The tyranny of non-inferiority trials.

Opportunities to decrease the toxicity and cost of approved treatment regimens with lower dose, less frequent, or shorter duration alternative regimens have been limited by the perception that alternatives must be non-inferior to approved regimens. Non-inferiority trials are large and expensive to do, because they must show statistically that the alternative and approved therapies differ in a single outcome, by a margin far smaller than that required to demonstrate superiority. Non-inferiority's flaws are manifest: it ignores variability expected to occur with repeated evaluation of the approved therapy, fails to recognise that a trial of similar design will be labelled as superiority or non-inferiority depending on whether it is done prior to or after initial registration of the approved treatment, and relegates endpoints such as toxicity and cost. For example, while a less toxic and less costly regimen of 3 months duration would typically be required to demonstrate efficacy that is non-inferior to that of a standard regimen of 6 months to displace it, the longer duration therapy has no such obligation to prove its superiority. This situation is the tyranny of the non-inferiority trial: its statistics perpetuate less cost-effective regimens, which are not patient-centred, even when less intensive therapies confer survival benefits nearly identical to those of the standard, by placing a disproportionately large burden of proof on the alternative. This approach is illogical. We propose that the designation of trials as superiority or non-inferiority be abandoned, and that randomised, controlled trials should henceforth be described simply as "comparative".

Evolution of heart transplant donor characteristics in the 21st century: A United States single center's experience.

Despite a record setting number of heart transplants performed annually, the national donor shortage continues to plague transplant teams across the United States. Here we describe the barriers to adaptation of numerous "non-traditional" orthotopic heart transplant donor characteristics including donors with hepatitis C virus, those meeting criteria for donation after cardiac death, donors with coronavirus disease 19 infection, donors with the human immunodeficiency virus, and grafts with left ventricular systolic dysfunction. Our center's objective was to increase our transplant volume by expanding our donor pool from "traditional" donors to these "non-traditional" donors. We detail how medical advances such as certain laboratory studies, pharmacologic interventions, and organ care systems have allowed our center to expand the donor pool thereby increasing transplantation volume without adverse effects on outcomes.

Deficiency in the mitophagy mediator Parkin accelerates murine skin allograft rejection.

Alterations in mitochondrial function and associated quality control programs, including mitochondrial-specific autophagy, termed mitophagy, are gaining increasing recognition in the context of disease. However, the role of mitophagy in organ transplant rejection remains poorly understood. Using mice deficient in Parkin, a ubiquitin ligase that tags damaged or dysfunctional mitochondria for autophagic clearance, we assessed the impact of Parkin-dependent mitophagy on skin-graft rejection. We observed accelerated graft loss in Parkin-deficient mice across multiple skin graft models. Immune cell distributions posttransplant were largely unperturbed compared to wild-type; however, the CD8+ T cells of Parkin-deficient mice expressed more T-bet, IFNγ, and Ki67, indicating greater priming toward effector function. This was accompanied by increased circulating levels of IL-12p70 in Parkin-deficient mice. Using a mixed leukocyte reaction, we demonstrated that naïve Parkin-deficient CD4+ and CD8+ T cells exhibit enhanced activation marker expression and proliferative responses to alloantigen, which were attenuated with administration of a pharmacological mitophagy inducer (p62-mediated mitophagy inducer), known to increase mitophagy in the absence of a functional PINK1-Parkin pathway. These findings indicate a role for Parkin-dependent mitophagy in curtailing skin-graft rejection.

Increasing Multiorgan Heart Transplantations From Donation After Circulatory Death Donors in the United States.

