HSV-1 LAT Promoter Deletion Viruses Exhibit Strain-Specific and LAT-Dependent Epigenetic Regulation of Latent Viral Genomes in Human Neurons.

Herpes simplex virus 1 (HSV-1) establishes latency in neurons and expresses long noncoding RNAs termed the latency-associated transcripts (LATs). Two previous studies using HSV-1 recombinants containing deletions in the LAT promoter revealed opposing effects of the promoter deletion regarding the heterochromatinization of latent viral genomes in mice ganglia. Confounding variables in these studies include viral strains utilized (17syn+ versus KOS), anatomical infection site (footpad versus eye) and infectious virus dose (500 versus 1 × 105 PFU), and to date the basis for the differences between the two studies remains unresolved. We recently reported that 17syn+ and KOS display distinct differences in heterochromatin levels during latency in human neurons. This raised the possibility that the discrepancy seen between the two previous studies could be explained by strain-specific differences within the LAT region. Here, we examine two recombinants containing orthologous 202 bp LAT promoter deletions, 17ΔPst and KOSΔPst, in a human neuronal model of latency and reactivation (LUHMES). We found that LUHMES neurons recapitulate previous observations in mice where deletion of the LAT promoter results in an increase in H3K27me3 deposition on the viral genome compared to the parental strain 17syn+ but a decrease compared to the parental strain KOS. We also found distinct strain-specific differences in the production of viral transcripts and proteins during latency. These results indicate that the function and/or regulation of the LATs differs between HSV-1 strains and may shed light on some discrepancies found in the literature when examining the function of the LATs. IMPORTANCE Herpes simplex virus 1 (HSV-1) establishes a lifelong infection in neuronal cells. Periodically, the virus reactivates from this latent state and causes recurrent disease. Mechanisms that control entry into and maintenance of latency are not well understood, though epigenetic posttranslational modification of histones associated with the viral genome are known to play an important role. During latency, the latency-associated transcript (LAT) is known to impact epigenetic marks, but the ultimate effect has been a point of controversy. Here, we utilize a human neuronal cell line model of HSV latency and reactivation (LUHMES) to characterize latency for two HSV-1 wild-type strains and their respective LAT promoter deletion viruses. We find that the LAT acts in a strain-specific manner to influence levels of chromatin marks, viral transcription, and viral protein production. This work highlights the need to account for strain-specific differences when characterizing the LAT's function and the dynamics of reactivation.

A viral lncRNA tethers HSV-1 genomes at the nuclear periphery to establish viral latency.

IMPORTANCE: Herpes simplex virus 1 (HSV-1) establishes lifelong latency in neuronal cells. Following a stressor, the virus reactivates from latency, virus is shed at the periphery and recurrent disease can occur. During latency, the viral lncRNA termed the latency-associated transcript (LAT) is known to accumulate to high abundance. The LAT is known to impact many aspects of latency though the molecular events involved are not well understood. Here, we utilized a human neuronal cell line model of HSV latency and reactivation (LUHMES) to identify the molecular-binding partners of the LAT during latency. We found that the LAT binds to both the cellular protein, TMEM43, and HSV-1 genomes in LUHMES cells. Additionally, we find that knockdown of TMEM43 prior to infection results in a decreased ability of HSV-1 to establish latency. This work highlights a potential mechanism for how the LAT facilitates the establishment of HSV-1 latency in human neurons.

Treatment of infection and inflammation associated with COVID-19, multi-drug resistant pneumonia and fungal sinusitis by nebulizing a nanosilver solution.

Solutions containing Ag0 nanoclusters, Ag+1, and higher oxidation state silver, generated from nanocrystalline silver dressings, were anti-inflammatory against porcine skin inflammation. The dressings have clinically-demonstrated broad-spectrum antimicrobial activity, suggesting application of nanosilver solutions in treating pulmonary infection. Nanosilver solutions were tested for antimicrobial efficacy; against HSV-1 and SARS-CoV-2; and nebulized in rats with acute pneumonia. Patients with pneumonia (ventilated), fungal sinusitis, burns plus COVID-19, and two non-hospitalized patients with COVID-19 received nebulized nanosilver solution. Nanosilver solutions demonstrated pH-dependent antimicrobial efficacy; reduced infection and inflammation without evidence of lung toxicity in the rat model; and inactivated HSV-1 and SARS-CoV-2. Pneumonia patients had rapidly reduced pulmonary symptoms, recovering pre-illness respiratory function. Fungal sinusitis-related inflammation decreased immediately with infection clearance within 21 days. Non-hospitalized patients with COVID-19 experienced rapid symptom remission. Nanosilver solutions, due to anti-inflammatory, antiviral, and antimicrobial activity, may be effective for treating respiratory inflammation and infections caused by viruses and/or microbes.

