc.)

We have developed and validated a sandwich chemiluminescence immunoassay (SCIA) which measures polycyclic aromatic hydrocarbon (PAH)-DNA adducts combining high throughput and adequate sensitivity, appropriate for evaluation of adduct levels in human population studies. Fragmented DNA is incubated with rabbit antiserum elicited against DNA modified with r7,t8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) and subsequently trapped by goat anti-rabbit IgG bound to a solid surface. Anti-single-stranded (ss) DNA antibodies binds in a quantity proportional to the adduct levels and is detected by chemiluminescence. The BPDE-DNA SCIA has a limit of detection of 3 adducts per 10(9) nucleotides with 5 μg DNA per well. We have validated the BPDE-DNA SCIA using DNA modified in vitro, DNA from benzo[a]pyrene (BP)-exposed cultured cells and mice. The levels of adduct measured by SCIA were lower (30-60%) than levels of bulky DNA adducts measured in the same samples by (32)P-postlabelling. The BPDE-DNA SCIA also detected adducts produced in vivo by PAHs other than BP. When blood DNA samples from maternal/infant pairs were assayed by BPDE-DNA SCIA, the adduct levels obtained were significantly correlated. However, there was no correlation between (32)P-postlabelling and SCIA values for the same samples. The SCIA can be extended to any DNA adduct and is expected to provide, when fully automated, a valuable high-throughput approach in large-scale population studies.

Background: Investigations of the presence of the precarcinogenic DNA adduct O⁶-methylguanine (O6-meG) in humans and its association with exposure or cancer risk have been hindered by the absence of analytic methods of adequate sensitivity and throughput. We report the development, validation, and application of an ELISA-type assay for O6-meG appropriate for large-scale population studies., Methods: In the new analytic method, restriction enzymes are used to digest DNA to fragments of size expected to contain no more than one O6-meG residue. Anti-adduct antisera are used to transfer O6-meG-containing fragments to a solid surface, where they are detected using anti-ssDNA antisera, the high ratio of normal nucleotides to adducts providing a strong signal enhancement., Results: An assay with a limit of detection of 1.5 adducts/10⁹ nucleotides using 10 μg of DNA, a dynamic range of approximately two orders of magnitude and satisfactory precision and accuracy characteristics was established and validated. Analysis of samples from 120 subjects from the Rhea mother-child cohort in Crete led to the detection of O6-meG in 70% of maternal and 50% of cord blood buffy coat samples at mean levels of 0.65 and 0.38 adducts/10⁸ nucleotides, respectively., Conclusions: The frequent observation of O6-meG in human DNA is compatible with dietary compounds (e.g. N-nitroso compounds or their precursors), or endogenous processes being responsible for the formation of this adduct., Impact: The new assay opens the way for large-scale population studies of O6-meG as a biomarker of exposure or risk. The approach used in this assay can, in principle, be extended to any DNA adduct for which suitable antisera are available., (©2011 AACR.)

Iron metabolism during pregnancy is biased toward maintaining the fetal supply, even at the cost of anemia in the mother. The mechanisms regulating this are not well understood. Here, we examine iron deficiency and supplementation on the hierarchy of iron supply and the gene expression of proteins that regulate iron metabolism in the rat. Dams were fed iron-deficient diets for 4 wk, mated, and either continued on the deficient diet or an iron-supplemented diet during either the first half or the second half of their pregnancy. A control group was maintained on normal iron throughout. They were killed at 0.5, 12.5, or 21.5 days of gestation, and tissues and blood samples were collected. Deficiency and supplementation had differential effects on maternal and fetal hematocrit and liver iron levels. From early in pregnancy, a hierarchy of iron supply is established benefiting the fetus to the detriment of the mother. Transferrin receptor, transferrin receptor 2, and hepcidin mRNA expression were regulated by both iron deficiency and supplementation. Expression patterns showed both organ and supplementation protocol dependence. Further analysis indicated that iron levels in the fetal, and not maternal, liver regulate the expression of liver transferrin receptor and hepcidin expression in the mother.

