Multiple early factors anticipate post-acute COVID-19 sequelae.

Post-acute sequelae of COVID-19 (PASC) represent an emerging global crisis. However, quantifiable risk factors for PASC and their biological associations are poorly resolved. We executed a deep multi-omic, longitudinal investigation of 309 COVID-19 patients from initial diagnosis to convalescence (2-3 months later), integrated with clinical data and patient-reported symptoms. We resolved four PASC-anticipating risk factors at the time of initial COVID-19 diagnosis: type 2 diabetes, SARS-CoV-2 RNAemia, Epstein-Barr virus viremia, and specific auto-antibodies. In patients with gastrointestinal PASC, SARS-CoV-2-specific and CMV-specific CD8+ T cells exhibited unique dynamics during recovery from COVID-19. Analysis of symptom-associated immunological signatures revealed coordinated immunity polarization into four endotypes, exhibiting divergent acute severity and PASC. We find that immunological associations between PASC factors diminish over time, leading to distinct convalescent immune states. Detectability of most PASC factors at COVID-19 diagnosis emphasizes the importance of early disease measurements for understanding emergent chronic conditions and suggests PASC treatment strategies.

Fundamental immune-oncogenicity trade-offs define driver mutation fitness.

Missense driver mutations in cancer are concentrated in a few hotspots1. Various mechanisms have been proposed to explain this skew, including biased mutational processes2, phenotypic differences3-6 and immunoediting of neoantigens7,8; however, to our knowledge, no existing model weighs the relative contribution of these features to tumour evolution. We propose a unified theoretical 'free fitness' framework that parsimoniously integrates multimodal genomic, epigenetic, transcriptomic and proteomic data into a biophysical model of the rate-limiting processes underlying the fitness advantage conferred on cancer cells by driver gene mutations. Focusing on TP53, the most mutated gene in cancer1, we present an inference of mutant p53 concentration and demonstrate that TP53 hotspot mutations optimally solve an evolutionary trade-off between oncogenic potential and neoantigen immunogenicity. Our model anticipates patient survival in The Cancer Genome Atlas and patients with lung cancer treated with immunotherapy as well as the age of tumour onset in germline carriers of TP53 variants. The predicted differential immunogenicity between hotspot mutations was validated experimentally in patients with cancer and in a unique large dataset of healthy individuals. Our data indicate that immune selective pressure on TP53 mutations has a smaller role in non-cancerous lesions than in tumours, suggesting that targeted immunotherapy may offer an early prophylactic opportunity for the former. Determining the relative contribution of immunogenicity and oncogenic function to the selective advantage of hotspot mutations thus has important implications for both precision immunotherapies and our understanding of tumour evolution.

Detection of classical Hodgkin lymphoma specific sequence in peripheral blood using a next-generation sequencing approach.

We applied a highly sensitive next-generation sequencing method to identify lymphoma-specific immunoglobulin gene segments in classical Hodgkin lymphoma (CHL) at initial diagnosis or recurrence, and assessed the ability of detecting such lymphoma-specific sequences in peripheral blood (PB). Seventeen CHL cases were tested and lymphoma-specific sequences were identified in 12 of the primary tumour biopsies. In 11 of these patients whose paired PB samples were available, tumour-specific clonotypes were detected in PB in eight patients. This data demonstrates the feasibility of detecting circulating tumour-specific sequences, creating an unprecedented opportunity to optimize the future treatment and monitoring strategies for patients with CHL.

Immunoglobulin and T cell receptor gene high-throughput sequencing quantifies minimal residual disease in acute lymphoblastic leukemia and predicts post-transplantation relapse and survival.