Herpes Simplex Virus 1 Strains 17syn+ and KOS(M) Differ Greatly in Their Ability To Reactivate from Human Neurons In Vitro.

Herpes simplex virus 1 (HSV-1) establishes a lifelong latent infection in peripheral nerve ganglia. Periodically, the virus reactivates from this latent reservoir and is transported to the original site of infection. Strains of HSV-1 have been noted to vary greatly in their virulence and reactivation efficiencies in animal models. While HSV-1 strain 17syn+ can be readily reactivated, strain KOS(M) shows little to no reactivation in the mouse and rabbit models of induced reactivation. Additionally, 17syn+ is markedly more virulent in vivo than KOS. This has raised questions regarding potential strain-specific differences in neuroinvasion and neurovirulence and their contribution to differences in the establishment of latency (or ability to spread back to the periphery) and to the reactivation phenotype. To determine if any difference in the ability to reactivate between strains 17syn+ and KOS(M) is manifest at the level of neurons, we utilized a recently characterized human neuronal cell line model of HSV latency and reactivation (LUHMES). We found that KOS(M) established latency with a higher number of viral genomes than strain 17syn+ Strikingly, we show that the KOS(M) viral genomes have a higher burden of heterochromatin marks than strain 17syn+ The increased heterochromatin profile for KOS(M) correlates with the reduced expression of viral lytic transcripts during latency and impaired induced reactivation compared to that of 17syn+ These results suggest that genomes entering neurons from HSV-1 infections with strain KOS(M) are more prone to rapid heterochromatinization than those of 17syn+ and that this results in a reduced ability to reactivate from latency.IMPORTANCE Herpes simplex virus 1 (HSV-1) establishes a lifelong infection in neuronal cells. The virus periodically reactivates and causes recurrent disease. Strains of HSV-1 vary greatly in their virulence and potential to reactivate in animal models. Although these differences are phenotypically well defined, factors contributing to the strains' abilities to reactivate are largely unknown. We utilized a human neuronal cell line model of HSV latency and reactivation (LUHMES) to characterize the latent infection of two HSV-1 wild-type strains. We find that strain-specific differences in reactivation are recapitulated in LUHMES. Additionally, these differences correlate with the degree of heterochromatinization of the latent genomes. Our data suggest that the epigenetic state of the viral genome is an important determinant of reactivation that varies in a strain-specific manner. This work also shows the first evidence of strain-specific differences in reactivation outside the context of the whole animal at a human neuronal cell level.

Creating and implementing a COVID-19 recruitment Data Mart.

The COVID-19 pandemic has resulted in an unprecedented strain on every aspect of the healthcare system, and clinical research is no exception. Researchers are working against the clock to ramp up research studies addressing every angle of COVID-19 - gaining a better understanding of person-to-person transmission, improving methods for diagnosis, and developing therapies to treat infection and vaccines to prevent it. The impact of the virus on research efforts is not limited to investigators and their teams. Potential participants also face unparalleled opportunities and requests to participate in research, which can result in a significant amount of participant fatigue. The Vanderbilt Institute for Clinical and Translational Research recognized early in the pandemic that a solution to assist researchers in the rapid identification of potential participants was critical, and thus developed the COVID-19 Recruitment Data Mart. This solution does not rest solely on technology; the addition of experienced project managers to support researchers and facilitate collaboration was essential. Since the platform and study support tools were launched on July 20, 2020, four studies have been onboarded and a total of 1693 potential participant matches have been shared. Each of these patients had agreed in advance to direct contact for COVID-19 research and had been matched to study-specific inclusion/exclusion criteria. Our innovative Data Mart system is scalable and looks promising as a generalizable solution for simultaneously recommending individuals from a pool of patients against a pool of time-sensitive trial opportunities.

Lund Human Mesencephalic (LUHMES) Neuronal Cell Line Supports Herpes Simplex Virus 1 Latency In Vitro.