BeWo cells are a placental cell line that has been widely used as an in vitro model for the placenta. The b30 subclone of these cells can be grown on permeable membranes in bicameral chambers to form confluent cell layers, enabling rates of both nutrient uptake into the cells from the apical surface and efflux from the basolateral membrane to be determined. The aim of this study was to evaluate structural and functional properties of confluent b30 BeWo cell layers grown in bicameral chambers, focusing on the potential application for studying receptor-mediated uptake and transport of transferrin (Tf)-bound iron (Fe-Tf). While it proved extremely difficult to establish and maintain an intact BeWo cell monolayer, it was possible to grow the cells to a confluent multilayer. Iron, applied as Fe-Tf, was rapidly transported across this cell layer; 9.3 +/- 0.5% of the total dose was transported after 8 h, equivalent to 38.8 +/- 2.1 pmol.cm(-2).h(-1). Transfer of Tf across the cell layer was much more limited; 2.4 +/- 0.2% of the total dose was transported after 8 h, equivalent to 5.0 +/- 0.4 pmol.cm(-2).h(-1). Compartmental modeling of these data suggested that iron was transported across the cell layer predominantly, if not exclusively, via a transcellular route, whereas Tf taken up into the cells was predominantly recycled back to the apical compartment. The results suggest that these cells are very efficient at transporting iron and, under carefully controlled conditions, can be a valuable tool for the study of iron transport in the placenta.

Background: Obesity is associated with a chronic and low-grade inflammation which may cause hypoferremia as seen in patients with chronic inflammatory diseases. The aim of the present study was to investigate the relationship between iron status and markers of inflammation in morbidly obese women and the effect of bariatric surgery., Methods: Our cohort of patients consisted of 178 morbidly obese females selected for bariatric surgery. Clinical and biochemical data were recorded before surgery, and histopathological studies were carried out on preoperative liver biopsy samples. Fifty-five patients have been followed up after bariatric surgery., Results: A high prevalence of iron depletion was present in this cohort, with 53% having a transferrin saturation ratio below 0.20. Iron depletion was significantly correlated with raised levels of indices of inflammation, C-reactive protein (CRP), orosomucoid and haptoglobin), and with the white blood cell count. In multivariate analysis, orosomucoid and CRP were independently associated with iron depletion. Moreover, 6 months after bariatric surgery, inflammation level decreased, which was inversely correlated with the increase in transferrin saturation., Conclusions: Iron depletion is common in morbidly obese women. Low-grade chronic inflammation associated with obesity could be a modulator of iron uptake and utilization. Bariatric surgery may reduce chronic inflammation and improve iron status.

The Ity/Lsh/Bcg locus encodes the macrophage protein Slc11a1/Nramp1, which protects inbred mice against infection by diverse intracellular pathogens including Leishmania, Mycobacterium, and Salmonella. Human susceptibility to infectious and inflammatory diseases, including rheumatoid arthritis, inflammatory bowel disease, and tuberculosis, shows allelic association with a highly polymorphic regulatory, Z-DNA-forming microsatellite of (GT/AC)n dinucleotides within the proximal SLC11A1 promoter. We surmised that cis-acting allelic polymorphisms may underlie heritable differences in SLC11A1 expression and phenotypic variation in disease risk. However, it is unclear what may underlie such variation in SLC11A1 allele expression. Here we show that hypoxia-inducible Factor 1 (HIF-1) regulates allelic variation in SLC11A1 expression by binding directly to the microsatellite during macrophage activation by infection or inflammation. Targeted Hif-1alpha ablation in murine macrophages attenuated Slc11a11 expression and responsiveness to S typhimurium infection. Our data also showed that HIF-1 may be functionally linked to complex prototypical inflammatory diseases associated with certain SLC11A1 alleles. As these alleles are highly polymorphic, our finding suggests that HIF-1 may influence heritable variation in SLC11A1-dependent innate resistance to infection and inflammation within and between populations. This report also suggests that microsatellites may play critical roles in the directional evolution of complex heritable traits by regulating gene expression phenotypes.

