Probing protein stability with unnatural amino acids.

Unnatural amino acid mutagenesis, in combination with molecular modeling and simulation techniques, was used to probe the effect of side chain structure on protein stability. Specific replacements at position 133 in T4 lysozyme included (i) leucine (wt), norvaline, ethylglycine, and alanine to measure the cost of stepwise removal of methyl groups from the hydrophobic core, (ii) norvaline and O-methyl serine to evaluate the effects of side chain solvation, and (iii) leucine, S,S-2-amino-4-methylhexanoic acid, and S-2-amino-3-cyclopentylpropanoic acid to measure the influence of packing density and side chain conformational entropy on protein stability. All of these factors (hydrophobicity, packing, conformational entropy, and cavity formation) significantly influence protein stability and must be considered when analyzing any structural change to proteins.

Ancillary risk information and pharmacogenetic tests: social and policy implications.

Some pharmacogenetic tests may provide ancillary disease risk information. To evaluate evidence and assess the social and policy implications of ancillary disease risk information associated with candidate pharmacogenetic variants, We conducted a literature search and abstract review of disease susceptibility studies for each of 42 gene variants potentially associated with drug response. Twenty-two variants (53%) had suggested association with disease risk in at least two studies, and sixteen (38%) were for diseases other than the pharmacogenetic indication. Seven variants (16%) were associated with risk for at least two different diseases. Pharmacogenetic tests have the potential to provide ancillary disease risk information, and this potential should be considered as pharmacogenetic tests are brought into clinical use. Implications will vary with each test but tests should be evaluated individually within a framework that outlines the potential implications of ancillary information.

Are Evidence Standards Different for Genomic- vs. Clinical-Based Precision Medicine? A Quantitative Analysis of Individualized Warfarin Therapy.

Evidence requirements for implementation of precision medicine (PM), whether informed by genomic or clinical data, are not well defined. Evidence requirements are driven by uncertainty and its attendant consequences; these aspects can be quantified by a novel technique in health economics: value of information analysis (VOI). We utilized VOI analysis to compare the evidence levels over time for warfarin dosing based on pharmacogenomic vs. amiodarone-warfarin drug-drug interaction information. The primary outcome was the expected value of perfect information (EVPI), which is an estimate of the upper limit of the societal value of conducting future research. Over the past decade, the EVPI for the pharmacogenomic strategy decreased from $1,550 to $140 vs. $1,220 to $280 per patient for the drug-interaction strategy. Evidence levels thus appear to be higher for pharmacogenomic-guided vs. drug-interaction-guided warfarin dosing. Clinical guidelines and reimbursement policies for warfarin PM could be informed by these findings.

The value of routine pharmacogenomic screening-Are we there yet? A perspective on the costs and benefits of routine screening-shouldn't everyone have this done?

Although there are several examples in which pharmacogenomic testing seems to provide clinical and economic value, use of pharmacogenomics as a tool to improve drug therapy through routine screening of unselected patients is currently tentative. An informal evaluation of the clinical benefits and economic costs of pharmacogenomic screening suggests that improving the evidence base, addressing uncertainty, and facilitating implementation can lead to practical and cost-effective pharmacogenomic screening programs.

The potential clinical and economic benefits of silver alloy urinary catheters in preventing urinary tract infection.

BACKGROUND: Catheter-associated urinary tract infection (UTI) is associated with increased morbidity, mortality, and costs. A recent meta-analysis concluded that silver alloy catheters reduce the incidence of UTI by 3-fold; however, clinicians must decide whether the efficacy of such catheters is worth the extra per unit cost of $5.30. OBJECTIVE: To assess the clinical and economic impact of using silver alloy urinary catheters in hospitalized patients. METHODS: The decision model, performed from the health care payer's perspective, evaluated a simulated cohort of 1000 hospitalized patients on general medical, surgical, urologic, and intensive care services requiring short-term urethral catheterization (2-10 days). We compared 2 catheterization strategies: silver alloy catheters and standard (noncoated) urinary catheters. Outcomes included the incidence of symptomatic UTI and bacteremia and direct medical costs. RESULTS: In the base-case analysis, use of silver-coated catheters led to a 47% relative decrease in the incidence of symptomatic UTI from 30 to 16 cases per 1000 patients (number needed to treat = 74) and a 44% relative decrease in the incidence of bacteremia from 4.5 to 2.5 cases per 1000 patients (number needed to treat = 500) compared with standard catheters. Use of silver alloy catheters resulted in estimated cost savings of $4.09 per patient compared with standard catheter use ($20.87 vs $16.78). In a multivariate sensitivity analysis using Monte Carlo simulation, silver-coated catheters provided clinical benefits over standard catheters in all cases and cost savings in 84% of cases. CONCLUSIONS: Using silver alloy catheters in hospitalized patients requiring short-term urinary catheterization reduces the incidence of symptomatic UTI and bacteremia, and is likely to produce cost savings compared with standard catheters.