Lund human mesencephalic (LUHMES) cells are human embryonic neuronal precursor cells that can be maintained as proliferating cells due to the expression of a tetracycline-regulatable (Tet-off) v-myc transgene. They can be differentiated to postmitotic neurons by the addition of tetracycline, glial cell-derived neurotrophic factor (GDNF), and dibutyryl cAMP. We demonstrate that these cells can be infected with herpes simplex virus 1 (HSV-1) at a multiplicity of infection (MOI) of 3 with the majority of cells surviving. By 6 days postinfection, there is a loss of lytic gene transcription and an increase in the numbers of neurons that express the latency-associated transcripts (LATs). Importantly, the virus can then be reactivated by the addition of a phosphoinositide 3-kinase inhibitor, which has previously been shown to reactivate HSV-1 in rat neuron cultures. While rodent primary culture neuron systems have been described, these are limited by their lack of scalability, as it is difficult to obtain more than 500,000 neurons to employ for a given experiment. Several recent papers have described a human dorsal root ganglion (DRG) neuron culture model and human induced pleuripotent stem cell (iPSC) neuron culture models that are scalable, but they require that the presence of an antiviral suppression be maintained following HSV-1 infection. The human LUHMES cell model of HSV-1 infection described here may be especially useful for studying HSV-1 latency and reactivation on account of its scalability, its amenability to maintenance of latency without the continual use of antiviral inhibitors, and its latent gene expression profile which mirrors many properties observed in vivo, importantly, the heterogeneity of cells expressing the LATs.IMPORTANCE Herpes simplex virus (HSV) is responsible for significant morbidity in humans due to its ability to cause oral and genital lesions, ocular disease, and encephalitis. While antivirals can attenuate the severity and frequency of disease, there is no vaccine or cure. Understanding the molecular details of HSV latency and reactivation is key to the development of new therapies. One of the difficulties in studying HSV latency has been the need to rely on establishment of latent infections in animal models. While rodent primary neuron culture models have shown promise, they yield relatively small numbers of latently infected neurons for biochemical and molecular analyses. Here we present the use of a human central nervous system (CNS)-derived conditionally proliferating cell line that can be differentiated into mature neurons and latently infected with HSV-1. This model shows promise as a scalable tool to study molecular and biochemical aspects of HSV-1 latency and reactivation in human neurons.

The CCCTC Binding Factor, CTRL2, Modulates Heterochromatin Deposition and the Establishment of Herpes Simplex Virus 1 Latency In Vivo.

The cellular insulator protein CTCF plays a role in herpes simplex virus 1 (HSV-1) latency through the establishment and regulation of chromatin boundaries. We previously found that the CTRL2 regulatory element downstream from the latency-associated transcript (LAT) enhancer was bound by CTCF during latency and underwent CTCF eviction at early times postreactivation in mice latently infected with 17syn+ virus. We also showed that CTRL2 was a functional enhancer-blocking insulator in both epithelial and neuronal cell lines. We hypothesized that CTRL2 played a direct role in silencing lytic gene expression during the establishment of HSV-1 latency. To test this hypothesis, we used a recombinant virus with a 135-bp deletion spanning only the core CTRL2 insulator domain (ΔCTRL2) in the 17syn+ background. Deletion of CTRL2 resulted in restricted viral replication in epithelial cells but not neuronal cells. Following ocular infection, mouse survival decreased in the ΔCTRL2-infected cohort, and we found a significant decrease in the number of viral genomes in mouse trigeminal ganglia (TG) infected with ΔCTRL2, indicating that the CTRL2 insulator was required for the efficient establishment of latency. Immediate early (IE) gene expression significantly increased in the number of ganglia infected with ΔCTRL2 by 31 days postinfection relative to the level with 17syn+ infection, indicating that deletion of the CTRL2 insulator disrupted the organization of chromatin domains during HSV-1 latency. Finally, chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) analyses of TG from ΔCTRL2-infected mice confirmed that the distribution of the repressive H3K27me3 (histone H3 trimethylated at K27) mark on the ΔCTRL2 recombinant genomes was altered compared to that of the wild type, indicating that the CTRL2 site modulates the repression of IE genes during latency.IMPORTANCE It is becoming increasingly clear that chromatin insulators play a key role in the transcriptional control of DNA viruses. The gammaherpesviruses Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV) utilize chromatin insulators to order protein recruitment and dictate the formation of three-dimensional DNA loops that spatially control transcription and latency. The contribution of chromatin insulators in alphaherpesvirus transcriptional control is less well understood. The work presented here begins to bridge that gap in knowledge by showing how one insulator site in HSV-1 modulates lytic gene transcription and heterochromatin deposition as the HSV-1 genome establishes latency.