P2Y receptors couple to G proteins and either mobilize intracellular Ca(2+) or alter cAMP levels to modulate the activity of Ca(2+)- and cAMP-sensitive ion channels. We hypothesize that increased ion transport into the lumen of MDCK cysts can osmotically drive fluid movement and increase cyst size. Furthermore, activation of the adenylate cyclase/cAMP pathway may trigger cell proliferation via an extracellular signal-related kinase cascade. To test this hypothesis, several P2Y receptor inhibitors were used on the MDCK in vitro model of renal cyst formation. The nonspecific P2 receptor inhibitors reactive blue 2 and suramin reduced cyst growth significantly, as did PPADS and, to a lesser extent, the P2Y(1)-specific antagonist MRS2179. Cyst growth was reduced by approximately 50% when ATP was removed from the culture medium with apyrase, although stable analogs of ATP failed to increase cyst size. The nonselective P2X receptor inhibitor Coomassie brilliant blue G was ineffective at reducing cyst growth, suggesting no involvement of P2X receptors. Finally, the presence of selective inhibitors of ERK activation (either PD98059 or U0126) greatly reduced cyst growth, whereas in untreated cysts ERK activity was observed to increase with time. We conclude that stimulation of endogenous P2Y receptors by extracellular ATP increases growth of MDCK cysts via cAMP-dependent activation of the ERK pathway. P2Y receptor antagonists may have therapeutic potential in reducing cyst size and slowing disease progression; although further studies in vitro and in vivo are needed to investigate the specificity and role of these P2Y receptors in renal cystic diseases.

Backgrounds & Aims: Hepcidin is an acute-phase response peptide. We have investigated the possible involvement of hepcidin in massive obesity, a state of chronic low-grade inflammation. Three groups of severely obese patients with or without diabetes or nonalcoholic steatohepatitis were investigated., Methods: Hepcidin expression was studied in liver and adipose tissue of these patients. Hepcidin regulation was investigated in vitro by adipose tissue explant stimulation studies., Results: Hepcidin was expressed not only in the liver but also at the messenger RNA (mRNA) and the protein levels in adipose tissue. Moreover, mRNA expression was increased in adipose tissue of obese patients. The presence of diabetes or NASH did not modify the hepcidin expression levels in liver and adipose tissue. In adipose tissue, mRNA expression correlated with indexes of inflammation, interleukin-6, and C-reactive protein. Interleukin-6 also promoted in vitro hepcidin expression. A low transferrin saturation ratio was observed in 68% of the obese patients; moreover, 24% of these patients presented with anemia. The observed changes in iron status could be due to the role of hepcidin as a negative regulator of intestinal iron absorption and macrophage iron efflux. Interestingly, a feedback control mechanism on hepcidin expression related to low transferrin saturation occurred in the liver but not in the adipose tissue., Conclusions: Hepcidin is a proinflammatory adipokine and may play an important role in hypoferremia of inflammation in obese condition.

We investigated the potential of a new family of lipidic peptide dendrimers in protein transduction into cultured cells. Dendrimer-protein interaction was determined by gel retardation assays using purified recombinant protein. To assess intracellular protein delivery, two marker proteins were used: recombinant firefly luciferase and a Cy3-labeled monoclonal antibody to the c-myc proto-oncogene. Protein delivery was determined by luciferase assays and fluorescence microscopy, respectively. While there was minimal delivery of luciferase or antibody in the absence of the dendrimers, the latter increased protein delivery substantially. Luciferase delivery was concentration and cell type-dependent; the efficiency of delivery also varied with the number of terminal amino groups on the dendrimers. In previous reports, we showed that these dendrimers could be used for gene and drug delivery; the data we report herein suggest that they may also be capable of intracellular protein delivery. This finding has important implications for the use of these dendrimers in protein therapeutics and vaccinology., ((c) 2006 Wiley-Liss, Inc. and the American Pharmacists Association)