Incidence and long-term cost of steroid-related side effects after renal transplantation.

Corticosteroids are an essential component of most immunosuppressive regimens currently used in renal transplantation because of their efficacy in reducing acute rejection and improving graft survival. Steroids, however, are associated with numerous side effects that lead to increased patient morbidity and mortality. The incidence and economic cost of steroid-related side effects have not been quantitatively assessed. Thus, based on a systematic review of the published literature, we estimated the incidence of steroid-related hypertension (15%), posttransplantation diabetes mellitus (10%), peripheral fractures (2% per year), avascular necrosis of the hip (8%), and cataracts (22%). In addition, we estimated that approximately 5% of patients who have cataracts or avascular necrosis of the hip require surgery. We used these literature-based estimates in a model to project the costs of treating side effects over a 10-year posttransplantation time frame for a 50-patient cohort that represented an average-sized renal transplant center. Steroid-induced hypertension and its complications were the most expensive side effect ($93,900), followed closely by posttransplantation diabetes ($89,700) and avascular necrosis of the hip ($61,700). Cataracts and peripheral bone fractures were less costly ($16,300 and $4,300, respectively). The cumulative projected 10-year cost of all side effects for the 50-patient cohort was $265, 900, or $5,300 per transplant patient. Steroid-related side effects add to the long-term cost of medical care of renal transplant recipients. These costs provide a rationale for further investigation of steroid-sparing immunosuppression protocols.

The clinical and economic consequences of nosocomial central venous catheter-related infection: are antimicrobial catheters useful?

Central venous catheters (CVCs) are essential for many hospitalized patients, but they are associated with important infectious complications. Recent studies have indicated that CVCs coated with antimicrobial agents reduce the incidence of catheter-related bloodstream infection (CR BSI). To estimate the clinical and economic consequences of short-term central venous catheter-related infection and the potential usefulness of antimicrobial-coated catheters, we reviewed and synthesized the available relevant literature. Statistical pooling was used to estimate the incidence of both catheter colonization and CR BSI. The attributable mortality of CR BSI was also evaluated. In addition, the economic consequences of both local and systemic catheter-related infection was estimated from literature reports that used micro-costing and other techniques. Among patients in whom standard, noncoated CVCs are in place for an average of 8 days, 24.7% are expected to develop catheter colonization (95% confidence interval [CI(95)], 22.0%-27.5%). Approximately 5.2% (CI(95), 3.9%-6.5%) will develop CR BSI. The attributable mortality of CR BSI remains unclear, but recent studies are consistent with a range from 4% to 20%. An episode of local catheter-related infection leads to an additional cost of approximately $400, whereas the additional cost of CR BSI ranges from approximately $6,005 to $9,738. Formal economic analyses indicate that CVCs coated with antibacterial agents (such as chlorhexidine-silver sulfadiazine or minocycline-rifampin) likely reduce infectious complications, yielding economic advantages. In light of the substantial clinical and economic burden of catheter-related infection, hospital personnel should adopt proven cost-effective methods to reduce this common and important nosocomial complication.

The use of meta-analysis in cost-effectiveness analysis. Issues and recommendations.

Meta-analysis is used to statistically pool the results from individual studies, usually randomised trials, to obtain an estimate of the summary effect size across studies. The summary measure from a meta-analysis is often used to derive the probability of treatment success in a cost-effectiveness analysis. Recently, LeLorier and colleagues questioned the ability of meta-analysis to accurately predict the results of a subsequent large-scale trial, implying that the use of a summary measure from a meta-analysis may be inappropriate in an economic evaluation. We comment on this potential shortcoming by first providing an outline of the use of meta-analysis results in a cost-effectiveness analysis. Then, using examples of discrepancies between meta-analyses and subsequent large trials noted by LeLorier and colleagues, we examine the potential impact of using the results from a small trial versus a meta-analysis. We found that the meta-analyses were comparable to or better than small trials at predicting the results of subsequent large trials. We, therefore, argue that a meta-analysis of homogeneous studies can provide a reasonable estimate of the treatment effect for use in a cost-effectiveness analysis when no large, definitive clinical trial has been performed. However, care must be taken not to over-interpret the precision of the estimate, since both the homogeneity and quality of the primary studies need to be considered. We conclude by providing guidance on the appropriate use of summary measures derived from meta-analyses for cost-effectiveness studies.