INTRODUCTION: Donation after circulatory death (DCD) donors are becoming an important source of organs for heart-transplantation (HT), but there are limited data regarding their use in multiorgan-HT. METHODS: Between January 2020 and June 2023, we identified 87 adult multiorgan-HTs performed using DCD-donors [77 heart-kidney, 6 heart-lung, 4 heart-liver] and 1494 multiorgan-HTs using donation after brain death (DBD) donors (1141 heart-kidney, 165 heart-lung, 188 heart-liver) in UNOS. For heart-kidney transplantations (the most common multiorgan-HT combination from DCD-donors), we also compared donor/recipient characteristics, and early outcomes, including 6-month mortality using Kaplan-Meier (KM) and Cox hazards-ratio (Cox-HR). RESULTS: Use of DCD-donors for multiorgan-HTs in the United States increased from 1% in January to June 2020 to 12% in January-June 2023 (p < 0.001); but there was a wide variation across UNOS regions and center volumes. Compared to recipients of DBD heart-kidney transplantations, recipients of DCD heart-kidney transplantations were less likely to be of UNOS Status 1/2 at transplant (35.06% vs. 69.59%) and had lower inotrope use (22.08% vs. 43.30%), lower IABP use (2.60% vs. 26.29%), but higher durable CF-LVAD use (19.48% vs. 12.97%), all p < 0.01. Compared to DBD-donors, DCD-donors used for heart-kidney transplantations were younger [28(22-34) vs. 32(25-39) years, p = 0.004]. Recipients of heart-kidney transplantations from DCD-donors and DBD-donors had similar 6-month survival using both KM analysis, and unadjusted and adjusted Cox-HR models, including in propensity matched cohorts. Rates of PGF and in-hospital outcomes were also similar. CONCLUSIONS: Use of DCD-donors for multiorgan-HTs has increased rapidly in the United States and early outcomes of DCD heart-kidney transplantations are promising.

Paradoxical SERCA dysregulation contributes to atrial fibrillation in a model of diet-induced obesity.

Obesity is a major risk factor for atrial fibrillation (AF) the most common serious cardiac arrhythmia, but the molecular mechanisms underlying diet-induced AF remain unclear. In this study, we subjected mice to a chronic high-fat diet and acute sympathetic activation ('two-hit' model) to study the mechanisms by which diet-induced obesity promotes AF. Surface electrocardiography revealed that diet-induced obesity and sympathetic activation synergize during intracardiac tachypacing to induce AF. At the cellular level, diet-induced obesity and acute adrenergic stimulation facilitate the formation of delayed afterdepolarizations in atrial myocytes, implicating altered Ca2+ dynamics as the underlying cause of AF. We found that diet-induced obesity does not alter the expression of major Ca2+-handling proteins in atria, including the sarcoplasmic reticulum Ca2+-ATPase (SERCA), a major component of beat-to-beat Ca2+ cycling in the heart. Paradoxically, obesity reduces phospholamban phosphorylation, suggesting decreased SERCA activity, yet atrial myocytes from obese mice showed a significantly increased Ca2+ transient amplitude and SERCA-mediated Ca2+ uptake. Adrenergic stimulation further increases the Ca2+ transient amplitude but does not affect Ca2+ reuptake in atrial myocytes from obese mice. Transcriptomics analysis showed that a high-fat diet prompts upregulation of neuronatin, a protein that has been implicated in obesity and is known to stimulate SERCA activity. We propose a mechanism in which obesity primes SERCA for paradoxical activation, and adrenergic stimulation facilitates AF conversion through a Ca2+-induced Ca2+ release gain in atrial myocytes. Overall, this study links obesity, altered Ca2+ signaling, and AF, and targeting this mechanism may prove effective for treating obesity-induced AF.

Lymphocyte-Directed Immunomodulation Remits Thymoma-Associated Autoimmune Pneumonitis.

BACKGROUND: Thymoma presents with several autoimmune manifestations and is associated with secondary autoimmune regulator (AIRE) deficiency. Pneumonitis has recently been described as an autoimmune manifestation associated with thymoma presenting with similar clinical, radiographic, histological, and autoantibody features as seen in patients with inherited AIRE deficiency who suffer from Autoimmune PolyEndocrinopathy-Candidiasis-Ectodermal Dystrophy (APECED) syndrome. OBJECTIVES: To treat two patients with biopsy-proven thymoma-associated pneumonitis with lymphocyte-directed immunomodulation. METHODS: Two patients with thymoma were enrolled on IRB-approved protocols at the NIH Clinical Center. We performed history and physical examination; laboratory, radiographic, histologic and pulmonary function evaluations; and measurement of the lung-directed autoantibodies KCNRG and BPIFB1 prior to and at 1- and 6-months following initiation of lymphocyte-directed immunomodulation with azathioprine with or without rituximab. RESULTS: Combination T- and B-lymphocyte-directed immunomodulation resulted in improvement of clinical, functional, and radiographic parameters at 6-month follow-up evaluations in both patients with sustained remission up to 12-36 months following treatment initiation. CONCLUSION: Lymphocyte-directed immunomodulation remitted autoimmune pneumonitis in two patients with thymoma.

Extrinsic outflow graft obstruction of the HeartMate 3 LVAD: A state-of-the-art review.

