Recycling of Significance Levels in Testing Multiple Hypotheses of Confirmatory Clinical Trials

ObjectivesOur objective is to assess the influence of amicus curiae briefs on judicial behavior on the U.S. Supreme Court. Our primary hypothesis is that amicus briefs have an impact on the justices across the ideological spectrum. Our secondary hypothesis is that this influence will be greater for justices nearer the ideological center.MethodsOur analysis is confined to the Roberts Court (2005 through 2014 terms, inclusive). The unit of analysis is the justice‐vote in each of the 793 full‐opinion decisions during this 10‐term period; thus, our data set contains 7,135 observations. We employ logistic regression to test the impact of amicus filings on the ideological direction of the vote cast by each justice in each case. We control for the direction of the lower court decision, the ideological orientations of the justices, the presence of the federal government (or agency or official) as party, and the presence of the solicitor general as amicus curiae.ResultsWe find statistical support for both the primary and secondary hypotheses. Amicus briefs appear to influence the justices across the ideological spectrum. The influence is somewhat greater among the more moderate justices, although the relationship between amicus influence and judicial moderation is a weak one.ConclusionsSupreme Court justices appear to respond positively to the persuasive attempts of amici. This impact is most noticeable for the justices in the middle of the Court—those who tend to be most influential in steering the Court's decision making.

Enhancement of histaminergic neurotransmission or histaminergic plus cholinergic neurotransmission may represent novel strategies for improving cognition in Alzheimer's disease. To evaluate the effects of a novel histamine H3 receptor inverse agonist (MK-3134), an acetylcholinesterase inhibitor (donepezil), and their combination in attenuating the cognitive impairment associated with scopolamine. Thirty-one subjects were randomized, and 28 completed this double-blind, placebo-controlled, five-period crossover study. Cognition was assessed using the Groton Maze Learning Task (GMLT) as the primary outcome measure. The two primary hypotheses were that donepezil 10 mg and MK-3134 25 mg, respectively, would attenuate scopolamine (0.5 mg)-induced impairment as measured by the GMLT over the first 12 h after scopolamine administration (AUC(1-12) (h)). A secondary hypothesis was that the combination of donepezil and MK-3134 would attenuate scopolamine-induced cognitive impairment to a greater extent than either agent alone as measured by the GMLT AUC(1-12 h). The primary and secondary hypotheses were not met. Upon examining the time course of the scopolamine effects (an exploratory objective), peak effects were generally observed around 2 h after scopolamine administration. Administration of MK-3134 or donepezil improved performance on the GMLT at the 2-h time point, rather than AUC(1-12 h), compared with scopolamine alone. Moreover, it appeared that the combination of MK-3134 and donepezil blunted the scopolamine effect to a greater extent than either drug alone. Exploratory analyses provide evidence for cognitive improvement through inverse agonism of the H3 histamine receptor and for cooperation between human cholinergic and histaminergic neurotransmitter systems. (ClinicalTrials.gov trial registration number: NCT01181310).

Questionable research practices (QRPs) are intentional and unintentional practices that can occur when designing, conducting, analysing, and reporting research, producing biased study results. Sport and exercise medicine (SEM) research is...

In confirmatory clinical trials the Type I error rate must be controlled for claims forming the basis for approval and labeling of a new drug. Strong control of the familywise error rate is usually needed for hypotheses related to the primary endpoint(s). For hypotheses related to secondary endpoint(s) which are only of interest if the corresponding “parent” primary null hypotheses have been rejected, less strict error rate control might be sufficient. We review and extend procedures for families of primary and secondary hypotheses when either at least one of the primary hypotheses or all coprimary hypotheses must be rejected to claim success for the trial. Such families of hypotheses arise naturally from comparing several treatments with a control, combined noninferiority and superiority testing for primary and secondary variables, the presence of multiple primary or secondary endpoints or any combination thereof. We show that many of the procedures proposed in the literature follow a common underlying principle and in some cases can be improved. In addition we present some general results on Type I error rates for the different families and subfamilies of hypotheses and their relation to group-sequential testing of multiple hypotheses.

