Stimulus-Controlled Anion Binding and Transport by Synthetic Receptors.

Anionic species are omnipresent and involved in many important biological processes. A large number of artificial anion receptors has therefore been developed. Some of these are capable of mediating transmembrane transport. However, where transport proteins can respond to stimuli in their surroundings, creation of synthetic receptors with stimuli-responsive functions poses a major challenge. Herein, we give a full overview of the stimulus-controlled anion receptors that have been developed thus far, including their application in membrane transport. In addition to their potential operation as membrane carriers, the use of anion recognition motifs in forming responsive membrane-spanning channels is discussed. With this review article, we intend to increase interest in transmembrane transport among scientists working on host-guest complexes and dynamic functional systems in order to stimulate further developments.

Photoswitchable Bis(amidopyrroles): Modulating Anion Transport Activity Independent of Binding Affinity.

Toward photocontrol of anion transport across the bilayer membrane, stiff-stilbene, which has dimethyl substituents in the five-membered rings, is functionalized with amidopyrrole units. UV-vis and 1H NMR studies show high photostability and photoconversion yields. Where the photoaddressable (E)- and (Z)-isomers exhibit comparable binding affinities, as determined by 1H NMR titrations, fluorescence-based transport assays reveal significantly higher transport activity for the (Z)-isomers. Changing the binding affinity is thus not a necessity for modulating transport. Additionally, transport can be triggered in situ by light.

Neurotrophic Factors as Regenerative Therapy for Neurodegenerative Diseases: Current Status, Challenges and Future Perspectives.

Neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), multiple sclerosis (MS), spinal cord injury (SCI), and amyotrophic lateral sclerosis (ALS), are characterized by acute or chronic progressive loss of one or several neuronal subtypes. However, despite their increasing prevalence, little progress has been made in successfully treating these diseases. Research has recently focused on neurotrophic factors (NTFs) as potential regenerative therapy for neurodegenerative diseases. Here, we discuss the current state of knowledge, challenges, and future perspectives of NTFs with a direct regenerative effect in chronic inflammatory and degenerative disorders. Various systems for delivery of NTFs, such as stem and immune cells, viral vectors, and biomaterials, have been applied to deliver exogenous NTFs to the central nervous system, with promising results. The challenges that currently need to be overcome include the amount of NTFs delivered, the invasiveness of the delivery route, the blood-brain barrier permeability, and the occurrence of side effects. Nevertheless, it is important to continue research and develop standards for clinical applications. In addition to the use of single NTFs, the complexity of chronic inflammatory and degenerative diseases may require combination therapies targeting multiple pathways or other possibilities using smaller molecules, such as NTF mimetics, for effective treatment.

Photomodulation of Transmembrane Transport and Potential by Stiff-Stilbene Based Bis(thio)ureas.

Membrane transport proteins fulfill important regulatory functions in biology with a common trait being their ability to respond to stimuli in the environment. Various small-molecule receptors, capable of mediating transmembrane transport, have been successfully developed. However, to confer stimuli-responsiveness on them poses a fundamental challenge. Here we demonstrate photocontrol of transmembrane transport and electric potential using bis(thio)ureas derived from stiff-stilbene. UV-vis and 1H NMR spectroscopy are used to monitor E-Z photoisomerization of these bis(thio)ureas and 1H NMR titrations reveal stronger binding of chloride to the (Z)-form than to the (E)-form. Additional insight into the binding properties is provided by single crystal X-ray crystallographic analysis and DFT geometry optimization. Importantly, the (Z)-isomers are much more active in transmembrane transport than the respective (E)-isomers as shown through various assays. As a result, both membrane transport and depolarization can be modulated upon irradiation, opening up new prospects toward light-based therapeutics as well as physiological and optopharmacological tools for studying anion transport-associated diseases and to stimulate neuronal activity, respectively.

Engineering of regulatory T cells by means of mRNA electroporation in a GMP-compliant manner.

