Editorial – Avoiding Unethical Helicobacter pylori Clinical Trials: Susceptibility‐Based Studies and Probiotics as Adjuvants

As a general rule, any clinical study where the result is already known or when the investigator(s) compares an assigned treatment against another assigned treatment known to be ineffective in the study population (e.g., in a population with known clarithromycin resistance) is unethical. As susceptibility‐based therapy will always be superior to empiric therapy in any population with a prevalence of antimicrobial resistance >0%, any trial that randomizes susceptibility‐based therapy with empiric therapy would be unethical. The journal Helicobacter welcomes susceptibility or culture‐guided studies, studies of new therapies, and studies of adjuvants and probiotics. However, the journal will not accept for review any study we judge to be lacking clinical equipoise or which assign subjects to a treatment known to be ineffective, such as a susceptibility‐based clinical trial with an empiric therapy comparator. To assist authors, we provide examples and suggestions regarding trial design for comparative studies, for susceptibility‐based studies, and for studies testing adjuvants or probiotics.     

Genotype‐by‐environment interactions due to antibiotic resistance and adaptation in Escherichia coli

Mutations that are beneficial in one environment can have different fitness effects in other environments. In the context of antibiotic resistance, the resulting genotype‐by‐environment interactions potentially make selection on resistance unpredictable in heterogeneous environments. Furthermore, resistant bacteria frequently fix additional mutations during evolution in the absence of antibiotics. How do these two types of mutations interact to determine the bacterial phenotype across different environments? To address this, I used Escherichia coli as a model system, measuring the effects of nine different rifampicin resistance mutations on bacterial growth in 31 antibiotic‐free environments. I did this both before and after approximately 200 generations of experimental evolution in antibiotic‐free conditions (LB medium), and did the same for the antibiotic‐sensitive wild type after adaptation to the same environment. The following results were observed: (i) bacteria with and without costly resistance mutations adapted to experimental conditions and reached similar levels of competitive fitness; (ii) rifampicin resistance mutations and adaptation to LB both indirectly altered growth in other environments; and (iii) resistant‐evolved genotypes were more phenotypically different from the ancestor and from each other than resistant‐nonevolved and sensitive‐evolved genotypes. This suggests genotype‐by‐environment interactions generated by antibiotic resistance mutations, observed previously in short‐term experiments, are more pronounced after adaptation to other types of environmental variation, making it difficult to predict long‐term selection on resistance mutations from fitness effects in a single environment.     

Furazolidone treatment for Helicobacter Pylori infection: A systematic review and meta‐analysis

Antibiotic resistance is a major cause of Helicobacter pylori (H. pylori) treatment failures. Because the resistance rate of H. pylori to furazolidone is low, we aimed to assess the efficacy and safety of furazolidone. We searched the PubMed, Web of Science, Cochrane Library, and Embase databases and included randomized controlled trials (RCT) that either compared furazolidone to other antibiotics or changed the administered dose of furazolidone. A total of 18 articles were included in the meta‐analysis. According to the intention‐to‐treat (ITT) analysis, the total eradication rates of furazolidone‐containing therapy were superior to those of other antibiotic‐containing therapies (relative risk [RR] 1.07, 95% confidence interval [CI] 1.01‐1.14) (13 RCTs). Specifically, the eradication rates of furazolidone‐containing therapy were better than those for metronidazole‐containing therapy (RR 1.10, 95% CI: 1.01‐1.21 for ITT). The eradication rate of furazolidone‐containing bismuth‐containing quadruple therapy was 92.9% (95% CI: 90.7%‐95.1%) (PP). In addition, a higher daily dose of furazolidone increased the eradication rate (RR 1.17, 95% CI: 1.05‐1.31). And the incidence of some adverse effects, such as fever and anorexia, was higher in the furazolidone group than in the control group, the overall incidences of total side effects and severe side effects showed no significant differences between the groups. Furazolidone‐containing treatments could achieve satisfactory eradication rates and did not increase the incidence of total or severe adverse effects, but the incidence of milder side effects, such as fever and anorexia, should be considered when prescribing furazolidone‐containing treatments to patients.     