BackgroundMolecular taxonomy studies and barcoding projects can provide rapid means of detecting cryptic diversity. Nevertheless, the use of molecular data for species delimitation should be undertaken with caution. Especially the single-gene approaches are linked with certain pitfalls for taxonomical inference. In the present study, recent and historical species descriptions based upon morphology were used as primary species hypotheses, which were then evaluated with molecular data (including in type and historical museum material) to form secondary species hypotheses. As an example of cryptic diversity and taxonomic controversy, the European Phoxinus phoxinus species complex was used.ResultsThe results of the revision showed that of the fourteen primary species hypotheses, three were rejected, namely P. ketmaieri, P. likai, and P. apollonicus. For three species (P. strandjae, P. strymonicus, P. morella), further investigation with increased data sampling was suggested, while two primary hypotheses, P. bigerri and P. colchicus, were supported as secondary species hypotheses. Finally, six of the primary species hypotheses (P. phoxinus, P. lumaireul, P. karsticus, P. septimanae, P. marsilii and P. csikii) were well supported by mitochondrial but only limitedly corroborated by nuclear data analysis.ConclusionThe approach has proven useful for revision of species complexes, and the study can serve as an overview of the Phoxinus genus in Europe, as well as a solid basis for further work.

TPS2098 Background: Liquid biopsy in glioblastoma (GBM) is hindered by a lack of requisite circulating tumor (ct) and cell-free (cf) DNA levels in blood due to the blood-brain barrier (BBB). This limits the identification of blood-based tumor biomarkers along with the development and use of biomarker-driven systemic therapies. Real-time image-guided low intensity focused ultrasound (LIFU) combined with intravenously administered microbubble oscillators, leads to non-invasive disruption of the BBB. This trial aims to evaluate the utility of LIFU for boosting blood ctDNA and cfDNA for enhanced liquid biopsy in GBM patients. Methods: LIBERATE is an ongoing, prospective, multi-center, self-controlled, pivotal trial evaluating safety and technical efficacy of LIFU-based BBB disruption for increasing blood ctDNA and cfDNA levels in adults (aged 18-80 years) with GBM. Patients with suspected GBM planned for tumor biopsy or resection at ten centers in the US are being enrolled. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered IV microbubbles for enhanced sonication effects, after which MR-guided BBB disruption using a 220 kHz LIFU device is performed with real-time acoustic feedback control for effective microbubble activation. Before and after procedure, phlebotomy and MRI brain are performed to evaluate outcomes. Primary study endpoint is defined, per subject, as ratio between their cfDNA level in blood 1-hour post-LIFU procedure compared to cfDNA level in blood pre-procedure. Primary study hypothesis is that BBB disruption with LIFU leads to a ≥2-fold rise in blood cfDNA. The secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in blood cfDNA sample obtained 1-hour post-LIFU and the biomarker pattern in tumor tissue obtained later. The trial has been powered to evaluate both primary and secondary hypotheses. Based on an assumed true agreement rate of 91% and a one-sided alpha of 0.025, an exact test for binomial proportions provides a sample of N = 50 with 84% power for the secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of known specific somatic mutations in ctDNA from blood samples collected before and after LIFU, (2) estimation of ctDNA levels post-LIFU in samples collected at 30 minutes, 1 hour, 2 hour, and 3 hour to determine the time of greatest yield, (3) correlation of MRI parameters related to grading of BBB disruption and ctDNA-based biomarkers from post-LIFU blood samples. Patient enrollment commenced in 2022 and 22 patients have been recruited by January 2024. Clinical trial information: NCT05383872 .