Background: The molecular identification and characterization of the adenosine triphosphate (ATP)-sensitive family of P2 receptors is comparatively new. There are two main subgroups, each with several subtypes and widespread tissue distribution, including the kidney. A unique member of the P2X subgroup of P2 receptors is the ATP-gated ion channel P2X(7), which on activation can cause cell blebbing, cytokine release, and cell death by necrosis or apoptosis. We report expression of this receptor in normal rat kidney and in two chronic models of glomerular injury: streptozotocin-induced (STZ) diabetes and ren-2 transgenic (TGR) hypertension., Methods: At different time points in these models, we used a polyclonal antibody to the P2X(7) receptor and immunohistochemistry to determine its expression and distribution. We also used Western blotting and real-time polymerase chain reaction (PCR) to detect changes in P2X(7) receptor protein and mRNA expression, respectively., Results: We found only low-level glomerular immuno-staining for the P2X(7) receptor in normal rat kidney, but intense P2X(7) receptor immunostaining of glomeruli in kidneys from diabetic animals at 6 and 9 weeks, and in hypertensive animals at 12 weeks. In diabetic animals, real-time PCR demonstrated a approximately tenfold increase in glomerular P2X(7) receptor mRNA relative to control, and Western blotting confirmed an increase in protein. Immunohistochemistry and immunoelectron microscopy showed staining of glomerular podocytes, which was both intracellular and at the plasma membrane., Conclusion: We conclude that the P2X(7) receptor is not expressed appreciably under normal conditions, but that following glomerular injury it is significantly up-regulated, mainly in podocytes, though also in endothelial and mesangial cells, of animals with STZ-induced diabetes mellitus or TGR hypertension. Although the exact function and regulation of this receptor remain unclear, its association with inflammatory cytokine release and cell death suggests that increased expression might be involved in the pathogenesis of glomerular cell injury or repair.

The relationship between haem biosynthesis and intestinal iron absorption in mice was investigated by ascertaining the effect of the haem synthesis inhibitor, griseofulvin, on duodenal iron absorption using both in vivo and in vitro measurements. Urinary 5-aminolaevulinic acid levels were increased within 24 hr of feeding mice with griseofulvin diet (2.5% w/w), with more marked increases seen after 3-7 days. Urinary porphobilinogen levels also showed a similar trend. In vivo intestinal iron absorption was significantly reduced (P<0.05) in experimental mice, mainly due to reduction in the transfer of 59Fe from the enterocytes to the portal circulation. In vitro studies using isolated duodenal fragments also exhibited marked decreases in both iron uptake and Fe (III) reduction. Changes in mucosal Divalent Metal Transporter 1 (DMT-1), Dcytb and Ireg1 (iron regulated protein 1) mRNA levels paralleled the changes in iron absorption. The reduction in iron absorption after griseofulvin treatment was normalised when mice were simultaneously injected with haem-arginate. These data support the hypothesis that intermediates in haem biosynthesis, particularly 5-aminolaevulinic acid, regulate intestinal iron absorption.

The metabolism of chlorogenic acid, naringin, and rutin, representative members of three common families of dietary polyphenols, the hydroxycinnamates, the flavanones, and the flavonols, respectively, was studied in an in vitro mixed culture model of the human colonic microflora. Time- and concentration-dependent degradation of all three compounds was observed, which was associated with the following metabolic events after cleavage of the ester or glycosidic bond: reduction of the aliphatic double bond of the resulting hydroxycinnamate caffeic acid residue; dehydroxylation and ring fission of the heterocyclic C-ring of the resulting deglycosylated flavanone, naringenin, and of the deglycosylated flavonol, quercetin (which differed depending on the substitution). The metabolic events, their sequences, and major phenolic end products, as identified by GC-MS or LC-MS/MS, were elucidated from the structural characteristics of the investigated compounds. The major phenolic end products identified were 3-(3-hydroxyphenyl)-propionic acid for chlorogenic acid, 3-(4-hydroxyphenyl)-propionic acid and 3-phenylpropionic acid for naringin, and 3-hydroxyphenylacetic acid and 3-(3-hydroxyphenyl)-propionic acid for rutin. The degree of degradation of the compounds studied was significantly influenced by the substrate concentration as well as individual variations in the composition of the fecal flora. The results support extensive metabolism of dietary polyphenols in the colon, depending on substrate concentration and residence time, with resultant formation of simple phenolics, which can be considered biomarkers of colonic metabolism if subsequently absorbed. It is also apparent that a relatively small number of phenolic degradation products are formed in the colon from the diverse group of natural polyphenols.