Predicting events in clinical trials using two time‐to‐event outcomes

In clinical trials with time‐to‐event outcomes, it is of interest to predict when a prespecified number of events can be reached. Interim analysis is conducted to estimate the underlying survival function. When another correlated time‐to‐event endpoint is available, both outcome variables can be used to improve estimation efficiency. In this paper, we propose to use the convolution of two time‐to‐event variables to estimate the survival function of interest. Propositions and examples are provided based on exponential models that accommodate possible change points. We further propose a new estimation equation about the expected time that exploits the relationship of two endpoints. Simulations and the analysis of real data show that the proposed methods with bivariate information yield significant improvement in prediction over that of the univariate method.     

Quantification of length‐bias in screening trials with covariate‐dependent test sensitivity

Length‐biased sampling exists in screening programs where longer duration disease is detected during the preclinical stage because a longer sojourn time (preclinical duration) has a higher probability of being screen detected. By modeling the course of disease, we quantify the effect of length‐biased sampling on clinical duration when cases are subject to periodic screening with variable test sensitivity. We use the highly flexible bivariate lognormal density to jointly model preclinical and clinical durations, and we model screening test sensitivity as a function of the sojourn time and number of previous false negative screens. We show that the mean clinical duration among screen‐detected cases can be up to 40% higher, with shrinking standard deviation, than those among nonscreen‐detected cases, due to biased sampling alone, irrespective of any possible benefit (increased survival time arising from earlier detection or reduction in mortality). These findings will aid in the design and interpretation of screening trials.     

Comparison of design strategies for a three‐arm clinical trial with time‐to‐event endpoint: Power,time‐to‐analysis,and operational aspects

To optimize resources, randomized clinical trials with multiple arms can be an attractive option to simultaneously test various treatment regimens in pharmaceutical drug development. The motivation for this work was the successful conduct and positive final outcome of a three‐arm randomized clinical trial primarily assessing whether obinutuzumab plus chlorambucil in patients with chronic lympocytic lymphoma and coexisting conditions is superior to chlorambucil alone based on a time‐to‐event endpoint. The inference strategy of this trial was based on a closed testing procedure. We compare this strategy to three potential alternatives to run a three‐arm clinical trial with a time‐to‐event endpoint. The primary goal is to quantify the differences between these strategies in terms of the time it takes until the first analysis and thus potential approval of a new drug, number of required events, and power. Operational aspects of implementing the various strategies are discussed. In conclusion, using a closed testing procedure results in the shortest time to the first analysis with a minimal loss in power. Therefore, closed testing procedures should be part of the statistician's standard clinical trials toolbox when planning multiarm clinical trials.     

Usefulness of resistant gene markers for predicting treatment outcome on second‐line anti‐tuberculosis drugs

Aim: Mutations in rrs [nucleotide (nt) 1401], gyrA gene (codons 90, 91 or 94), tlyA, ethA and thyA genes of Mycobacterium tuberculosis (MTB) were evaluated for their usefulness in predicting treatment outcome of kanamycin (KM), capreomycin (CPM), ofloxacin (OFX), ethionamide (ETH) and para‐aminosalicylic acid (PAS). Methods and Results: DNA sequence analyses of these genes were performed against 188 MTB isolates obtained from patients put on second‐line anti‐TB drugs (SLDs) with well‐documented clinical history and treatment outcome. Mutations in rrs and gyrA have 100% positive predictive value (PPV) in predicting treatment failure for KM and OFX, while 88·9 and 80% were obtained, respectively, when tlyA and rrs mutations were considered in CPM. For ETH and PAS, the PPV of using ethA and thyA mutations to predict treatment failure was 82·5 and 89·3%, respectively. Conclusions: Our study demonstrated high specificities of gene mutations in predicting poor treatment outcome; however, further technical advancement is required to make the molecular detection of resistances to other SLDs feasible in clinical laboratories. Significance and Impact of the Study: This is the first study to correlate different polymorphisms of major SLD resistance gene markers with predicted treatment outcome, using an international set of well‐documented clinical MTB strains.     

Knock‐on community impacts of a novel vector: spillover of emerging DWV‐B from Varroa‐infested honeybees to wild bumblebees

Novel transmission routes can directly impact the evolutionary ecology of infectious diseases, with potentially dramatic effect on host populations and knock‐on effects on the wider host community. The invasion of Varroa destructor, an ectoparasitic viral vector in Western honeybees, provides a unique opportunity to examine how a novel vector affects disease epidemiology in a host community. This specialist honeybee mite vectors deformed wing virus (DWV), an important re‐emerging honeybee pathogen that also infects wild bumblebees. Comparing island honeybee and wild bumblebee populations with and without V. destructor, we show that V. destructor drives DWV prevalence and titre in honeybees and sympatric bumblebees. Viral genotypes are shared across hosts, with the potentially more virulent DWV‐B overtaking DWV‐A in prevalence in a current epidemic. This demonstrates disease emergence across a host community driven by the acquisition of a specialist novel transmission route in one host, with dramatic community level knock‐on effects.     