Modeling Herpes Simplex Virus 1 Infections in Human Central Nervous System Neuronal Cells Using Two- and Three-Dimensional Cultures Derived from Induced Pluripotent Stem Cells.

Herpes simplex virus 1 (HSV-1) establishes latency in both peripheral nerve ganglia and the central nervous system (CNS). The outcomes of acute and latent infections in these different anatomic sites appear to be distinct. It is becoming clear that many of the existing culture models using animal primary neurons to investigate HSV-1 infection of the CNS are limited and not ideal, and most do not recapitulate features of CNS neurons. Human induced pluripotent stem cells (hiPSCs) and neurons derived from them are documented as tools to study aspects of neuropathogenesis, but few have focused on modeling infections of the CNS. Here, we characterize functional two-dimensional (2D) CNS-like neuron cultures and three-dimensional (3D) brain organoids made from hiPSCs to model HSV-1-human-CNS interactions. Our results show that (i) hiPSC-derived CNS neurons are permissive for HSV-1 infection; (ii) a quiescent state exhibiting key landmarks of HSV-1 latency described in animal models can be established in hiPSC-derived CNS neurons; (iii) the complex laminar structure of the organoids can be efficiently infected with HSV, with virus being transported from the periphery to the central layers of the organoid; and (iv) the organoids support reactivation of HSV-1, albeit less efficiently than 2D cultures. Collectively, our results indicate that hiPSC-derived neuronal platforms, especially 3D organoids, offer an extraordinary opportunity for modeling the interaction of HSV-1 with the complex cellular and architectural structure of the human CNS.IMPORTANCE This study employed human induced pluripotent stem cells (hiPSCs) to model acute and latent HSV-1 infections in two-dimensional (2D) and three-dimensional (3D) CNS neuronal cultures. We successfully established acute HSV-1 infections and infections showing features of latency. HSV-1 infection of the 3D organoids was able to spread from the outer surface of the organoid and was transported to the interior lamina, providing a model to study HSV-1 trafficking through complex neuronal tissue structures. HSV-1 could be reactivated in both culture systems; though, in contrast to 2D cultures, it appeared to be more difficult to reactivate HSV-1 in 3D cultures, potentially paralleling the low efficiency of HSV-1 reactivation in the CNS of animal models. The reactivation events were accompanied by dramatic neuronal morphological changes and cell-cell fusion. Together, our results provide substantive evidence of the suitability of hiPSC-based neuronal platforms to model HSV-1-CNS interactions in a human context.

The ATM and Rad3-Related (ATR) Protein Kinase Pathway Is Activated by Herpes Simplex Virus 1 and Required for Efficient Viral Replication.

The ATM and Rad3-related (ATR) protein kinase and its downstream effector Chk1 are key sensors and organizers of the DNA damage response (DDR) to a variety of insults. Previous studies of herpes simplex virus 1 (HSV-1) showed no evidence for activation of the ATR pathway. Here we demonstrate that both Chk1 and ATR were phosphorylated by 3 h postinfection (h.p.i.). Activation of ATR and Chk1 was observed using 4 different HSV-1 strains in multiple cell types, while a specific ATR inhibitor blocked activation. Mechanistic studies point to early viral gene expression as a key trigger for ATR activation. Both pATR and pChk1 localized to the nucleus within viral replication centers, or associated with their periphery, by 3 h.p.i. Significant levels of pATR and pChk1 were also detected in the cytoplasm, where they colocalized with ICP4 and ICP0. Proximity ligation assays confirmed that pATR and pChk1 were closely and specifically associated with ICP4 and ICP0 in both the nucleus and cytoplasm by 3 h.p.i., but not with ICP8 or ICP27, presumably in a multiprotein complex. Chemically distinct ATR and Chk1 inhibitors blocked HSV-1 replication and infectious virion production, while inhibitors of ATM, Chk2, and DNA-dependent protein kinase (DNA-PK) did not. Together our data show that HSV-1 activates the ATR pathway at early stages of infection and that ATR and Chk1 kinase activities play important roles in HSV-1 replication fitness. These findings indicate that the ATR pathway may provide insight for therapeutic approaches.IMPORTANCE Viruses have evolved complex associations with cellular DNA damage response (DDR) pathways, which sense troublesome DNA structures formed during infection. The first evidence for activation of the ATR pathway by HSV-1 is presented. ATR is activated, and its downstream target Chk1 is robustly phosphorylated, during early stages of infection. Both activated proteins are found in the nucleus associated with viral replication compartments and in the cytoplasm associated with viral proteins. We also demonstrate that both ATR and Chk1 kinase activities are important for viral replication. The findings suggest that HSV-1 activates ATR and Chk1 during early stages of infection and utilizes the enzymes to promote its own replication. The observation may be exploitable for antiviral approaches.