Minimal residual disease (MRD) quantification is an important predictor of outcome after treatment for acute lymphoblastic leukemia (ALL). Bone marrow ALL burden ≥ 10(-4) after induction predicts subsequent relapse. Likewise, MRD ≥ 10(-4) in bone marrow before initiation of conditioning for allogeneic (allo) hematopoietic cell transplantation (HCT) predicts transplantation failure. Current methods for MRD quantification in ALL are not sufficiently sensitive for use with peripheral blood specimens and have not been broadly implemented in the management of adults with ALL. Consensus-primed immunoglobulin (Ig), T cell receptor (TCR) amplification and high-throughput sequencing (HTS) permit use of a standardized algorithm for all patients and can detect leukemia at 10(-6) or lower. We applied the LymphoSIGHT HTS platform (Sequenta Inc., South San Francisco, CA) to quantification of MRD in 237 samples from 29 adult B cell ALL patients before and after allo-HCT. Using primers for the IGH-VDJ, IGH-DJ, IGK, TCRB, TCRD, and TCRG loci, MRD could be quantified in 93% of patients. Leukemia-associated clonotypes at these loci were identified in 52%, 28%, 10%, 35%, 28%, and 41% of patients, respectively. MRD ≥ 10(-4) before HCT conditioning predicted post-HCT relapse (hazard ratio [HR], 7.7; 95% confidence interval [CI], 2.0 to 30; P = .003). In post-HCT blood samples, MRD ≥10(-6) had 100% positive predictive value for relapse with median lead time of 89 days (HR, 14; 95% CI, 4.7 to 44, P < .0001). The use of HTS-based MRD quantification in adults with ALL offers a standardized approach with sufficient sensitivity to quantify leukemia MRD in peripheral blood. Use of this approach may identify a window for clinical intervention before overt relapse.

Massive evolution of the immunoglobulin heavy chain locus in children with B precursor acute lymphoblastic leukemia.

The ability to distinguish clonal B-cell populations based on the sequence of their rearranged immunoglobulin heavy chain (IgH) locus is an important tool for diagnosing B-cell neoplasms and monitoring treatment response. Leukemic precursor B cells may continue to undergo recombination of the IgH gene after malignant transformation; however, the magnitude of evolution at the IgH locus is currently unknown. We used next-generation sequencing to characterize the repertoire of IgH sequences in diagnostic samples of 51 children with B precursor acute lymphoblastic leukemia (B-ALL). We identified clonal IgH rearrangements in 43 of 51 (84%) cases and found that the number of evolved IgH sequences per patient ranged dramatically from 0 to 4024. We demonstrate that the evolved IgH sequences are not the result of amplification artifacts and are unique to leukemic precursor B cells. In addition, the evolution often follows an allelic exclusion pattern, where only 1 of 2 rearranged IgH loci exhibit ongoing recombination. Thus, precursor B-cell leukemias maintain evolution at the IgH locus at levels that were previously underappreciated. This finding sheds light on the mechanisms associated with leukemic clonal evolution and may fundamentally change approaches for monitoring minimal residual disease burden.

Contextual cueing effects across the lifespan.

The authors evaluated age-related variations in contextual cueing, which reflects the extent to which visuospatial regularities can facilitate search for a target. Previous research produced inconsistent results regarding contextual cueing effects in young children and in older adults, and no study has investigated the phenomenon across the life span. Three groups (6, 20, and 70 years old) were compared. Participants located a designated target stimulus embedded in a context of distractor stimuli. During exposure, the location of the target could be predicted from the location of the distracters in each display. During test, these predictable displays were intermixed with new displays that did not predict the target location. Response times to locating predictable relative to unpredictable targets were compared. All groups exhibited facilitation effects greater than 0 (95% CIs [.02, .11], d = .4; [.01, .12], d = .4; and [.01, .10], d = .4, for the children, young adults, and older adults, respectively) indicating that contextual cueing is robust across a wide age range. The relative magnitude of contextual cueing effects was essentially identical across the age range tested, F(2, 103) = 1.71, eta rho2 = .02. The authors argue that a mechanism that uses environmental covariation is available to all age ranges, but the expression of the contextual cueing may depend on the way it is measured.