Macrolide use in the previous years is associated with failure to eradicate Helicobacter pylori with clarithromycin‐containing regimens

Background

There is some evidence that prior use of macrolide antibiotics is a useful predictor of the likelihood of standard triple therapy failure in Helicobacter pylori eradication. In this study, we have evaluated whether previous intake of macrolides correlates with failure to eradicate H. pylori using two different first‐line clarithromycin‐containing regimens.

Materials and Methods

Retrospective study of 212 patients with H. pylori infection treated with one of two first‐line clarithromycin‐containing regimens: 108 patients treated with triple therapy for 10 days and 104 patients treated with concomitant therapy for 10 days. The intake of macrolides (clarithromycin, azithromycin, and other macrolides) prior to the eradication therapy was obtained from the electronic medical record, which contains information regarding all the medication prescribed to the patients since the year 2004.

Results

One hundred of 212 patients (47.2%) had received at least one treatment with macrolides during the years prior to the eradication therapy. H. pylori eradication rates were significantly lower in patients with previous use compared to patients without previous use of macrolides, both with triple therapy (60.8% vs 92.9%; P < .0001) and with concomitant therapy (85.7% vs 98.2%; P = .024).

Conclusions

Previous use of macrolides correlates with a low H. pylori eradication rate with triple and concomitant clarithromycin‐containing regimens. In addition, our study shows that in patients without previous use of macrolides, triple therapy achieves per‐protocol eradication rates over 90%.     

Selective RAF inhibitor impairs ERK1/2 phosphorylation and growth in mutant NRAS,vemurafenib‐resistant melanoma cells

The RAF inhibitor vemurafenib achieves remarkable clinical responses in mutant BRAF melanoma patients. However, vemurafenib is burdened by acquired drug resistance and by the side effects associated with its paradoxical activation of the ERK1/2 pathway in wild‐type BRAF cells. This paradoxical effect has driven the development of a new class of RAF inhibitors. Here, we tested one of these selective, non‐paradox‐inducing RAF inhibitors termed paradox‐breaker‐04 (PB04) or PLX7904. Consistent with its design, PB04 is able to efficiently inhibit activation of ERK1/2 in mutant BRAF melanoma cells but does not hyperactivate ERK1/2 in mutant RAS‐expressing cells. Importantly, PB04 inhibited ERK1/2 phosphorylation in mutant BRAF melanoma cells with acquired resistance to vemurafenib/PLX4720 that is mediated by a secondary mutation in NRAS. Consistent with ERK1/2 reactivation driving the re‐acquisition of malignant properties, PB04 promoted apoptosis and inhibited entry into S phase and anchorage‐independent growth in mutant N‐RAS‐mediated vemurafenib‐resistant cells. These data indicate that paradox‐breaker RAF inhibitors may be clinically effective as a second‐line option in a cohort of acquired vemurafenib‐resistant patients.     

Delayed egg‐laying and shortened incubation duration of Arctic‐breeding shorebirds coincide with climate cooling

Biological impacts of climate change are exemplified by shifts in phenology. As the timing of breeding advances, the within‐season relationships between timing of breeding and reproductive traits may change and cause long‐term changes in the population mean value of reproductive traits. We investigated long‐term changes in the timing of breeding and within‐season patterns of clutch size, egg volume, incubation duration, and daily nest survival of three shorebird species between two decades. Based on previously known within‐season patterns and assuming a warming trend, we hypothesized that the timing of clutch initiation would advance between decades and would be coupled with increases in mean clutch size, egg volume, and daily nest survival rate. We monitored 1,378 nests of western sandpipers, semipalmated sandpipers, and red‐necked phalaropes at a subarctic site during 1993–1996 and 2010–2014. Sandpipers have biparental incubation, whereas phalaropes have uniparental incubation. We found an unexpected long‐term cooling trend during the early part of the breeding season. Three species delayed clutch initiation by 5 days in the 2010s relative to the 1990s. Clutch size and daily nest survival showed strong within‐season declines in sandpipers, but not in phalaropes. Egg volume showed strong within‐season declines in one species of sandpiper, but increased in phalaropes. Despite the within‐season patterns in traits and shifts in phenology, clutch size, egg volume, and daily nest survival were similar between decades. In contrast, incubation duration did not show within‐season variation, but decreased by 2 days in sandpipers and increased by 2 days in phalaropes. Shorebirds demonstrated variable breeding phenology and incubation duration in relation to climate cooling, but little change in nonphenological components of traits. Our results indicate that the breeding phenology of shorebirds is closely associated with the temperature conditions on breeding ground, the effects of which can vary among reproductive traits and among sympatric species.     