While notable improvements in survival, the incidence of hemocompatibility-related adverse events, hospitalizations, and cost have been demonstrated with the only commercially available durable left ventricular assist device, a category of pump malfunctions characterized by outflow graft obstruction has been noted with broader use and clinical follow-up of recipients of this technology. Of particular concern is the accumulation of acellular biodebris between the outflow graft and bend relief covering the outflow graft at its origin with the pump (which we term extrinsic outflow graft obstruction at the bend relief). This process tends to be insidious, occurs late in the postoperative course, can be challenging to diagnose, and can result in significant morbidity and mortality. Herein, we provide a review of this complication and outline diagnostic, treatment, and preventive strategies.

Brain Cell-based Genetic Subtyping and Drug Repositioning for Alzheimer Disease.

Alzheimer's Disease (AD) is characterized by its complex and heterogeneous etiology and gradual progression, leading to high drug failure rates in late-stage clinical trials. In order to better stratify individuals at risk for AD and discern potential therapeutic targets we employed a novel procedure utilizing cell-based co-regulated gene networks and polygenic risk scores (cbPRSs). After defining genetic subtypes using extremes of cbPRS distributions, we evaluated correlations of the genetic subtypes with previously defined AD subtypes defined on the basis of domain-specific cognitive functioning and neuroimaging biomarkers. Employing a PageRank algorithm, we identified priority gene targets for the genetic subtypes. Pathway analysis of priority genes demonstrated associations with neurodegeneration and suggested candidate drugs currently utilized in diabetes, hypertension, and epilepsy for repositioning in AD. Experimental validation utilizing human induced pluripotent stem cell (hiPSC)-derived astrocytes demonstrated the modifying effects of estradiol, levetiracetam, and pioglitazone on expression of APOE and complement C4 genes, suggesting potential repositioning for AD.

Donation After Circulatory Death Heart Transplant: Current State and Future Directions.

Orthotopic heart transplant is the gold standard therapeutic intervention for patients with end-stage heart failure. Conventionally, heart transplant has relied on donation after brain death for organ recovery. Donation after circulatory death (DCD) is the donation of the heart after confirming that circulatory function has irreversibly ceased. DCD-orthotopic heart transplant differs from donation after brain death-orthotopic heart transplant in ways that carry implications for widespread adoption, including differences in organ recovery, storage and ethical considerations surrounding normothermic regional perfusion with DCD. Despite these differences, DCD has shown promising early outcomes, augmenting the donor pool and allowing more individuals to benefit from orthotopic heart transplant. This review aims to present the current state and future trajectory of DCD-heart transplant, examine key differences between DCD and donation after brain death, including clinical experiences and innovations in methodologies, and address the ongoing ethical challenges surrounding the new frontier in heart transplant with DCD donors.

iGATE analysis improves the interpretability of single-cell immune landscape of influenza infection.

Influenza poses a persistent health burden worldwide. To design equitable vaccines effective across all demographics, it is essential to better understand how host factors such as genetic background and aging affect the single-cell immune landscape of influenza infection. Cytometry by time-of-flight (CyTOF) represents a promising technique in this pursuit, but interpreting its large, high-dimensional data remains difficult. We have developed a new analytical approach, in silico gating annotating training elucidating (iGATE), based on probabilistic support vector machine classification. By rapidly and accurately "gating" tens of millions of cells in silico into user-defined types, iGATE enabled us to track 25 canonical immune cell types in mouse lung over the course of influenza infection. Applying iGATE to study effects of host genetic background, we show that the lower survival of C57BL/6 mice compared with BALB/c was associated with a more rapid accumulation of inflammatory cell types and decreased IL-10 expression. Furthermore, we demonstrate that the most prominent effect of aging is a defective T cell response, reducing survival of aged mice. Finally, iGATE reveals that the 25 canonical immune cell types exhibited differential influenza infection susceptibility and replication permissiveness in vivo, but neither property varied with host genotype or aging. The software is available at https://github.com/UmichWenLab/iGATE.

Biomarkers from subcutaneous engineered tissues predict acute rejection of organ allografts.