Human papillomavirus episome stability is reduced by aphidicolin and controlled by DNA damage response pathways.

A highly reproducible quantitative PCR (Q-PCR) assay was used to study the stability of human papillomavirus (HPV) in undifferentiated keratinocytes that maintain viral episomes. The term "stability" refers to the ability of episomes to persist with little copy number variation in cells. In investigating the mechanism of action of PA25, a previously published compound that destabilizes HPV episomes, aphidicolin was also found to markedly decrease episome levels, but via a different pathway from that of PA25. Since aphidicolin is known to activate DNA damage response (DDR) pathways, effects of inhibitors and small interfering RNAs (siRNAs) acting within DDR pathways were investigated. Inhibitors of Chk1 and siRNA directed against ataxia-telangiectasia mutated (ATM) and ataxia-telangiectasia Rad3-related (ATR) pathways significantly reduced viral episomes, suggesting that these pathways play a role in maintaining HPV episome stability. Inhibitors of Chk2 and DNA-PK had no effect on episome levels. Pharmacological inhibition of ATM proteins had no effect on episome levels, but ATM knockdown by siRNA significantly reduced episome levels, suggesting that ATM proteins are playing an important role in HPV episome stability that does not require kinase activity. These results outline two pathways that trigger episome loss from cells and suggest the existence of a little-understood mechanism that mediates viral DNA elimination. Together, our results also indicate that HPV episomes have a stability profile that is remarkably similar to that of fragile sites; these similarities are outlined and discussed. This close correspondence may influence the preference of HPV for integration into fragile sites.

Impact of financial compensation on enrollment and participation in a remote, mobile-app based research study.

Background: There is no consensus on how to determine appropriate financial compensation for research recruitment. Selecting incentive amounts that are reasonable and respectful, without undue inducement, remains challenging. Previously, we demonstrated that incentive amount significantly impacts participants' willingness to complete various hypothetical research activities. Here we further explore this relationship in a mock decentralized study. Methods: Adult ResearchMatch volunteers were invited to join a prospective study where interested individuals were given an opportunity to view details for a study along with participation requirements, then offered a randomly generated compensation amount between $0 and $50 to enroll and participate. Individuals agreeing to participate were then asked to complete tasks using a remote mobile application (MyCap), for two weeks. Tasks included a weekly survey, a daily gratitude journal and daily phone tapping task. Results: Willingness to participate was 85% across all incentive levels but not significantly impacted by amount. Task completion appeared to increase as a function of compensation until a plateau at $25. While participants described the study as low burden and reported that compensation was moderately important to their decision to join, only 31% completed all study tasks. Conclusion: While offering compensation in this study did not have a strong effect on enrollment rate, this work provides insight into participant motivation when joining and participating in studies employing mobile applications.

Taking AIM at serious illness: implementing an access to investigational medicines expanded access program.

When seriously ill patients have exhausted all treatment options available as part of usual care, the use of investigational agents may be warranted. Food and Drug Administration's (FDA) Expanded Access (EA) pathway provides a mechanism for these patient's physicians to pursue use of an investigational agent outside of a clinical trial when trial enrollment is not a feasible option. Though FDA has recently implemented processes to significantly streamline the regulatory portion of the process, the overall pathway has several time-consuming components including communication with the pharmaceutical company and the associated institutional requirements for EA use (contracting, Institutional Review Board [IRB], pharmacy, billing). Here, we present our experience building infrastructure at the Vanderbilt University Medical Center (VUMC) to support physicians and patients in pursuing EA, called the Access to Investigational Medicines (AIM) Platform, aligning the needs and responsibilities of institutional stakeholders and streamlining to ensure efficiency and regulatory compliance. Since its launch, the AIM team has experienced steady growth, supporting 40 EA cases for drugs/biologics, including both single patient cases and intermediate-size EA protocols in the emergent and non-emergent setting. As the EA pathway is a complex process that requires expert facilitation, we propose prioritizing EA support infrastructure at major academic medical centers as an essential regulatory knowledge function.

