Preclinical studies show that Co-STARs combine the advantages of chimeric antigen and T cell receptors for the treatment of tumors with low antigen densities.

Two types of engineered T cells have been successfully used to treat patients with cancer, one with an antigen recognition domain derived from antibodies [chimeric antigen receptors (CARs)] and the other derived from T cell receptors (TCRs). CARs use high-affinity antigen-binding domains and costimulatory domains to induce T cell activation but can only react against target cells with relatively high amounts of antigen. TCRs have a much lower affinity for their antigens but can react against target cells displaying only a few antigen molecules. Here, we describe a new type of receptor, called a Co-STAR (for costimulatory synthetic TCR and antigen receptor), that combines aspects of both CARs and TCRs. In Co-STARs, the antigen-recognizing components of TCRs are replaced by high-affinity antibody fragments, and costimulation is provided by two modules that drive NF-κB signaling (MyD88 and CD40). Using a TCR-mimic antibody fragment that targets a recurrent p53 neoantigen presented in a common human leukocyte antigen (HLA) allele, we demonstrate that T cells equipped with Co-STARs can kill cancer cells bearing low densities of antigen better than T cells engineered with conventional CARs and patient-derived TCRs in vitro. In mouse models, we show that Co-STARs mediate more robust T cell expansion and more durable tumor regressions than TCRs similarly modified with MyD88 and CD40 costimulation. Our data suggest that Co-STARs may have utility for other peptide-HLA antigens in cancer and other targets where antigen density may limit the efficacy of engineered T cells.

What is the Utilization and Impact of Advanced Imaging for Tibial Tubercle Fractures? An Analysis of 598 Patients From the Tibial Tubercle Study (TITUS) Group.

BACKGROUND: Given the rare nature of tibial tubercle fractures, previous studies are mostly limited to small, single-center series. This results in practice variation. Previous research has shown poor surgeon agreement on utilization of advanced imaging, but improved evidence-based indications may help balance clinical utility with resource utilization. The purpose of this study is to quantify diagnostic practices for tibial tubercle fractures in a large, multicenter cohort, with attention to the usage and impact of advanced imaging. METHODS: This is a retrospective series of pediatric tibial tubercle fractures from 7 centers between 2007 and 2022. Exclusion criteria were age above 18 years, missing demographic and pretreatment data, closed proximal tibial physis and tubercle apophysis, or a proximal tibia fracture not involving the tubercle. Demographic and injury data were collected. Fracture classifications were derived from radiographic evaluation. The utilization of advanced imaging was recorded as well as the presence of findings not identified on radiographs. Standard descriptive statistics were reported, and χ2 tests were performed (means reported±SD). RESULTS: A total of 598 patients satisfied the inclusion criteria, of which 88.6% (530/598) were male with a mean age of 13.8±1.9 years. Internal oblique x-rays were obtained in 267 patients (44.6%), computed tomography (CT) in 158 (26.4%), and magnetic resonance imaging (MRI) in 64 (10.7%). There were significant differences in the frequency at which CT (7.2% to 79.4%, P<0.001) and MRI were obtained (1.5% to 54.8%, P<0.001). CT was obtained most frequently for Ogden type IV fractures (50/99, 50.5%), and resulted in novel findings that were not visualized on radiographs in a total of 37/158 patients (23.4%). The most common finding on CT was intra-articular fracture extension (25/37). MRI was obtained most frequently for Ogden type V fractures (13/35, 37.1%), and resulted in novel findings in a total of 31/64 patients (48.4%). The most common finding was patellar tendon injury (11/64), but only 3 of these patients required tendon repair. CONCLUSIONS: Substantial variation exists in the diagnostic evaluation of tibial tubercle fractures. CT was most helpful in clarifying intra-articular involvement, while MRI can identify patellar tendon injury, periosteal sleeve avulsion, or a nondisplaced fracture. This study quantifies variation in diagnostic practices for tibial tubercle fractures, highlighting the need for evidence-based indications for advanced imaging. LEVEL OF EVIDENCE: Level III.

Low neighborhood socioeconomic status is associated with poor outcomes in young adults with colorectal cancer.