BACKGROUND Liquid biopsy in glioblastoma (GBM) is hindered by insufficient requisite circulating-tumor (ct) and cell-free (cf) DNA in blood due to blood-brain barrier (BBB). This limits the identification of blood-based tumor biomarkers along with biomarker-driven systemic therapies. Real-time image-guided low-intensity focused ultrasound (LIFU) plus IV microbubble oscillators, leads to non-invasive BBB disruption. METHODS LIBERATE is an ongoing, prospective, multi-center, self-controlled, pivotal trial investigating safety and technical efficacy of LIFU-based BBB disruption for increasing blood ctDNA and cfDNA levels in adult GBM patients (aged 18-80 years). Patients with suspected GBM planned for biopsy/resection at ten US centers are being enrolled. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered IV microbubbles for enhanced sonication effects, after which MR-guided BBB disruption using 220kHz LIFU device is performed with real-time acoustic-feedback-control for effective microbubble activation. Pre- and post-LIFU, phlebotomy and MRI brain are performed. Primary study endpoint is defined, per subject, as ratio between their blood cfDNA level 1-hour post-LIFU procedure compared to blood cfDNA pre-procedure. Primary study hypothesis is that BBB disruption leads to ≥2-fold rise in blood cfDNA. Secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in blood cfDNA sample obtained 1-hour post-LIFU and biomarker pattern in tumor tissue obtained later. Trial is powered to evaluate both primary and secondary hypotheses. Based on assumed true agreement rate of 91% and one-sided alpha=0.025, an exact test for binomial proportions provides N=50 with 84% power for secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of specific somatic mutations in ctDNA from blood collected pre- and post-LIFU, (2) ctDNA estimation in samples collected at 30-minutes, 1-hour, 2-hour, and 3-hour post-LIFU to determine time of greatest yield, (3) correlation of MRI parameters related to grading of BBB disruption and ctDNA-based biomarkers from post-LIFU blood samples. Patient enrollment commenced in 2022 and remains ongoing (NCT05383872).

BACKGROUND Liquid biopsy in glioblastoma (GBM) is hindered by a lack of requisite circulating tumor (ct) and cell-free (cf) DNA levels in blood due to the blood-brain barrier (BBB). This limits the identification of blood-based tumor biomarkers along with the development and use of biomarker-driven systemic therapies. Real-time image-guided low intensity focused ultrasound (LIFU) combined with intravenously administered microbubble oscillators, leads to non-invasive disruption of the BBB. This trial aims to evaluate the utility of LIFU for boosting blood ctDNA and cfDNA for enhanced liquid biopsy in GBM patients. MATERIAL AND METHODS LIBERATE is an ongoing, prospective, multi-center, self-controlled, pivotal trial evaluating safety and technical efficacy of LIFU-based BBB disruption for increasing blood ctDNA and cfDNA levels in adults, aged 18-80 years with GBM. Patients with suspected GBM planned for tumor biopsy or resection at ten centers in the US are being enrolled. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered IV microbubbles for enhanced sonication effects, after which MR-guided BBB disruption using a 220 kHz LIFU device is performed with real-time acoustic feedback control for effective microbubble activation. Before and after procedure, phlebotomy and MRI brain are performed to evaluate outcomes. Primary study endpoint is defined, per subject, as ratio between their cfDNA level in blood 1-hour post-LIFU procedure compared to cfDNA level in blood pre-procedure. Primary study hypothesis is that BBB disruption with LIFU leads to a ≥2-fold rise in blood cfDNA. The secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in blood cfDNA sample obtained 1-hour post-LIFU and the biomarker pattern in tumor tissue obtained later. The trial has been powered to evaluate both primary and secondary hypotheses. Based on an assumed true agreement rate of 91% and a one-sided alpha of 0.025, an exact test for binomial proportions provides a sample of N = 50 with 84% power for the secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of known specific somatic mutations in ctDNA from blood samples collected before and after LIFU, (2) estimation of ctDNA levels post-LIFU in samples collected at 30 minutes, 1 hour, 2 hour, and 3 hour to determine the time of greatest yield, (3) correlation of MRI parameters related to grading of BBB disruption and ctDNA-based biomarkers from post-LIFU blood samples. RESULTS Patient enrollment commenced in 2022, and 22 patients have been recruited by 05/2024. CONCLUSION Findings from this trial will be critical to liquid biopsy in GBM (Clinical trial information: NCT05383872).