Economic evaluation of systemic treatments for cytomegalovirus retinitis in patients with AIDS.

OBJECTIVE: To determine the cost of using systemic therapy to treat newly diagnosed cytomegalovirus (CMV) retinitis in persons with AIDS. DESIGN: Incidence-based simulation model of CMV treatment from a government payer perspective. SETTING: Swiss healthcare system. PATIENTS AND PARTICIPANTS: Patients with AIDS and newly diagnosed CMV retinitis. INTERVENTIONS: Patients were assigned to 1 of 4 treatment regimens for induction and maintenance therapy: (i) intravenous (IV) cidofovir induction and maintenance (cidofovir IV/IV); (ii) IV foscarnet induction and maintenance (foscarnet IV/IV); (iii) IV ganciclovir induction and maintenance (ganciclovir IV/IV); and (iv) IV ganciclovir induction and oral (PO) ganciclovir maintenance (ganciclovir IV/PO). Following a second relapse, patients were assigned to one of the other regimens. MAIN OUTCOME MEASURES: Time to first and subsequent progression, duration of maintenance treatment and direct medical expenditures [1998 Swiss francs (SwF)] . RESULTS: The median time to first progression was longest for cidofovir IV/IV, followed by foscarnet IV/IV, ganciclovir IV/IV and ganciclovir IV/PO. Mean survival was 13 months and mean costs for this period in the base case were lowest in those initially treated with cidofovir (SwF146,742), followed by initial treatment with foscarnet IV/IV (SwF194,809), ganciclovir IV/PO (SwF195,190) and ganciclovir IV/IV (SwF243,964). Costs were most sensitive to changes in efficacy estimates. CONCLUSIONS: Of the regimens studied, initiation of treatment with systemic cidofovir appears least costly over a 13-month period.

Assessing the cost-effectiveness of pharmacogenomics.

The use of pharmacogenomics to individualize drug therapy offers the potential to improve drug effectiveness, reduce adverse side effects, and provide cost-effective pharmaceutical care. However, the combinations of disease, drug, and genetic test characteristics that will provide clinically useful and economically feasible therapeutic interventions have not been clearly elucidated. The purpose of this paper was to develop a framework for evaluating the potential cost-effectiveness of pharmacogenomic strategies that will help scientists better understand the strategic implications of their research, assist in the design of clinical trials, and provide a guide for health care providers making reimbursement decisions. We reviewed concepts of cost-effectiveness analysis and pharmacogenomics and identified 5 primary characteristics that will enhance the cost-effectiveness of pharmacogenomics: 1) there are severe clinical or economic consequence that are avoided through the use of pharmacogenomics, 2) monitoring drug response using current methods is difficult, 3) a well-established association between genotype and clinical phenotype exists, 4) there is a rapid and relatively inexpensive genetic test, and 5) the variant gene is relatively common. We used this framework to evaluate several examples of pharmacogenomics. We found that pharmacogenomics offers great potential to improve patients' health in a cost-effective manner. However, pharmacogenomics will not be applied to all currently marketed drugs, and careful evaluations are needed on a case-by-case basis before investing resources in research and development of pharmacogenomic-based therapeutics and making reimbursement decisions.

Genetic risk factors for major bleeding in patients treated with warfarin in a community setting.