Invasive graft biopsies assess the efficacy of immunosuppression through lagging indicators of transplant rejection. We report on a microporous scaffold implant as a minimally invasive immunological niche to assay rejection before graft injury. Adoptive transfer of T cells into Rag2-/- mice with mismatched allografts induced acute cellular allograft rejection (ACAR), with subsequent validation in wild-type animals. Following murine heart or skin transplantation, scaffold implants accumulate predominantly innate immune cells. The scaffold enables frequent biopsy, and gene expression analyses identified biomarkers of ACAR before clinical signs of graft injury. This gene signature distinguishes ACAR and immunodeficient respiratory infection before injury onset, indicating the specificity of the biomarkers to differentiate ACAR from other inflammatory insult. Overall, this implantable scaffold enables remote evaluation of the early risk of rejection, which could potentially be used to reduce the frequency of routine graft biopsy, reduce toxicities by personalizing immunosuppression, and prolong transplant life.

Deactivation of LVAD support for myocardial recovery-surgical perspectives.

Left ventricular assist devices (LVADs) are excellent therapies for advanced heart failure patients either bridged to transplant or for lifetime use. LVADs also allow for reverse remodeling of the failing heart that is often associated with functional improvement. Indeed, growing enthusiasm exists to better understand this population of patients, whereby the LVAD is used as an adjunct to mediate myocardial recovery. When patients achieve benchmarks suggesting that they no longer need LVAD support, questions related to the discontinuation of LVAD therapy become front and center. The purpose of this review is to provide a surgical perspective on the practical and technical issues surrounding LVAD deactivation.

The landscape of regional missense mutational intolerance quantified from 125,748 exomes.

Missense variants can have a range of functional impacts depending on factors such as the specific amino acid substitution and location within the gene. To interpret their deleteriousness, studies have sought to identify regions within genes that are specifically intolerant of missense variation 1-12 . Here, we leverage the patterns of rare missense variation in 125,748 individuals in the Genome Aggregation Database (gnomAD) 13 against a null mutational model to identify transcripts that display regional differences in missense constraint. Missense-depleted regions are enriched for ClinVar 14 pathogenic variants, de novo missense variants from individuals with neurodevelopmental disorders (NDDs) 15,16 , and complex trait heritability. Following ClinGen calibration recommendations for the ACMG/AMP guidelines, we establish that regions with less than 20% of their expected missense variation achieve moderate support for pathogenicity. We create a missense deleteriousness metric (MPC) that incorporates regional constraint and outperforms other deleteriousness scores at stratifying case and control de novo missense variation, with a strong enrichment in NDDs. These results provide additional tools to aid in missense variant interpretation.

The Scalable Variant Call Representation: Enabling Genetic Analysis Beyond One Million Genomes.

The Variant Call Format (VCF) is widely used in genome sequencing but scales poorly. For instance, we estimate a 150,000 genome VCF would occupy 900 TiB, making it both costly and complicated to produce and analyze. The issue stems from VCF's requirement to densely represent both reference-genotypes and allele-indexed arrays. These requirements lead to unnecessary data duplication and, ultimately, very large files. To address these challenges, we introduce the Scalable Variant Call Representation (SVCR). This representation reduces file sizes by ensuring they scale linearly with samples. SVCR achieves this by adopting reference blocks from the Genomic Variant Call Format (GVCF) and employing local allele indices. SVCR is also lossless and mergeable, allowing for N+1 and N+K incremental joint-calling. We present two implementations of SVCR: SVCR-VCF, which encodes SVCR in VCF format, and VDS, which uses Hail's native format. Our experiments confirm the linear scalability of SVCR-VCF and VDS, in contrast to the super-linear growth seen with standard VCF files. We also discuss the VDS Combiner, a scalable, open-source tool for producing a VDS from GVCFs and unique features of VDS which enable rapid data analysis. SVCR, and VDS in particular, ensure the scientific community can generate, analyze, and disseminate genetics datasets with millions of samples.

Use of Extracorporeal Membrane Oxygenation for Primary Graft Dysfunction After Cardiac Transplantation: Results of an A Priori Ventless Approach.

Primary graft dysfunction (PGD) after cardiac transplantation is a devastating complication with increasing frequency lately in the setting of donation after circulatory death (DCD). Severe PGD is commonly treated with extracorporeal membrane oxygenation (ECMO) using central or peripheral cannulation. We retrospectively reviewed the outcomes of PGD after cardiac transplantation requiring ECMO support at our center from 2015 to 2020, focused on our now preferential approach using peripheral cannulation without a priori venting. During the study period, 255 patients underwent heart transplantation at our center and 26 (10.2%) of them required ECMO for PGD. Of 24 patients cannulated peripherally 19 (79%) were alive at 30 days and 17 (71%) 1 year after transplant; two additional patients underwent central ECMO cannulation due to unfavorable size of femoral vessels and concern for limb ischemia. Successful decannulation with full graft function recovery occurred in 22 of 24 (92%) patients cannulated peripherally. Six of them had an indwelling intra-aortic balloon pump placed before the transplantation. None of the other 18 patients received a ventricular vent. In conclusion, the use of an a priori peripheral and ventless ECMO approach in patients with PGD after heart transplant is an effective strategy associated with high rates of graft recovery and survival.