Regulatory T cells (Tregs) are crucial in inducing and maintaining tolerance. This unique capacity of Tregs, in combination with proof-of-principle in preclinical studies, highlights the potential clinical use of Tregs for the treatment of autoimmunity and transplant rejection. Although proven to be safe and well tolerated in the first clinical trials, only modest clinical results were observed. In this regard, it has been hypothesized that current challenges lie in the development of antigen-specific Tregs. Here, we present an innovative, good manufacturing practices (GMP)-compliant manufacturing protocol for Tregs applicable in a clinical-grade setting, allowing efficient and safe redirection of Treg specificity. First, a soluble polymer conjugated with antibodies to CD3 and CD28 and high amounts of exogenous IL-2 for in vitro Treg expansion resulted in a >70-fold and 185-fold increase of a pure population of CD4+CD127-CD25hi Tregs and CD4+CD127-CD25+CD45RA+ Tregs, respectively. Next, as a proof-of-principle, expanded Tregs were engineered by means of TCR-encoding mRNA electroporation to generate antigen-specific Tregs. This resulted in an expression of the newly introduced TCR in up to 85% of Tregs. Moreover, we did not observe a negative effect on the phenotype of Tregs, as demonstrated by the expression of FOXP3, Helios, CTLA-4 and CCR4, nor on the TSDR methylation status. Importantly, mRNA-engineered Tregs were still able to induce in vitro suppression of effector T cells and produced anti-inflammatory, but not pro-inflammatory, cytokines when activated. In conclusion, our findings demonstrate that high numbers of stable and functional Tregs can be obtained with high purity and successfully engineered for gain of function, in a GMP-compliant manner. We envisage that this clinical-grade protocol will provide solid basis for future clinical application of mRNA-engineered Tregs.

Are Cell-Based Therapies Safe and Effective in the Treatment of Neurodegenerative Diseases? A Systematic Review with Meta-Analysis.

Over the past two decades, significant advances have been made in the field of regenerative medicine. However, despite being of the utmost clinical urgency, there remains a paucity of therapeutic strategies for conditions with substantial neurodegeneration such as (progressive) multiple sclerosis (MS), spinal cord injury (SCI), Parkinson's disease (PD) and Alzheimer's disease (AD). Different cell types, such as mesenchymal stromal cells (MSC), neuronal stem cells (NSC), olfactory ensheathing cells (OEC), neurons and a variety of others, already demonstrated safety and regenerative or neuroprotective properties in the central nervous system during the preclinical phase. As a result of these promising findings, in recent years, these necessary types of cell therapies have been intensively tested in clinical trials to establish whether these results could be confirmed in patients. However, extensive research is still needed regarding elucidating the exact mechanism of action, possible immune rejection, functionality and survival of the administered cells, dose, frequency and administration route. To summarize the current state of knowledge, we conducted a systematic review with meta-analysis. A total of 27,043 records were reviewed by two independent assessors and 71 records were included in the final quantitative analysis. These results show that the overall frequency of serious adverse events was low: 0.03 (95% CI: 0.01-0.08). In addition, several trials in MS and SCI reported efficacy data, demonstrating some promising results on clinical outcomes. All randomized controlled studies were at a low risk of bias due to appropriate blinding of the treatment, including assessors and patients. In conclusion, cell-based therapies in neurodegenerative disease are safe and feasible while showing promising clinical improvements. Nevertheless, given their high heterogeneity, the results require a cautious approach. We advocate for the harmonization of study protocols of trials investigating cell-based therapies in neurodegenerative diseases, adverse event reporting and investigation of clinical outcomes.

Hepatocyte-specific IKK-ß activation enhances VLDL-triglyceride production in APOE*3-Leiden mice.