Lack of evidence to favor specific preventive interventions in psychosis: a network meta‐analysis

Preventing psychosis in patients at clinical high risk may be a promising avenue for pre‐emptively ameliorating outcomes of the most severe psychiatric disorder. However, information on how each preventive intervention fares against other currently available treatment options remains unavailable. The aim of the current study was to quantify the consistency and magnitude of effects of specific preventive interventions for psychosis, comparing different treatments in a network meta‐analysis. PsycINFO, Web of Science, Cochrane Central Register of Controlled Trials, and unpublished/grey literature were searched up to July 18, 2017, to identify randomized controlled trials conducted in individuals at clinical high risk for psychosis, comparing different types of intervention and reporting transition to psychosis. Two reviewers independently extracted data. Data were synthesized using network meta‐analyses. The primary outcome was transition to psychosis at different time points and the secondary outcome was treatment acceptability (dropout due to any cause). Effect sizes were reported as odds ratios and 95% confidence intervals (CIs). Sixteen studies (2,035 patients, 57% male, mean age 20.1 years) reported on risk of transition. The treatments tested were needs‐based interventions (NBI); omega‐3 + NBI; ziprasidone + NBI; olanzapine + NBI; aripiprazole + NBI; integrated psychological interventions; family therapy + NBI; D‐serine + NBI; cognitive behavioural therapy, French & Morrison protocol (CBT‐F) + NBI; CBT‐F + risperidone + NBI; and cognitive behavioural therapy, van der Gaag protocol (CBT‐V) + CBT‐F + NBI. The network meta‐analysis showed no evidence of significantly superior efficacy of any one intervention over the others at 6 and 12 months (insufficient data were available after 12 months). Similarly, there was no evidence for intervention differences in acceptability at either time point. Tests for inconsistency were non‐significant and sensitivity analyses controlling for different clustering of interventions and biases did not materially affect the interpretation of the results. In summary, this study indicates that, to date, there is no evidence that any specific intervention is particularly effective over the others in preventing transition to psychosis. Further experimental research is needed.     

Using expert knowledge to incorporate uncertainty in cause‐of‐death assignments for modeling of cause‐specific mortality

Implicit and explicit use of expert knowledge to inform ecological analyses is becoming increasingly common because it often represents the sole source of information in many circumstances. Thus, there is a need to develop statistical methods that explicitly incorporate expert knowledge, and can successfully leverage this information while properly accounting for associated uncertainty during analysis. Studies of cause‐specific mortality provide an example of implicit use of expert knowledge when causes‐of‐death are uncertain and assigned based on the observer's knowledge of the most likely cause. To explicitly incorporate this use of expert knowledge and the associated uncertainty, we developed a statistical model for estimating cause‐specific mortality using a data augmentation approach within a Bayesian hierarchical framework. Specifically, for each mortality event, we elicited the observer's belief of cause‐of‐death by having them specify the probability that the death was due to each potential cause. These probabilities were then used as prior predictive values within our framework. This hierarchical framework permitted a simple and rigorous estimation method that was easily modified to include covariate effects and regularizing terms. Although applied to survival analysis, this method can be extended to any event‐time analysis with multiple event types, for which there is uncertainty regarding the true outcome. We conducted simulations to determine how our framework compared to traditional approaches that use expert knowledge implicitly and assume that cause‐of‐death is specified accurately. Simulation results supported the inclusion of observer uncertainty in cause‐of‐death assignment in modeling of cause‐specific mortality to improve model performance and inference. Finally, we applied the statistical model we developed and a traditional method to cause‐specific survival data for white‐tailed deer, and compared results. We demonstrate that model selection results changed between the two approaches, and incorporating observer knowledge in cause‐of‐death increased the variability associated with parameter estimates when compared to the traditional approach. These differences between the two approaches can impact reported results, and therefore, it is critical to explicitly incorporate expert knowledge in statistical methods to ensure rigorous inference.     