The influence of warfarin pharmacogenomics on major bleeding risk has been little studied in long-term users and non-specialist care settings. We conducted a case-control study to evaluate associations between CYP2C9*2/*3, VKORC1(1173), and CYP4F2*3 variants and major bleeding among patients treated with warfarin in a community setting. We calculated major bleeding odds ratios, adjusting for race, duration of warfarin use, age, gender, and body mass index. In 265 cases and 305 controls with 3.4 and 3.7 mean years of warfarin use, respectively, CYP4F2*3 was associated with decreased major bleeding risk (odds ratio: 0.62; 95% confidence interval: 0.43-0.91). CYP2C9*2/*3 and VKORC1(1173) had null associations overall, but there was a nonsignificant increase in major bleeding risk in patients with duration <6 months (odds ratio: 1.30; 95% confidence interval: 0.60-2.83; odds ratio: 1.23; 95% confidence interval: 0.57-2.64, respectively). In summary, in the largest study of warfarin pharmacogenomics and major bleeding to date, we found a 38% lower risk in patients with CYP4F2*3, potentially reflecting interaction with warfarin and dietary vitamin K intake and warranting additional evaluation.

A risk-benefit assessment of prasugrel, clopidogrel, and genotype-guided therapy in patients undergoing percutaneous coronary intervention.

The objective of this study was to quantitatively evaluate the clinical benefits and harms of prasugrel, clopidogrel, and a CYP2C19 genotype-guided drug selection strategy for patients with acute coronary syndrome (ACS) and planned percutaneous coronary intervention (PCI). We used decision-analytic techniques to model the risks and benefits of alternative antiplatelet strategies. Sensitivity and scenario analyses were conducted to assess the uncertainty of the results. Prasugrel demonstrated little difference in net benefit as compared with clopidogrel (+0.02 quality-adjusted life-years (QALYs); 95% confidence range (CR), -0.23 to 0.21). The genotype-guided strategy had a 93% probability of greater net benefit as compared with clopidogrel (+0.05 QALYs; 95% CR, -0.02 to 0.11), and 66% probability of greater net benefit as compared with prasugrel (+0.03 QALYs; 95% CR, -0.13 to 0.24). Prasugrel and clopidogrel differ in their risk-benefit profiles but appear to offer similar net benefit on average. Use of patient-specific factors such as CYP2C19 genotype offers promise for developing a personalized medicine approach to antiplatelet treatment regimens.

Can modeling of health outcomes facilitate regulatory decision making? The benefit-risk tradeoff for rosiglitazone in 1999 vs. 2007.

Rosiglitazone was initially approved for type 2 diabetes monotherapy. We tested health-outcomes modeling as an aid to regulatory decision making by quantifying the incremental net benefit (INB) value of rosiglitazone (relative to a comparator), both at the time of first approval (1999) and at the FDA advisory committee review (2007). Using 1999 data, rosiglitazone was projected to provide an additional 0.639 years of life (0.373 quality-adjusted life years (QALYs)) relative to placebo but a loss of 0.312 years (0.173 QALYs) relative to glyburide, with uncertainty in reduction of hemoglobin A(1c) (HbA(1c)) level having the greatest impact on the benefit-risk profile. By 2007, rosiglitazone was projected to provide an additional 0.222 years (0.091 QALYs) vs. glyburide and 0.026 years vs. metformin (0.009 QALYs). Modeling suggested that the use of rosiglitazone as monotherapy was not initially warranted, given the uncertainty with regard to benefit. Despite similar net benefit (NB) as metformin shown in postmarketing data, residual cardiovascular (CV) concerns did not support the use of rosiglitazone as first-line therapy. We adapted a mathematical diabetes model to estimate NB and uncertainty of diabetes monotherapy.

Evaluating anti-viral drug selection and treatment duration in HBeAg-negative chronic hepatitis B: a cost-effectiveness analysis.

BACKGROUND: Several anti-viral treatments are now available for HBeAg-negative chronic hepatitis B (CHB), but the clinical and economic outcomes of potential treatment strategies and durations are unclear. AIM: To examine the clinical and economic outcomes of potential treatment strategies and durations for HBeAg-negative CHB. METHODS: We conducted a cost-utility analysis from a payer perspective over a lifetime time horizon. Disease progression probabilities, costs and quality of life data were derived from the literature. We evaluated 5-year, 10-year, lifetime and 5 on-1 off treatment durations. For each of these treatment durations, we evaluated initial therapy with entecavir, lamivudine or adefovir, with addition of adefovir or entecavir for patients who developed virological breakthrough because of resistance (12 strategies total). RESULTS: Increasing treatment duration improved quality-adjusted life-years (QALYs) and was generally cost-effective for all three drugs. However, a 5 on-1 off strategy was the most cost-effective: lifetime vs. 5 on-1 off entecavir had an ICER of $148,200/QALY. In probabilistic sensitivity analyses, entecavir 5 on-1 off was the preferred strategy over the range of commonly reimbursed cost-effectiveness thresholds. Lifetime treatment was preferred to a 5 on-1 off strategy, if treatment durability was < 10%. CONCLUSION: The results of our analysis suggest that in HBeAg-negative CHB infection, a 5 on-1 off treatment strategy with entecavir improves health outcomes in a cost-effective manner compared to alternative strategies.