Low-grade inflammation in different tissues, including activation of the nuclear factor κB pathway in liver, is involved in metabolic disorders such as type 2 diabetes and cardiovascular diseases (CVDs). In this study, we investigated the relation between chronic hepatocyte-specific overexpression of IkB kinase (IKK)-ß and hypertriglyceridemia, an important risk factor for CVD, by evaluating whether activation of IKK-ß only in the hepatocyte affects VLDL-triglyceride (TG) metabolism directly. Transgenic overexpression of constitutively active human IKK-ß specifically in hepatocytes of hyperlipidemic APOE*3-Leiden mice clearly induced hypertriglyceridemia. Mechanistic in vivo studies revealed that the hypertriglyceridemia was caused by increased hepatic VLDL-TG production rather than a change in plasma VLDL-TG clearance. Studies in primary hepatocytes showed that IKK-ß overexpression also enhances TG secretion in vitro, indicating a direct relation between IKK-ß activation and TG production within the hepatocyte. Hepatic lipid analysis and hepatic gene expression analysis of pathways involved in lipid metabolism suggested that hepatocyte-specific IKK-ß overexpression increases VLDL production not by increased steatosis or decreased FA oxidation, but most likely by carbohydrate-responsive element binding protein-mediated upregulation of Fas expression. These findings implicate that specific activation of inflammatory pathways exclusively within hepatocytes induces hypertriglyceridemia. Furthermore, we identify the hepatocytic IKK-ß pathway as a possible target to treat hypertriglyceridemia.

Cost-effectiveness of tipranavir versus comparator protease inhibitor regimens in HIV infected patients previously exposed to antiretroviral therapy in the Netherlands.

BACKGROUND: This study compares the costs and effects of a regimen with ritonavir-boosted tipranavir (TPV/r) to a physician-selected genotypically-defined standard-of-care comparator protease inhibitor regimen boosted with ritonavir (CPI/r) in HIV infected patients that were previously exposed to antiretroviral therapy in the Netherlands. METHODS: We compared the projected lifetime costs and effects of two theoretical groups of 1000 patients, one receiving a standard of care regimen with TPV/r as a component and the other receiving a standard of care regimen with CPI/r. A 3-stage Markov model was formulated to represent three different consecutive HAART regimens. The model uses 12 health states based on viral load and CD4+ count to simulate disease progression. The transition probabilities for the Markov model were derived from a United States cohort of treatment experienced HIV patients. Furthermore, the study design was based on 48-week data from the RESIST-2 clinical trial and local Dutch costing data. Cost and health effects were discounted at 4% and 1.5% respectively according to the Dutch guideline. The analysis was conducted from the Dutch healthcare perspective using 2006 unit cost prices. RESULTS: Our model projects an accumulated discounted cost to the Dutch healthcare system per patient receiving the TPV/r regimen of euro167,200 compared to euro145,400 for the CPI/r regimen. This results in an incremental cost of euro21,800 per patient. The accumulated discounted effect is 7.43 life years or 6.31 quality adjusted life years (QALYs) per patient receiving TPV/r, compared to 6.91 life years or 5.80 QALYs per patient receiving CPI/r. This translates into an incremental effect of TPV/r over CPI/r of 0.52 life years gained (LYG) or 0.51 QALYs gained. The corresponding incremental cost effectiveness ratios (iCERs) are euro41,600 per LYG and euro42,500 per QALY. CONCLUSION: We estimated the iCER for TPV/r compared to CPI/r at approximately euro40,000 in treatment experienced HIV-1 infected patients in the Netherlands. This ratio may well be in range of what is acceptable and warrants reimbursement for new drug treatments in the Netherlands, in particular in therapeutic areas as end-stage oncology and HIV and other last-resort health-care interventions.

Combination vaccine against invasive meningococcal B and pneumococcal infections: potential epidemiological and economic impact in the Netherlands.