Iron (Fe) deficiency anaemia during pregnancy results in an increased risk of perinatal mortality and morbidity and is a significant factor for increased risk of disease in later life. Consequently we have developed a rat model to study the relationship between maternal Fe deficiency and postnatal growth and blood pressure in the offspring. Weanlings were fed a control or Fe-deficient diet prior to and throughout pregnancy. At term, all pups were cross-fostered to control fed dams and weaned onto control diet. At birth, pups from deficient dams had a greater mortality rate, were smaller and had reduced haematocrit and liver Fe levels. They also had larger hearts, smaller kidneys and spleens and unchanged livers (relative organ weight). The pups grew normally. At 6 weeks, male pups from deficient dams had a higher and females a lower blood pressure than their normal counterparts. At 10 and 16 weeks, blood pressure in the males from deficient dams was still raised and in the females was now greater than controls. The haematocrit was lower in males throughout the 16 weeks and in females until 10 weeks of age. There was no significant difference in the offsprings' liver Fe stores at 6, 10 or 16 weeks. Duodenal Fe uptake in both the Fe-deficient mother and newborn offspring was significantly increased. By cross-fostering, we have eliminated confounding factors, such as maternal anaemia during lactation and show, unequivocally, that prenatal nutrition is critical for the development of normal postnatal function.

The effect of Hfe (haemochromatosis) gene deletion on the hypoxic response of iron absorption was investigated. Hfe knock-out mice were exposed to 0.5 atmospheres hypoxia for 3 d before in vivo iron absorption was measured. Both wild-type and Hfe knock-out mice had similar (two- to threefold) increases in iron absorption in response to hypoxia. We conclude that the Hfe gene product is not required for mice to increase iron absorption rates in response to hypoxia. The data further support the hypothesis that at least two independent mechanisms for the regulation of iron absorption exist, only one of which requires Hfe.

During pregnancy, nutrients are transferred from mother to fetus across the placenta. The mechanisms whereby this occurs, and the adaptations that occur in response to deficiency or overload of iron (Fe) and copper (Cu) are examined in this review. Fe deficiency during pregnancy is common and has serious consequences both in the short and the long term such as fetal growth retardation and cardiovascular problems in the adult offspring. Similarly, Cu deficiency, although not so common, also has deleterious effects. The placenta minimizes the effect of the deficiency by up-regulating the proteins involved in Fe transfer. For example, transferrin receptor levels increase inversely to maternal Fe levels. Divalent metal transporter 1 (DMT1) mRNA in the iron-responsive element (IRE) regulated, but not the non-IRE regulated form is increased, as is the placenta Cu oxidase. Conversely, iron-regulated gene 1 (IREG1) expression is not affected. Fe deficiency increases Cu levels in maternal liver, serum and placenta, but has much less effect in the fetal serum and liver. Apart from maternal ceruloplasmin, mRNA levels of Cu-related proteins are not changed. The Cu oxidase, which we suggest fulfils the function of hephaestin in placenta, is regulated by Cu as well as by Fe. Fe deficiency also has marked effects on cytokine levels in the placenta. Tumor necrosis factor alpha (TNFalpha) and TNFalpha receptor 1 (TNFalphaR1) levels both increase. The data show that altering Fe status has a marked effect on metabolism of other metals and of other important mediators of cell function. This is particularly important during pregnancy, when the developing fetus is very vulnerable to inappropriate micronutrient status.

Hereditary hemochromatosis is a common iron-loading disorder found in populations of European descent. It has been proposed that mutations causing loss of function of HFE gene result in reduced iron incorporation into immature duodenal crypt cells. These cells then overexpress genes for iron absorption, leading to inappropriate cellular iron balance, a persistent iron deficiency of the duodenal mucosa, and increased iron absorption. The objective was to measure duodenal iron content in Hfe knock-out mice to test whether the mutation causes a persistent decrease in enterocyte iron concentration. In both normal and Hfe knock-out mice, duodenal nonheme iron content was found to correlate with liver iron stores (P <.001, r = 0.643 and 0.551, respectively), and this effect did not depend on dietary iron levels. However, duodenal iron content was reduced in Hfe knock-out mice for any given content of liver iron stores (P <.001).