Clinical applications of palifermin: amelioration of oral mucositis and other potential indications

Mucositis is one of the most significant toxicities in cancer patients undergoing cytotoxic treatment. It can have a negative impact on both quality of life and health economics. Severe oral mucositis can contribute to hospitalization, need for narcotic analgesics, total parentral nutrition, suboptimal delivery of anti‐neoplastic treatment, and morbidity and mortality. Palifermin, a recombinant derivative of human keratinocyte growth factor, is the first active agent approved by the FDA for the prevention of severe oral mucositis in patients undergoing haematopoietic stem cell transplantation (HSCT). Several studies have also shown significant reduction in the incidence, severity and/or duration of oral mucositis in other high‐risk settings such as concurrent chemoradiotherapy (CT/RT) for patients with head and neck cancer, and use of mucotoxic chemotherapeutic agents such as doxorubicin in sarcoma and fluorouracil for the treatment of colorectal cancer. The reduction in mucositis has translated into amelioration of symptoms and improvement in daily functioning as measured by patient‐reported outcome in multiple studies. The clinical response to palifermin appears to be related in part to epithelial proliferation and mucosal thickening. Palifermin also has other potential clinical applications including the acceleration of immune reconstitution and inhibition of graft‐versus‐host disease in patients undergoing HSCT, and mitigation of dysphagia in lung cancer patients treated with concurrent CT/RT. Palifermin is generally well tolerated with mild‐to‐moderate skin and oral adverse events. Future studies may expand the use of palifermin into other areas that would benefit from its cytoprotective and regenerative effects.     

The Use of Two‐Way Linear Mixed Models in Multitreatment Meta‐Analysis

Summary Meta‐analysis summarizes the results of a series of trials. When more than two treatments are included in the trials and when the set of treatments tested differs between trials, the combination of results across trials requires some care. Several methods have been proposed for this purpose, which feature under different labels, such as network meta‐analysis or mixed treatment comparisons. Two types of linear mixed model can be used for meta‐analysis. The one expresses the expected outcome of treatments as a contrast to a baseline treatment. The other uses a classical two‐way linear predictor with main effects for treatment and trial. In this article, we compare both types of model and explore under which conditions they give equivalent results. We illustrate practical advantages of the two‐way model using two published datasets. In particular, it is shown that between‐trial heterogeneity as well as inconsistency between different types of trial is straightforward to account for.     

High Efficacy of 14‐Day Triple Therapy‐Based,Bismuth‐Containing Quadruple Therapy for Initial Helicobacter pylori Eradication

Background: The success rate of currently recommended 7‐day triple therapy with a PPI plus amoxicillin and clarithromycin has fallen into the unacceptable range. It is urgent to look for a new strategy to treat the infection of Helicobacter pylori. Aims: To observe the efficacy of triple therapy‐based, bismuth‐containing quadruple therapy for H. pylori treatment. Methods: A total of 160 patients with functional dyspepsia who were Hp+ were randomly assigned into two groups. Regimen: Omeprazole 20 mg, Amoxicillin 1.0 g, Clarithromycin 500 mg and Bismuth Potassium Citrate 220 mg, twice a day. Eighty patients received 7‐day quadruple therapy and 80 patients received the same therapy for 14 days. Six weeks after treatment, H. pylori eradication was assessed by ¹³C‐urea breath test. Minimal inhibitory concentrations of metronidazole, clarithromycin and amoxicillin of clinical isolates were determined by the twofold agar dilution method. Results: Fourteen‐day therapy led to a significant increase of H. pylori eradication success when compared to 7‐day therapy in the intention‐to‐treat analysis (93.7 vs 80.0%; p = .01), and the per‐protocol analysis (97.4 vs 82.0%; p = .0016). The H. pylori resistance rates to metronidazole, clarithromycin and amoxicillin were 42.1, 18.0 and 0%. Fourteen‐day therapy was significantly more effective in patients with clarithromycin‐resistant strains. Incidences of adverse events were comparable. Conclusions: Addition bismuth and prolonging treatment duration can overcome H. pylori resistance to clarithromycin and decrease the bacterial load. Fourteen‐day triple therapy‐based, bismuth‐containing quadruple therapy achieved ITT success rate 93% and could be recommended as the first line eradication regimen.     