Multimodal, broadly neutralizing antibodies against SARS-CoV-2 identified by high-throughput native pairing of BCRs from bulk B cells.

TruAB Discovery is an approach that integrates cellular immunology, high-throughput immunosequencing, bioinformatics, and computational biology in order to discover naturally occurring human antibodies for prophylactic or therapeutic use. We adapted our previously described pairSEQ technology to pair B cell receptor heavy and light chains of SARS-CoV-2 spike protein-binding antibodies derived from enriched antigen-specific memory B cells and bulk antibody-secreting cells. We identified approximately 60,000 productive, in-frame, paired antibody sequences, from which 2,093 antibodies were selected for functional evaluation based on abundance, isotype and patterns of somatic hypermutation. The exceptionally diverse antibodies included RBD-binders with broad neutralizing activity against SARS-CoV-2 variants, and S2-binders with broad specificity against betacoronaviruses and the ability to block membrane fusion. A subset of these RBD- and S2-binding antibodies demonstrated robust protection against challenge in hamster and mouse models. This high-throughput approach can accelerate discovery of diverse, multifunctional antibodies against any target of interest.

Bivalent intra-spike binding provides durability against emergent Omicron lineages: Results from a global consortium.

The SARS-CoV-2 Omicron variant of concern (VoC) and its sublineages contain 31-36 mutations in spike and escape neutralization by most therapeutic antibodies. In a pseudovirus neutralization assay, 66 of the nearly 400 candidate therapeutics in the Coronavirus Immunotherapeutic Consortium (CoVIC) panel neutralize Omicron and multiple Omicron sublineages. Among natural immunoglobulin Gs (IgGs), especially those in the receptor-binding domain (RBD)-2 epitope community, nearly all Omicron neutralizers recognize spike bivalently, with both antigen-binding fragments (Fabs) simultaneously engaging adjacent RBDs on the same spike. Most IgGs that do not neutralize Omicron bind either entirely monovalently or have some (22%-50%) monovalent occupancy. Cleavage of bivalent-binding IgGs to Fabs abolishes neutralization and binding affinity, with disproportionate loss of activity against Omicron pseudovirus and spike. These results suggest that VoC-resistant antibodies overcome mutagenic substitution via avidity. Hence, vaccine strategies targeting future SARS-CoV-2 variants should consider epitope display with spacing and organization identical to trimeric spike.

Self responses along cingulate cortex reveal quantitative neural phenotype for high-functioning autism.

Attributing behavioral outcomes correctly to oneself or to other agents is essential for all productive social exchange. We approach this issue in high-functioning males with autism spectrum disorder (ASD) using two separate fMRI paradigms. First, using a visual imagery task, we extract a basis set for responses along the cingulate cortex of control subjects that reveals an agent-specific eigenvector (self eigenmode) associated with imagining oneself executing a specific motor act. Second, we show that the same self eigenmode arises during one's own decision (the self phase) in an interpersonal exchange game (iterated trust game). Third, using this exchange game, we show that ASD males exhibit a severely diminished cingulate self response when playing the game with a human partner. This diminishment covaries parametrically with their behaviorally assessed symptom severity, suggesting its value as an objective endophenotype. These findings may provide a quantitative assessment tool for high-functioning ASD.

Reporter alleles that inform on differences in Cre recombinase expression.

Alleles that express reporters after Cre recombination allow for fate-mapping studies when used in combination with appropriate cre alleles. In this study, we describe two fluorescent reporter alleles that differentially mark populations of cells as a function of their level of expression of Cre recombinase. Mice carrying these alleles were generated and used to demonstrate the usefulness of the reporter alleles for informing on prior Cre recombinase expression in lymphocytes. The alleles expand the range of genetic tools available for understanding how differences in gene expression result in divergent developmental fates during the development and differentiation of lymphocytes and other cells.

MLL5 contributes to hematopoietic stem cell fitness and homeostasis.