Ghrelin is a 28-residue peptide hormone that is principally released from the stomach during fasting and prior to eating. Two forms are present in human plasma: the unmodified peptide and a less abundant acylated version, in which octanoic acid is attached to the third residue, a serine, via an ester linkage. The acylated form of ghrelin acts as a ligand for the growth hormone secretagogue receptor and can stimulate the release of growth hormone from the pituitary gland. It also initiates behavioral and metabolic adaptations to fasting. Here we show that an immobilized form of ghrelin specifically binds a species of high density lipoprotein associated with the plasma esterase, paraoxonase, and clusterin. Both free ghrelin and paraoxon, a substrate for paraoxonase, can inhibit this interaction. An endogenous species of ghrelin is found to co-purify with high density lipoprotein during density gradient centrifugation and subsequent gel filtration. This interaction links the orexigenic peptide hormone ghrelin to lipid transport and metabolism. Furthermore, the interaction of the esterified hormone ghrelin with a species of HDL containing an esterase suggests a possible mechanism for the conversion of ghrelin to des-acyl ghrelin.

Hephaestin was implicated in mammalian iron homeostasis following its identification as the defective gene in murine sex-linked anaemia. It is a member of the family of copper oxidases that includes mammalian ceruloplasmin, factors V and VIII, yeast fet3 and fet5 and bacterial ascorbate oxidase. Hephaestin is different from ceruloplasmin, a soluble ferroxidase, in having a membrane-spanning region towards the C-terminus. Here we report the gene structure, spanning approximately 100 kb, of the human homologue of mouse hephaestin. The sequence was assembled from the cDNA clones and the chromosome X genomic sequence data available at the Sanger Centre. It has an open reading frame that encodes a protein of 1158 residues, 85% identical with the murine homologue. A model of the N-terminal ecto-domain has been built based on the known three-dimensional structure of human ceruloplasmin. The overall tertiary structure for the hephaestin and the putative residues involved in binding copper and iron appear to be highly conserved between these proteins, which suggests they share the same fold and a conserved function.

The levels of bulky DNA adducts were measured by (32)P-post-labelling in lymphocytes of 194 non-smoking students living in the city of Athens and the region of Halkida, Greece, once in the winter and again in the following summer. Personal exposures to particulate-bound polycyclic aromatic hydrocarbons (PAH) were significantly higher in Athens subjects during both seasons. There was hardly any diagonal radioactive zone in the pattern of DNA adducts observed. Highest adduct levels were observed in a sub-group of subjects living in or near the Halkida Institute campus, which was located in rural surroundings with a minimal burden of urban air pollution. The remaining Halkida subjects had intermediate levels, while Athens subjects showed the lowest levels. This trend, which was observed over both monitoring seasons, consistently paralleled the variation in three markers of exposure to environmental tobacco smoke (ETS), namely (i) declared times of exposure to ETS during the 24 h prior to blood donation, (ii) plasma cotinine levels and (iii) chrysene/benzo[g,h,i]perylene ratios in the profile of personal PAH exposure. Furthermore, among the Halkida campus area subjects (but not the remaining subjects) positive correlations were observed between DNA adducts and (i) measured personal exposures to chrysene or benzo[a]pyrene, (ii) time of declared ETS exposure and (iii) chrysene/benzo[g,h,i] perylene ratios. These correlations suggest that, for a group suffering minimal exposure to urban air pollution, exposure to ETS was a significant determinant of the observed DNA damage. Gender had a consistent and significant effect on adduct levels (males having higher levels), which remained significant even after multiple regression analysis. Habitual consumption of roasted meat was significantly associated with an enhancement of adduct levels and the effect was strengthened when only individuals unexposed to ETS were taken into consideration. No significant effects were observed for other dietary parameters or factors reflecting exposure to air pollution.