BACKGROUND Liquid biopsy in glioblastoma (GBM) is hindered by a lack of requisite circulating-free DNA (cfDNA) levels in blood due to the blood-brain barrier (BBB). This results in challenges to the identification of blood-based biomarkers and the development of novel biomarker-driven systemic therapies. Real-time image-guided low intensity focused ultrasound (LIFU) combined with intravenously administered microbubble oscillators non-invasively causes BBB disruption (BBBD). This clinical trial aimed to evaluate the utility of LIFU for increasing cfDNA in blood for liquid biopsy in GBM. METHODS LIBERATE is a prospective, multi-center, self-controlled, ongoing, pivotal trial evaluating the safety and technical efficacy of LIFU for BBBD to increase cfDNA in blood for GBM. Patients aged 18-80 years with suspected GBM planned for tumor biopsy or resection at ten centers in the US are being included. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered intravenous oscillating microbubbles for enhancing sonication, after which MR-guided BBBD using a 220 kHz LIFU device is performed with real-time acoustic feedback for effective cavitation. Before and after the procedure, phlebotomies and MRI brain are performed to evaluate outcomes. The primary study endpoint is defined, per subject, as the ratio between their cfDNA level in blood 1-hour post-LIFU procedure compared to cfDNA level in blood pre-procedure. The primary study hypothesis is that BBBD with LIFU leads to a ≥2-fold increase in cfDNA in blood. The secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in cfDNA sample from 1-hour post-LIFU sample and biomarker pattern in tumor tissue obtained later. The trial has been powered to evaluate both primary and secondary hypotheses. Based on an assumed true agreement rate of 91% and a one-sided alpha of 0.025, an exact test for binomial proportions provides a sample of N=50 with 84% power for the secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of known somatic mutations in cfDNA from blood samples collected before and after LIFU, (2) estimation of cfDNA levels post-LIFU in samples collected at 30 minutes, 1 hour, 2 hours, and 3 hours to determine the time of greatest yield, (3) correlation of MRI parameters related to grading of BBBD and biomarkers positive in cfDNA from post-LIFU blood samples. RESULTS Patient enrollment commenced in 2022, and 8 patients have been recruited by 04/11/2023. CONCLUSION Findings from this trial will be critical to liquid biopsy in GBM (Clinical trial information: NCT05383872).

TPS2084 Background: Liquid biopsy in glioblastoma (GBM) is hindered by a lack of requisite circulating-free DNA (cfDNA) levels in blood due to the blood-brain barrier (BBB). This results in challenges to the identification of blood-based biomarkers and the development of novel biomarker-driven systemic therapies. Real-time image-guided low intensity focused ultrasound (LIFU) combined with IV microbubble oscillators (DEFINITY), non-invasively causes BBB disruption (BBBD). This clinical trial aims to evaluate the utility of LIFU for increasing cfDNA in blood for liquid biopsy in GBM. Methods: LIBERATE is a prospective, multi-center, self-controlled, ongoing, pivotal trial evaluating safety and technical efficacy of LIFU for BBBD to increase cfDNA in blood for GBM. Patients aged >18-80 years with suspected GBM planned for tumor biopsy or resection at eleven centers in North America are being included. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered IV oscillating microbubbles for enhancing sonication, after which MR-guided BBBD using a 220 kHz LIFU device is performed with real-time acoustic feedback for effective cavitation. Before and after procedure, phlebotomies and MRI brain are performed to evaluate outcomes. The primary study endpoint is defined, per subject, as the ratio between their cfDNA level in blood 1-hour post-LIFU compared to cfDNA level in blood pre-procedure. The primary study hypothesis is that BBBD with LIFU leads to ≥2-fold increase in cfDNA in blood. The secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in cfDNA sample from 1-hour post-LIFU sample and biomarker pattern in tumor tissue obtained later. The trial has been powered to evaluate both primary and secondary hypotheses. Based on an assumed true agreement rate of 91% and a one-sided alpha of 0.025, an exact test for binomial proportions provides a sample of N=50 with 84% power for the secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of known somatic mutations in cfDNA from blood samples collected before and after LIFU, (2) estimation of cfDNA levels post-LIFU in samples collected at 30 minutes, 1 hour, 2 hour, and 3 hour to determine time of greatest yield, (3) correlation of MRI parameters related to grading of BBBD and biomarkers positive in cfDNA from post-LIFU blood samples. Patient enrollment commenced in 2022 and 7 patients have been recruited by 02/13/2023. Clinical trial information: NCT05383872 .