Use of pharmacogenetic and clinical factors to predict the therapeutic dose of warfarin.

Initiation of warfarin therapy using trial-and-error dosing is problematic. Our goal was to develop and validate a pharmacogenetic algorithm. In the derivation cohort of 1,015 participants, the independent predictors of therapeutic dose were: VKORC1 polymorphism -1639/3673 G>A (-28% per allele), body surface area (BSA) (+11% per 0.25 m(2)), CYP2C9(*)3 (-33% per allele), CYP2C9(*)2 (-19% per allele), age (-7% per decade), target international normalized ratio (INR) (+11% per 0.5 unit increase), amiodarone use (-22%), smoker status (+10%), race (-9%), and current thrombosis (+7%). This pharmacogenetic equation explained 53-54% of the variability in the warfarin dose in the derivation and validation (N= 292) cohorts. For comparison, a clinical equation explained only 17-22% of the dose variability (P < 0.001). In the validation cohort, we prospectively used the pharmacogenetic-dosing algorithm in patients initiating warfarin therapy, two of whom had a major hemorrhage. To facilitate use of these pharmacogenetic and clinical algorithms, we developed a nonprofit website, http://www.WarfarinDosing.org.

Cost effectiveness of inhaled steroid withdrawal in outpatients with chronic obstructive pulmonary disease.

BACKGROUND: The evidence for the effectiveness and safety of inhaled corticosteroids (ICS) in chronic obstructive pulmonary disease (COPD) is inconclusive. This study determined the cost effectiveness of withdrawing fluticasone propionate (FP) in outpatients with COPD. METHODS: The cost effectiveness analysis was based on a randomised, placebo controlled FP withdrawal study. After a 4 month run in period on FP, patients were randomly assigned to continue FP 500 microg twice daily or to receive placebo for 6 months. A decision analytical model evaluated the 6 month incremental cost effectiveness of the ICS versus ICS withdrawal strategy. One way sensitivity analyses and a Monte Carlo simulation were performed to evaluate the robustness of the findings. RESULTS: The average patient with COPD in the FP group generated 511 in direct medical costs, including 238 for FP. The cost of the placebo strategy was 456. The higher direct drug cost of 212 per patient for the FP strategy during the 6 month follow up period compared with the placebo group was partially offset by a lower exacerbation and hospital admission cost of 157. The 6 month incremental cost effectiveness of the FP strategy compared with placebo was 110 per exacerbation prevented and 1286 per hospital admission prevented. CONCLUSIONS: Over a 6 month period, withdrawing FP in a pre-selected trial population of COPD patients led to absolute cost savings but with a higher rate of exacerbations and hospital admissions.

Modeling protein stability: a theoretical analysis of the stability of T4 lysozyme mutants.

Free energy calculations were conducted to determine the relative stability of the unnatural amino acid mutants of T4 lysozyme norvaline (Nvl) and O-methyl-serine (Mse) and of alanine at residue 133, which is leucine in the native sequence. These calculations were performed both to assess the validity of the methodology and to gain a better understanding of the forces which contribute to protein stability. Peptides of different length were used to model the denatured state. Restraints were employed to force sampling of the side chain chi1 dihedral of the perturbed side chain, and the effect of protein repacking in response to mutation was studied through the use of different constraint sets. In addition, the convergence behavior and hysteresis of the simulations in the folded and unfolded states were determined. The calculated results agree well with experiment, + 1.84 versus + 1.56 kcal/mol for Mse-->Nvl and -3.48 versus -2.2 to -3.6 kcal/mol for Nvl-->Ala. We find that free energy calculations can provide useful insights to protein stability when conducted carefully on a well chosen system. Our results suggest that loss of packing interactions in the native state is a major source of destabilization for mutants which decrease the amount of buried nonpolar surface area and that subtle responses of the backbone affect the magnitude of the loss of stability. We show that the conformational freedom of the chi1 dihedral has a noticeable effect on protein stability and that the solvation of amino acid side chains is strongly influenced by interactions with the peptide backbone.