BACKGROUND: Streptococcus pneumoniae and Neisseria meningitidis group B are among the main causes of invasive bacterial meningitis infections in infants. Worldwide, these diseases lead to significant mortality, morbidity and costs. The societal impact is especially severe since the majority of cases occur in very young infants. A combination vaccine consisting of 9-valent conjugated pneumococcal and meningococcal B components is currently being developed. The aim of this study was to estimate the potential impact and cost effectiveness from the societal perspective of vaccinating infants in The Netherlands with this combination pneumococcal and meningococcal B vaccine versus no vaccination. METHODS: A Markov cycle model was developed using epidemiological and healthcare resource use data from 1996 to 2001. This model was used to project the annual costs, benefits and health gains associated with vaccinating all newborns. The base year for the costing was 2003 and all costs and health effects were discounted at 4%. The results of the analysis are expressed in costs per QALY and both probabilistic and univariate sensitivity analyses were used to identify the robustness of the results. RESULTS: Annually, an average of 755 cases of invasive pneumococcal and meningococcal B infection occurred in infants aged 0-10 years in The Netherlands. Introduction of the combination vaccine would prevent 201 cases of meningococcal B meningitis and 165 cases of invasive pneumococcal disease per year. Additionally, 3410 cases of pneumococcal pneumonia and 46,350 cases of otitis media would be prevented. Vaccination would save 35 lives per year and prevent 71 cases of severe sequelae. This translates into 860 life-years gained, or 1128 QALYs gained. Alongside these health gains, vaccination would prevent euro 17,681,370 of direct medical and indirect costs attributable to meningococcal and pneumococcal infections in The Netherlands. Depending on vaccine price, cost effectiveness varied from euro 3160 (vaccine price per dose euro 20) to euro 32,170 (vaccine price euro 60 per dose) per QALY. Base-case cost effectiveness (vaccine price euro 40) was euro 17,700 per QALY. The model was most sensitive to changes in incidence, vaccine price and duration of protective efficacy. CONCLUSION: Our results suggest that the introduction of a combination meningococcal B and pneumococcal vaccine into the Dutch infant vaccination programme is potentially cost effective compared with no vaccination.

Routine HIV screening of sexually transmitted disease clinic attenders has favourable cost-effectiveness ratio in low HIV prevalence settings.

HIV screening for attenders of clinics for sexually transmitted disease (STD) may identify individuals with high-risk sexual behaviour and avert HIV infections in partners. Extending our previous analysis in AIDS, we performed an economic evaluation of HIV screening of STD-clinic attenders in Rotterdam, the Netherlands. The results, at euro 2987 per life-year gained, display a favourable economic profile. Routine HIV screening in STD clinics should be considered as a highly cost-effective prevention measure even in countries with a low HIV prevalence.

Epidemiologic impact and cost-effectiveness of universal infant vaccination with a 7-valent conjugated pneumococcal vaccine in the Netherlands.

BACKGROUND: Streptococcus pneumoniae is one of the main causes of bacterial meningitis, bacteremia, pneumonia, and otitis media in the Netherlands. These diseases lead to substantial mortality, morbidity, and costs. The societal impact is especially severe because most cases occur in very young infants. OBJECTIVE: The aim of this study was to estimate the epidemiological impact and cost-effectiveness of universal infant vaccination with a 7-valent conjugated pneumococcal vaccine in the Netherlands. METHODS: Decision analysis was performed using epidemiological data and data on health care resource use from 1996 to 2001. A model was used to project the impact of pneumococcal vaccination on the incidence of pneumococcal infections in infants and children from birth to age 10 years. Costs, benefits, and health gains were estimated, and cost-effectiveness was calculated. All analyses were performed from a societal perspective. RESULTS: On average, 339 cases per year of invasive pneumococcal infection occurred in infants and children from birth to age 10 years in the Netherlands from 1996 to 2001. The model predicted that introduction of the 7-valent conjugated pneumococcal vaccine would prevent 48 cases of bacterial meningitis and 88 cases of pneumococcal bacteremia per year, as well as 42,695 cases of pneumococcal otitis media and 3411 cases of invasive pneumococcal pneumonia. The model also predicted that vaccination would save 13 lives per year and prevent 31 cases of lifelong sequelae, rendering 382 discounted quality-adjusted life-years (QALYs) gained or 329 discounted life-years gained per year. Considering these health gains, vaccination would prevent Euro 9,453,600 of direct and indirect medical costs of meningococcal and pneumococcal infections in the Netherlands, including acute medical care, management of sequelae, and lost time at work. With a vaccine price of Euro 40 per dose, the base-case cost-effectiveness ratio would be Euro 71,250 per QALY. The model was sensitive to changes in incidence of infections, vaccine effectiveness, and vaccine price. CONCLUSIONS: Our analytic model predicted that universal pneumococcal vaccination of infants in the Netherlands could prevent a large number of pneumococcal infections and considerably reduce related mortality and morbidity. However, the baseline cost-effectiveness ratio of such a vaccination program would be relatively unfavorable compared with other interventions implemented in the Netherlands.