MLL5 is a novel trithorax group gene and a candidate tumor suppressor gene located within a 2.5-Mb interval of chromosome band 7q22 that frequently is deleted in human myeloid malignancy. Here we show that inactivation of the Mll5 gene in mice results in a 30% reduction in the average representation of hematopoietic stem cells and in functional impairment of long-term hematopoietic repopulation potential under competitive conditions. Bone marrow cells from Mll5-deficient mice were defective in spleen colony-forming assays, and the mutant mice showed enhanced susceptibility to 5-fluorouracil-induced myelosuppression. Heterozygous and homozygous Mll5 mutant mice did not spontaneously develop hematologic cancers, and loss of Mll5 did not alter the phenotype of a fatal myeloproliferative disorder induced by oncogenic Kras in vivo. Collectively, the data reveal an important role for Mll5 in HSC homeostasis and provide a basis for further studies to explore its role in leukemogenesis.

Thymic OX40 expression discriminates cells undergoing strong responses to selection ligands.

OX40 is a member of the TNF receptor family expressed on activated and regulatory T (Treg) cells. Using an Ox40-cre allele for lineage marking, we found that a subpopulation of naive T cells had also previously expressed OX40 in the thymus. Ox40-cre was induced in a small fraction of thymocytes that were OX40(+), some of which were CD25(high) Treg cell precursors. Thymic OX40 expression distinguished cells experiencing a strong signaling response to positive selection. Naive T cells that had previously expressed OX40 demonstrated a partially activated phenotype that was distinct from that of most naive T cells. The results are consistent with the selection of Treg cells and a minor subpopulation of naive T cells being dependent on strong signaling responses to thymic self ligands.

Examining Environmental Predictors of Community Participation for Adults with Autism Spectrum Disorder Using Geographic Information Systems (GIS).

A geographic information system (GIS) approach systematically assessed whether population density and distribution of community resources contributed to caregiver reported community participation outcomes for 124 adults with autism spectrum disorder (ASD). Regression analyses examined whether GIS measures predicted community participation in areas of social activities and use of services, while also accounting for adult age, conversation ability, and daily living skills (DLS). Results indicated that in addition to person factors of greater DLS and better conversation ability, access to specific community features, such as bus stops, contributed to improved participation. Unexpectedly, population density where one lived made minimal contribution to participation outcomes, except in getting together with friends outside of organized activities.

Assessing the convergence of self-report and informant measures for adults with autism spectrum disorder.

LAY ABSTRACT: Self-report measures are frequently used for research and clinical assessments of adults with autism spectrum disorder. However, there has been little research examining agreement between self-report and informant-report in this population. Valid self-report measures are essential for conducting research with and providing high quality clinical services for adults with autism spectrum disorder. This study collected measures from 40 pairs of adults with autism spectrum disorder and their caregivers on measures of symptom severity, daily living skills, quality of life, and unmet service needs. Caregiver and self-report responses were highly associated with one another on all measures, though there were significant gaps between scores on the measures of daily living skills and quality of life. It is also important to understand how each informant's responses relate to outcomes in the areas of employment and independent living. Using self-report and caregiver-report together better predicted outcomes for the adult with autism spectrum disorder than scores from either individual reporter alone. These findings show that there is unique and valuable information provided by both adults with autism spectrum disorder and their caregivers; a multi-informant approach is important for obtaining the most comprehensive picture of current functioning, identifying unmet service needs, and creating treatment plans. This research also highlights the importance of including and prioritizing self-report perspectives in shaping service planning.

A large-scale database of T-cell receptor beta (TCRß) sequences and binding associations from natural and synthetic exposure to SARS-CoV-2.

We describe the establishment and current content of the ImmuneCODE™ database, which includes hundreds of millions of T-cell Receptor (TCR) sequences from over 1,400 subjects exposed to or infected with the SARS-CoV-2 virus, as well as over 135,000 high-confidence SARS-CoV-2-specific TCRs. This database is made freely available, and the data contained in it can be downloaded and analyzed online or offline to assist with the global efforts to understand the immune response to the SARS-CoV-2 virus and develop new interventions.