Itaconate suppresses atherosclerosis by activating a Nrf2-dependent antiinflammatory response in macrophages in mice.

Itaconate has emerged as a critical immunoregulatory metabolite. Here, we examined the therapeutic potential of itaconate in atherosclerosis. We found that both itaconate and the enzyme that synthesizes it, aconitate decarboxylase 1 (Acod1, also known as immune-responsive gene 1 [IRG1]), are upregulated during atherogenesis in mice. Deletion of Acod1 in myeloid cells exacerbated inflammation and atherosclerosis in vivo and resulted in an elevated frequency of a specific subset of M1-polarized proinflammatory macrophages in the atherosclerotic aorta. Importantly, Acod1 levels were inversely correlated with clinical occlusion in atherosclerotic human aorta specimens. Treating mice with the itaconate derivative 4-octyl itaconate attenuated inflammation and atherosclerosis induced by high cholesterol. Mechanistically, we found that the antioxidant transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), was required for itaconate to suppress macrophage activation induced by oxidized lipids in vitro and to decrease atherosclerotic lesion areas in vivo. Overall, our work shows that itaconate suppresses atherogenesis by inducing Nrf2-dependent inhibition of proinflammatory responses in macrophages. Activation of the itaconate pathway may represent an important approach to treat atherosclerosis.

Projecting complete redaction of clinical trial protocols (RAPTURE): redacted cross sectional study.

OBJECTIVES: To characterise redactions in clinical trials and estimate a time when all protocols are fully removed (RAPTURE). DESIGN: Redacted cross sectional study. SETTING: Published phase 3 randomised controlled trials from 1 January 2010 to ██████████████. PARTICIPANTS: New England Journal of Medicine, ██████████, and Journal of the American Medical Association. MAIN OUTCOME MEASURES: █████ ████████ ██████████████ ██████ ██████████ ████████ ████████ ██████████ ███████████ ████████████ ████████████ ████████████████████████ ██████████████████ RESULTS: ████████████████████ met the inclusion criteria, with 268 (56.7%) research protocols available and accessible. The rate of redactions in protocols has increased from 0 in 2010 to 60.8% in 2021 (P<0.001). The degree of data redaction has also increased, with the average cumulative redactions among industry funded trials rising from 0 in 2010 to 3.5 pages in 2021 (P<0.001). Modelling predicts that RAPTURE is expected to occur between 2073 and 2136. Redactions featured predominantly in ████████ sponsored trials and mostly occurred in the statistical design. CONCLUSIONS: This study highlights the rise in protocol redactions and predicts that, ██████████████████████████████████████████ will be entirely redacted between 2073 and 2136. A legitimate rationale for the redactions could ███ be found. A multipronged strategy against protocol redactions is required to maintain the integrity of science. AVAILABILITY: This paper is partially redacted, but for the sake of ███████████, a version without any redactions can be found in the supplementary material.

Clonally expanded memory CD8+ T cells accumulate in atherosclerotic plaques and are pro-atherogenic in aged mice.

Aging is a strong risk factor for atherosclerosis and induces accumulation of memory CD8+ T cells in mice and humans. Biological changes that occur with aging lead to enhanced atherosclerosis, yet the role of aging on CD8+ T cells during atherogenesis is unclear. In this study, using femle mice, we found that depletion of CD8+ T cells attenuated atherogenesis in aged, but not young, animals. Furthermore, adoptive transfer of splenic CD8+ T cells from aged wild-type, but not young wild-type, donor mice significantly enhanced atherosclerosis in recipient mice lacking CD8+ T cells. We also characterized T cells in healthy and atherosclerotic young and aged mice by single-cell RNA sequencing. We found specific subsets of age-associated CD8+ T cells, including a Granzyme K+ effector memory subset, that accumulated and was clonally expanded within atherosclerotic plaques. These had transcriptomic signatures of T cell activation, migration, cytotoxicity and exhaustion. Overall, our study identified memory CD8+ T cells as therapeutic targets for atherosclerosis in aging.