Valuing prevention through economic evaluation: some considerations regarding the choice of discount model for health effects with focus on infectious diseases.

In cost-effectiveness analysis, the valuing of costs and health effects over time remains a controversial issue. The debate mostly focuses on whether the discount rates for health and money should be equal and which discounting model and time preferences are most appropriate. In this paper we add to the debate by arguing that the assessment of effectiveness of a preventive intervention may influence the choice of the discounting procedure for health. Health effects in cost-effectiveness analysis are commonly expressed in life-years gained, QALYs gained or lives saved. These denominators are only indirect and partial measures of the effects of a preventive intervention. The actual effect of the intervention is a reduction of the risk of mortality and morbidity in a given period of time. This risk reduction will not always coincide with the moment at which the impact on (quality-adjusted) life-years gained is made (i.e. at risk exposure), for example when preventing chronic disease with an asymptomatic stage. In this paper we show that truly acknowledging the origin of health benefits could have implications for the discounting procedure. We present a discounting model that adequately focuses on the reduction of risk. This model recognises the potential interpretation of risk reduction for infection as an economic good to be acquired with associated mortality reductions as later indirect effects. This implies that our suggested discounting model focuses on the moment(s) of risk reduction. A numerical example illustrates our approach. We discuss the associated potential implications for public health policy and discuss how the effects of the intervention can be additionally corrected for societal preferences.

Methods for reducing peak pressure in laparoscopic grasping.

During tissue retraction with a laparoscopic grasper, tissue-damaging pressures can occur. Past research suggests that peak pressures can be considerably reduced by rounding the edges or covering the tip of the end effector with a silicon sleeve. To identify grasping methods that limit tissue damage, the effects of (a) Young's modulus of the end effector, (b) curvature of the end effector, and (c) angle with which the tissue is pulled relative to the plane of the end effector, on the pressure generated on the tissue were investigated. Artificial skin was placed between two non-serrated jaws, a pressure-sensitive film was interposed between the skin and upper jaw, and the end effector was loaded with 13 N. End effectors with Young's moduli of 0.09, 0.67, 1.49 MPa, and 69 GPa, and with non-rounded and 5 mm rounded edges were tested under pulling angles of 25°, 50°, and 75°. For non-rounded end effectors, the maximum pressure and the area across which pressure exceeded the safety threshold for tissue damage increased with Young's modulus and pulling angle. For rounded end effectors, maximum pressure did not increase monotonically with Young's modulus. Instead, the end effector with the second lowest Young's modulus yielded significantly lower maximum pressure than the end effector with the lowest Young's modulus. For rounded end effectors, pressures were below the safety threshold for all Young's moduli. This indicates that to prevent tissue damage, soft graspers may not be needed; rounding the edges of metal graspers could suffice for preventing tissue damage.

Towards a healthier discount procedure.