Quantitative assessment of the impact of standard agreement templates on multisite clinical trial start up time.

Contracting delays remain a challenge to the successful initiation of multisite clinical research in the US. The Clinical and Translational Science Awards (CTSA) Contracts Processing Study showed average contract negotiation duration of > 100 days for industry-sponsored or investigator-initiated contracts. Such delays create enormous costs to sponsors and to patients waiting to use new evidence-based treatments. With support from the National Institutes of Health's National Center for Advancing Translational Sciences, the Accelerated Clinical Trial Agreement (ACTA) was developed by 25 major academic institutions and medical centers engaged in clinical research in collaboration with the University-Industry Demonstration Partnership and with input from pharmaceutical companies. The ACTA also informed the development of subsequent agreements, including the Federal Demonstration Partnership Clinical Trial Subaward Agreement (FDP-CTSA); both ACTA and the FDP-CTSA are largely non-negotiable agreements that represent pre-negotiated compromises in contract terms agreed upon by industry and/or medical center stakeholders. When the involved parties agree to use the CTSA-developed and supported standard agreement templates as a starting point for negotiations, there can be significant time savings for trials. Use of the ACTA resulted in an average savings of 48 days and use of the FDP-CTSA saved an average of 57 days of negotiation duration.

Using a multicultural and multilingual awareness-raising strategy to enhance enrollment of racially underrepresented minoritized communities - the PassITON trial.

Racially and ethnically minoritized populations have been historically excluded and underrepresented in research. This paper will describe best practices in multicultural and multilingual awareness-raising strategies used by the Recruitment Innovation Center to increase minoritized enrollment into clinical trials. The Passive Immunity Trial for Our Nation will be used as a primary example to highlight real-world application of these methods to raise awareness, engage community partners, and recruit diverse study participants.

The RIC Recruitment & Retention Materials Toolkit - a resource for developing community-informed study materials.

Clinical trials face many challenges with meeting projected enrollment and retention goals. A study's recruitment materials and messaging convey necessary key information and therefore serve as a critical first impression with potential participants. Yet study teams often lack the resources and skills needed to develop engaging, culturally tailored, and professional-looking recruitment materials. To address this gap, the Recruitment Innovation Center recently developed a Recruitment & Retention Materials Content and Design Toolkit, which offers research teams guidance, actionable tips, resources, and customizable templates for creating trial-specific study materials. This paper seeks to describe the creation and contents of this new toolkit.

Approaches for enhancing the informativeness and quality of clinical trials: Innovations and principles for implementing multicenter trials from the Trial Innovation Network.

One challenge for multisite clinical trials is ensuring that the conditions of an informative trial are incorporated into all aspects of trial planning and execution. The multicenter model can provide the potential for a more informative environment, but it can also place a trial at risk of becoming uninformative due to lack of rigor, quality control, or effective recruitment, resulting in premature discontinuation and/or non-publication. Key factors that support informativeness are having the right team and resources during study planning and implementation and adequate funding to support performance activities. This communication draws on the experience of the National Center for Advancing Translational Science (NCATS) Trial Innovation Network (TIN) to develop approaches for enhancing the informativeness of clinical trials. We distilled this information into three principles: (1) assemble a diverse team, (2) leverage existing processes and systems, and (3) carefully consider budgets and contracts. The TIN, comprised of NCATS, three Trial Innovation Centers, a Recruitment Innovation Center, and 60+ CTSA Program hubs, provides resources to investigators who are proposing multicenter collaborations. In addition to sharing principles that support the informativeness of clinical trials, we highlight TIN-developed resources relevant for multicenter trial initiation and conduct.

Decentralized clinical trials in the trial innovation network: Value, strategies, and lessons learned.