Comparison of Linear,Nonlinear and Semiparametric Mixed‐effects Models for Estimating HIV Dynamic Parameters

The potency of antiretroviral agents in AIDS clinical trials can be assessed on the basis of an early viral response such as viral decay rate or change in viral load (number of copies of HIV RNA) of the plasma. Linear, parametric nonlinear, and semiparametric nonlinear mixed‐effects models have been proposed to estimate viral decay rates in viral dynamic models. However, before applying these models to clinical data, a critical question that remains to be addressed is whether these models produce coherent estimates of viral decay rates, and if not, which model is appropriate and should be used in practice. In this paper, we applied these models to data from an AIDS clinical trial of potent antiviral treatments and found significant incongruity in the estimated rates of reduction in viral load. Simulation studies indicated that reliable estimates of viral decay rate were obtained by using the parametric and semiparametric nonlinear mixed‐effects models. Our analysis also indicated that the decay rates estimated by using linear mixed‐effects models should be interpreted differently from those estimated by using nonlinear mixed‐effects models. The semiparametric nonlinear mixed‐effects model is preferred to other models because arbitrary data truncation is not needed. Based on real data analysis and simulation studies, we provide guidelines for estimating viral decay rates from clinical data. (© 2004 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

High pH reversed‐phase chromatography as a superior fractionation scheme compared to off‐gel isoelectric focusing for complex proteome analysis

MS/MS is the technology of choice for analyzing complex protein mixtures. However, due to the intrinsic complexity and dynamic range present in higher eukaryotic proteomes, prefractionation is an important step to maximize the number of proteins identified. Off‐gel IEF (OG‐IEF) and high pH RP (Hp‐RP) column chromatography have both been successfully utilized as a first‐dimension peptide separation technique in shotgun proteomic experiments. Here, a direct comparison of the two methodologies was performed on ex vivo peripheral blood mononuclear cell lysate. In 12‐fraction replicate analysis, Hp‐RP resulted in more peptides and proteins identified than OG‐IEF fractionation. Distributions of peptide pIs and hydropathy did not reveal any appreciable bias in either technique. Resolution, defined here as the ability to limit a specific peptide to one particular fraction, was significantly better for Hp‐RP. This leads to a more uniform distribution of total and unique peptides for Hp‐RP across all fractions collected. These results suggest that fractionation by Hp‐RP over OG‐IEF is the better choice for typical complex proteome analysis.     

Estrogens, lipoproteins, and cardiovascular risk factors: an update following the randomized placebo-controlled trials of hormone-replacement therapy

PURPOSE OF REVIEW: The effects of hormone-replacement therapy on cardiovascular risk factors are examined. In an attempt to explain the results of recent randomized controlled trials in which no benefit of hormone-replacement therapy for postmenopausal women has been observed, RECENT FINDINGS: Changes in lipoproteins in response to hormone-replacement therapy have now been analysed for both primary and secondary prevention studies. In none of the large randomized controlled trials was there any effect of hormone-induced changes in low-density lipoprotein, high-density lipoprotein, or triglyceride on clinical outcome. Further detailed studies of lipoprotein metabolism have not revealed any adverse effect of hormone-replacement therapy. Recent analysis of the Heart Estrogen/Progestin-Replacement Study data suggests hormone-replacement therapy reduces the risk of developing diabetes. The effect of hormone-replacement therapy on inflammatory markers and on flow-mediated dilatation is largely beneficial, although the effect on flow-mediated dilatation is modulated according to endothelial function, which is adversely affected by known risk factors, including age and presence of atherosclerosis. In this respect the work on polymorphisms of estrogen receptor-alpha may in due course help to define those women who would benefit most from use of estrogen. Crucially, oral but not transdermal hormone-replacement therapy increases activated protein C resistance independently of the presence of factor V Leiden. This effect increases the risk of venous thromboembolic events, which is reflected in the results of a hospital case control study of thromboembolism. SUMMARY: Despite the outcome of the hormone-replacement therapy trials, recent work has confirmed the putative antiatherogenic effects of hormone-replacement therapy on lipoprotein metabolism. Metabolic differences of route of administration of estrogen, particularly on haemostatic variables, may explain this clinical paradox, which continues to be an important research area.