Well-designed and carefully conducted confirmatory clinical trials when successful usually provide evidence of efficacy of new treatments. Regulatory agencies normally release them for patient use if they are convinced that these treatments, in addition to exhibiting such evidence of efficacy, are also safe for patient use and provide clinically relevant benefits to patients. Trials for this purpose generally include multiple endpoints as primary and secondary for testing hypotheses on them. Primary endpoint hypotheses are considered more important; if the trial wins for at least one of them, then depending on the level of evidence desired for such a win, one can characterize clinically relevant benefits of the new treatment. The last two decades have witnessed several innovative statistical test procedures which for testing multiple primary and secondary endpoint hypotheses of a trial allow greater power of the tests. Some of these procedures are quite advanced and pioneering, but are mostly developed for fixed-sample trials without interim looks of the accumulating data of the trial. Extensions of these approaches to group sequential trials which permit interim looks for stopping the trial early for efficacy or futility are mostly new. The purpose of this book chapter is to review relevant historical methods and this new literature for group sequential trials for the demonstration of efficacy of new treatments.

ABSTRACTWe consider a confirmatory clinical trial where a primary and a secondary endpoints are tested hierarchically to control the family-wise error rate at level α. The trial uses a group sequential design with an interim and a final analysis. When the information times at the interim analysis are the same for the primary and the secondary hypotheses, it has been shown in the literature that the secondary hypothesis has to be tested using a group sequential boundary at a level no more than α. In many event-driven trials, however, the information times are usually different because of different event rates for the two endpoints. The information times may also be different for a noninferiority hypothesis and a superiority hypothesis due to different analysis sets. We consider this general setup and derive a sharp upper bound on the probability of rejecting the secondary hypothesis at an interim and a final analysis. This bound suggests that the secondary boundary can be refined so that the group sequential design can be tested at a significance level greater than α while still controlling the family-wise error rate at level α. We carry out a simulation study to illustrate the power gain by using the refined boundary for different choices of boundaries for the two hypotheses. The proposed approach is illustrated in two oncology clinical trials with more than two analyses.

Major depressive disorder (MDD) trials - investigating either non-pharmacological or pharmacological interventions - have shown mixed results. Many reasons explain this heterogeneity, but one that stands out is the trial design due to specific challenges in the field. We aimed therefore to review the methodology of non-invasive brain stimulation (NIBS) trials and provide a framework to improve clinical trial design. We performed a systematic review for randomized, controlled MDD trials whose intervention was transcranial magnetic stimulation (rTMS) or transcranial direct current stimulation (tDCS) in MEDLINE and other databases from April 2002 to April 2008. We created an unstructured checklist based on CONSORT guidelines to extract items such as power analysis, sham method, blinding assessment, allocation concealment, operational criteria used for MDD, definition of refractory depression and primary study hypotheses. Thirty-one studies were included. We found that the main methodological issues can be divided in to three groups: (1) issues related to phase II/small trials, (2) issues related to MDD trials and, (3) specific issues of NIBS studies. Taken together, they can threaten study validity and lead to inconclusive results. Feasible solutions include: estimating the sample size a priori; measuring the degree of refractoriness of the subjects; specifying the primary hypothesis and statistical tests; controlling predictor variables through stratification randomization methods or using strict eligibility criteria; adjusting the study design to the target population; using adaptive designs and exploring NIBS efficacy employing biological markers. In conclusion, our study summarizes the main methodological issues of NIBS trials and proposes a number of alternatives to manage them.