New technologies and disruptions related to Coronavirus disease-2019 have led to expansion of decentralized approaches to clinical trials. Remote tools and methods hold promise for increasing trial efficiency and reducing burdens and barriers by facilitating participation outside of traditional clinical settings and taking studies directly to participants. The Trial Innovation Network, established in 2016 by the National Center for Advancing Clinical and Translational Science to address critical roadblocks in clinical research and accelerate the translational research process, has consulted on over 400 research study proposals to date. Its recommendations for decentralized approaches have included eConsent, participant-informed study design, remote intervention, study task reminders, social media recruitment, and return of results for participants. Some clinical trial elements have worked well when decentralized, while others, including remote recruitment and patient monitoring, need further refinement and assessment to determine their value. Partially decentralized, or "hybrid" trials, offer a first step to optimizing remote methods. Decentralized processes demonstrate potential to improve urban-rural diversity, but their impact on inclusion of racially and ethnically marginalized populations requires further study. To optimize inclusive participation in decentralized clinical trials, efforts must be made to build trust among marginalized communities, and to ensure access to remote technology.

Development, implementation, and dissemination of operational innovations across the trial innovation network.

Improving the quality and conduct of multi-center clinical trials is essential to the generation of generalizable knowledge about the safety and efficacy of healthcare treatments. Despite significant effort and expense, many clinical trials are unsuccessful. The National Center for Advancing Translational Science launched the Trial Innovation Network to address critical roadblocks in multi-center trials by leveraging existing infrastructure and developing operational innovations. We provide an overview of the roadblocks that led to opportunities for operational innovation, our work to develop, define, and map innovations across the network, and how we implemented and disseminated mature innovations.

The Trial Innovation Network Liaison Team: building a national clinical and translational community of practice.

In 2016, the National Center for Advancing Translational Science launched the Trial Innovation Network (TIN) to address barriers to efficient and informative multicenter trials. The TIN provides a national platform, working in partnership with 60+ Clinical and Translational Science Award (CTSA) hubs across the country to support the design and conduct of successful multicenter trials. A dedicated Hub Liaison Team (HLT) was established within each CTSA to facilitate connection between the hubs and the newly launched Trial and Recruitment Innovation Centers. Each HLT serves as an expert intermediary, connecting CTSA Hub investigators with TIN support, and connecting TIN research teams with potential multicenter trial site investigators. The cross-consortium Liaison Team network was developed during the first TIN funding cycle, and it is now a mature national network at the cutting edge of team science in clinical and translational research. The CTSA-based HLT structures and the external network structure have been developed in collaborative and iterative ways, with methods for shared learning and continuous process improvement. In this paper, we review the structure, function, and development of the Liaison Team network, discuss lessons learned during the first TIN funding cycle, and outline a path toward further network maturity.

Leveraging the Expertise of the CTSA Program to Increase the Impact and Efficiency of Clinical Trials.

Importance: Multicenter clinical trials play a critical role in the translational processes that enable new treatments to reach all people and improve public health. However, conducting multicenter randomized clinical trials (mRCT) presents challenges. The Trial Innovation Network (TIN), established in 2016 to partner with the Clinical and Translational Science Award (CTSA) Consortium of academic medical institutions in the implementation of mRCTs, consists of 3 Trial Innovation Centers (TICs) and 1 Recruitment Innovation Center (RIC). This unique partnership has aimed to address critical roadblocks that impede the design and conduct of mRCTs, in expectation of accelerating the translation of novel interventions to clinical practice. The TIN's challenges and achievements are described in this article, along with examples of innovative resources and processes that may serve as useful models for other clinical trial networks providing operational and recruitment support. Observations: The TIN has successfully integrated more than 60 CTSA institution program hubs into a functional network for mRCT implementation and optimization. A unique support system for investigators has been created that includes the development and deployment of novel tools, operational and recruitment services, consultation models, and rapid communication pathways designed to reduce delays in trial start-up, enhance recruitment, improve engagement of diverse research participants and communities, and streamline processes that improve the quality, efficiency, and conduct of mRCTs. These resources and processes span the clinical trial spectrum and enable the TICs and RIC to serve as coordinating centers, data centers, and recruitment specialists to assist trials across the National Institutes of Health and other agencies. The TIN's impact has been demonstrated through its response to both historical operational challenges and emerging public health emergencies, including the national opioid public health crisis and the COVID-19 pandemic. Conclusions and Relevance: The TIN has worked to reduce barriers to implementing mRCTs and to improve mRCT processes and operations by providing needed clinical trial infrastructure and resources to CTSA investigators. These resources have been instrumental in more quickly and efficiently translating research discoveries into beneficial patient treatments.