Most national guidelines for pharmacoeconomic research prescribe discounting, mostly of money and health against the same rate. There is much debate on whether this is adequate. Two theoretical arguments, the consistency argument of Weinstein and Stason, and the paralyzing paradox of Keeler and Cretin, are mostly responsible for the current standards. However, more recently, several authors have indicated that the basis to claim the necessity of using similar discount rates is rather weak, both practically and theoretically. In terms of finding a new theoretical basis on which to base discount rates for money and, in particular, health, Van Hout has made an important suggestion arguing that the discount rate for health could be based on the expected growth in life expectancy and the diminishing marginal utility related to such additional health. Similarly, Gravelle and Smith argue that if the value of health grows over time, discount rates that are used for costs cannot directly be applied to effects, but should be adjusted downwards.

Adherence of pharmacoeconomic studies to national guidelines in the Netherlands.

OBJECTIVE: This study examines the adherence of Dutch pharmacoeconomic studies to the national guidelines of conducting a pharmacoeconomic evaluation. METHODS: Dutch guidelines for pharmacoeconomic research were issued in 1999. All Dutch pharmacoeconomic studies that were published in English during 2000-2002 were selected for our review. Two reviewers examined each study for relevance and compared each study with the nine methodological guidelines selected. RESULTS: It was found that 29 studies satisfied the inclusion criteria. The societal perspective was taken in 13 out of the 29 studies (45%), an adequate time period of analysis was chosen in 21 (72%), effectiveness was explicitly differentiated from efficacy in 17 (59%), an incremental analysis was performed in 23 (79%), costs, benefits and health gains were discounted in 24 (83%), effectiveness was expressed in LYGs or QALYs in 16 (55%), reference prices were used in 8 (28%), subgroup analysis was presented in 13 (45%) and sensitivity analysis was included in 26 (90%). CONCLUSIONS: In this review we found that the adherence of studies to some of the Dutch guidelines for pharmacoeconomic studies is fair. However, major improvements are required with respect to the adoption of the societal perspective, presentation of adequate subgroup analyses and application of reference prices.

Drug costs developments after patent expiry of enalapril, fluoxetine and ranitidine: a study conducted for the Netherlands.

BACKGROUND: In order to increase price competition, government regulations focus on controlling drug costs. Drug costs after patent expiry are an area of particular interest because the substitution of branded medication with generics represents an opportunity for lowering drug costs. However, drug costs may not decrease after patent expiry, because of a lack of price competition and different national pricing systems. AIM: The aim of this study was to investigate the trends in the use of generics after patent expiry for enalapril, fluoxetine and ranitidine and the subsequent changes, if any, in the costs of these medications. METHODS: A drug-utilisation study was performed using data from a large sample of Dutch pharmacies. Both volumes (measured as defined daily doses [DDD] per 1000 population) as well as drug costs (calculated per DDD) prior to and after patent expiry were calculated. Costs per DDD were compared using trend-line analysis. In addition, the relative market shares of the different trade channels (branded, parallel imported and generic) were compared before and after patent expiry. RESULTS: The costs per DDD decreased for all three drugs and, as expected, these costs decrease more rapidly after patent expiry. Significant differences in the trend lines were found for enalapril and fluoxetine. CONCLUSIONS: Despite relatively high reimbursement prices for generics in the Netherlands, this example from the Dutch pharmaceutical market demonstrates the benefit of generic substitution for containing pharmaceutical costs, which contrasts with concerns raised by the Dutch government.

The use of modeling in the economic evaluation of vaccines.

As a consequence of the increased role of pharmacoeconomics in policy-making, economic evaluations are performed at more and more early stages in the development of a therapeutic. This implies the development of models to assess the future impact of an intervention and to account for the level of uncertainty in the associated parameters. This also applies for economic evaluations of vaccines, where not only progression of disease and associated costs are important, but the transmission of the causing agent in the target population also has to be modelled. In this review, we provide an overview of the models that have been used in recent publications on the pharmacoeconomics of vaccines and go deeper into some of the methodological issues associated with the use of models in the economic evaluation of vaccines.