Magnitude and Dynamics of the T-Cell Response to SARS-CoV-2 Infection at Both Individual and Population Levels.

T cells are involved in the early identification and clearance of viral infections and also support the development of antibodies by B cells. This central role for T cells makes them a desirable target for assessing the immune response to SARS-CoV-2 infection. Here, we combined two high-throughput immune profiling methods to create a quantitative picture of the T-cell response to SARS-CoV-2. First, at the individual level, we deeply characterized 3 acutely infected and 58 recovered COVID-19 subjects by experimentally mapping their CD8 T-cell response through antigen stimulation to 545 Human Leukocyte Antigen (HLA) class I presented viral peptides (class II data in a forthcoming study). Then, at the population level, we performed T-cell repertoire sequencing on 1,815 samples (from 1,521 COVID-19 subjects) as well as 3,500 controls to identify shared "public" T-cell receptors (TCRs) associated with SARS-CoV-2 infection from both CD8 and CD4 T cells. Collectively, our data reveal that CD8 T-cell responses are often driven by a few immunodominant, HLA-restricted epitopes. As expected, the T-cell response to SARS-CoV-2 peaks about one to two weeks after infection and is detectable for at least several months after recovery. As an application of these data, we trained a classifier to diagnose SARS-CoV-2 infection based solely on TCR sequencing from blood samples, and observed, at 99.8% specificity, high early sensitivity soon after diagnosis (Day 3-7 = 85.1% [95% CI = 79.9-89.7]; Day 8-14 = 94.8% [90.7-98.4]) as well as lasting sensitivity after recovery (Day 29+/convalescent = 95.4% [92.1-98.3]). These results demonstrate an approach to reliably assess the adaptive immune response both soon after viral antigenic exposure (before antibodies are typically detectable) as well as at later time points. This blood-based molecular approach to characterizing the cellular immune response has applications in clinical diagnostics as well as in vaccine development and monitoring.

Safety Evaluation of a 405-nm LED Device for Direct Antimicrobial Treatment of the Murine Brain.

Antimicrobial resistance is a growing problem in human medicine that extends to biomedical research. Compared with chemical-based therapies, light-based therapies present an alternative to traditional pharmaceuticals and are less vulnerable to acquired bacterial resistance. Due to immunologic privilege and relative tissue sensitivity to topical antibiotics, the brain poses a unique set of difficulties with regard to antimicrobial therapy. This study focused on 405-nm 'true violet' light-which has been shown to kill multiple clinically relevant bacterial species in vitro yet leave mammalian cells unscathed-and its effect on the murine brain. We built a 405-nm LED array, validated its power and efficacy against a clinical bacterial isolate in vitro, and then, at the time of craniotomy, treated mice with various doses of 405-nm light (36, 45, and 54 J/cm²). The selected doses caused no behavioral derangements postoperatively or any observable brain pathology as determined postmortem by histologic evaluation and immunofluorescence staining for caspase 3 and glial fibrillary acidic protein, markers of apoptosis and necrosis. True-violet light devices may present an inexpensive refinement to current practices for maintaining open craniotomy sites or reducing bacterial loads in contaminated surgical sites.

Understanding Service Usage and Needs for Adults with ASD: The Importance of Living Situation.

With the increasing prevalence of adults with autism spectrum disorder (ASD), research examining the service experiences of this population is greatly needed. The current study investigated service use, unmet needs, and obstacles to service access for a large sample of adults with ASD. After accounting for various demographic factors known to impact service usage and needs, living situation was a significant predictor of service use, needs, and obstacles to services. Adults with ASD living with family reported less service use, higher unmet need, and more obstacles to accessing services. With more than half of this adult sample living with family, results have clear public policy implications to support the increasing population of adults with ASD living with aging caregivers.