Objective. The intent of the present study was to test two hypotheses. The primary hypothesis was that there would be differences between blood serum individual free fatty acids (SIFFA) and serum individual total fatty acids (SITFA) in terms of their different relationships (correlations) to each of homeostatic model assessment-individual insulin resistance (HOMA-IR) and homeostatic model assessment-individual insulin resistance-percentage β-cell function (HOMA-% β) remaining in human type 2 diabetic patients with pre-flaxseed oil (FXO) and pre-safflower oil (SFO) administration. The secondary hypothesis was that FXO (rich in alpha-linolenic acid, ALA) supplementation would alter these correlations differently in the SIFFA and STIFFA pools in comparison with the placebo SFO (poor in ALA). Methods. Patients were recruited via a newspaper advertisement and two physicians. All patients came to visit 1 and three months later to visit 2. At visit 2, the subjects were randomly assigned (double-blind) to flaxseed or safflower oil (placebo) treatment for three months until visit 3. Results. There were pre-intervention differences in the SIFFA and STIFA pool's relationships with each of HOMA-IR and HOMA-% β. These relatioships remained either unchanged or became significant after intervention (treatment or placebo). There was a negative correlation found between HOMA-IR and serum free ALA (SFALA) mol % after FXO. Serum total ALA (STALA) mol % had no significant correlations with HOMA-IR and HOMA- % β before and after flaxseed oil administration. Conclusions. The SIFFA and SITFA pools have different relationships with HOMA-IR and HOMA-% β for each of pre- and post-intervention. It is concluded that the data support both the primary and the secondary hypotheses indicating that they are correct.

BackgroundMotion can exacerbate headache during a migraine attack, potentially leading to avoidance of routine physical activity. Advances in wrist-worn actigraphy facilitate objectively analyzing how headache episodes affect physical activity in everyday settings. The primary hypothesis was hypoactivity during daytime headache events. Secondary hypotheses are hypoactivity during the prodromal and postdromal hours closest to the headache event.MethodsDuring a 90-day prospective observational study, participants diagnosed with migraine wore an actigraphy device on their non-dominant wrist during daily life and work, while also logging migraine-related data in a dedicated smartphone application. There were no restrictions on use of acute and preventive headache treatments. Data from the wrist-worn accelerometer were used to (i) calculate activity energy expenditure, and (ii) predict types of human activities. These metrics were used to compare daytime prodromal, ictal, and postdromal phases of headache events with time-matched intervals during non-headache periods.ResultsA significant reduction in daytime physical activity was observed during the ictal phase of headache attacks, as evidenced by decreases in both activity energy expenditure and human activity recognition prediction metrics. A reduction in movement was also observed during evening hours (18:00–24:00) on headache days. However, no significant physical activity changes were noted in the prodromal and postdromal phases. Reduced physical activity was more pronounced during the ictal phase when acute treatments were ineffective.ConclusionsThis study is the first to examine the impact of headache on physical activity levels during daytime headache events by assessing changes in daily activities and activity energy expenditure in individuals with migraine, within their habitual environments and without restrictions on acute medication use. Our findings confirm reduced movement during the ictal phase of migraine attacks, supporting the primary hypothesis. Wrist-worn actigraphy further indicated that this reduction is more pronounced when patients experience movement sensitivity. Evening hypoactivity is also observed on headache days. Furthermore, attacks with ineffective acute treatment or moderate-to-high intensity were associated with more pronounced reductions in movement. In contrast, our data did not support the secondary hypothesis that physical activity would decrease during daytime prodromal and postdromal periods.Trial registrationNCT04983186 (www.ClinicalTrials.gov).Graphical