BACKGROUND: The incidence of early-onset colorectal cancer has increased markedly over the past decade. Although established for older adults, there are limited data on socioeconomic and racial disparities in screening, treatment, and outcomes in this distinct group. METHODS: Adults with primary colorectal cancer diagnosed at age <50 were identified from the Surveillance, Epidemiology, and End Results database. The exposure of interest was neighborhood socioeconomic status based on the Yost Index, a census-tract level composite score of neighborhood economic health. Univariate analysis was performed with χ2 analyses. Logistic regression models were created to evaluate the association of neighborhood socioeconomic status (Yost Index quintile) with metastasis at presentation and surgical intervention. Kaplan-Meier and Cox proportional hazards models were created. RESULTS: In total, 45,660 early-onset colorectal cancer patients were identified; 16.8% (7,679) were in the lowest quintile of neighborhood socioeconomic status. Patients with the lowest neighborhood socioeconomic status were 1.13 times (95% confidence interval 1.06-1.21) more likely to present with metastases and had lower survival (hazard ratio 1.45, 95% confidence interval 1.37-1.53) compared to those with the highest neighborhood socioeconomic status. Non-Hispanic Black patients were more likely to present with metastatic disease (odds ratio 1.11, 95% confidence interval 1.05-1.19), less likely to undergo surgery for localized or regional disease (odds ratio 0.48, 95% confidence interval 0.43-0.53), and had lower survival (hazard ratio 1.21, 95% confidence interval 1.15-1.27) than non-Hispanic White patients. CONCLUSION: Socioeconomic and racial disparities in early-onset colorectal cancer span diagnosis, treatment, and survival. As the disease burden of early-age onset colorectal cancer increases, interventions to boost early diagnosis and access to surgery are necessary to improve survival among minorities and patients with low neighborhood socioeconomic status.

Disparities in air pollution attributable mortality in the US population by race/ethnicity and sociodemographic factors.

There are large differences in premature mortality in the USA by race/ethnicity, education, rurality and social vulnerability index groups. Using existing concentration-response functions, published particulate matter (PM2.5) air pollution estimates, population estimates at the census tract level and county-level mortality data from the US National Vital Statistics System, we estimated the degree to which these mortality discrepancies can be attributed to differences in exposure and susceptibility to PM2.5. We show that differences in PM2.5-attributable mortality were consistently more pronounced by race/ethnicity than by education, rurality or social vulnerability index, with the Black American population having the highest proportion of deaths attributable to PM2.5 in all years from 1990 to 2016. Our model estimates that over half of the difference in age-adjusted all-cause mortality between the Black American and non-Hispanic white population was attributable to PM2.5 in the years 2000 to 2011. This difference decreased only marginally between 2000 and 2015, from 53.4% (95% confidence interval 51.2-55.9%) to 49.9% (95% confidence interval 47.8-52.2%), respectively. Our findings underscore the need for targeted air quality interventions to address environmental health disparities.

Inflammatory and Autoimmune Aspects of Multisystem Inflammatory Syndrome in Children (MIS-C): A Prospective Cohort Study.

Multisystem Inflammatory Syndrome in Children (MIS-C) is a potentially life-threatening complication of COVID-19. The pathophysiological mechanisms leading to severe disease are poorly understood. This study leveraged clinical samples from a well-characterized cohort of children hospitalized with COVID-19 or MIS-C to compare immune-mediated biomarkers. Our objective was to identify selected immune molecules that could explain, in part, why certain SARS-CoV-2-infected children developed MIS-C. We hypothesized that type-2 helper T cell-mediated inflammation can elicit autoantibodies, which may account for some of the differences observed between the moderate-severe COVID-19 (COVID+) and MIS-C cohort. We enumerated blood leukocytes and measured levels of selected serum cytokines, chemokines, antibodies to COVID-19 antigens, and autoantibodies in children presenting to an academic medical center in Connecticut, United States. The neutrophil/lymphocyte and eosinophil/lymphocyte ratios were significantly higher in those in the MIS-C versus COVID+ cohort. IgM and IgA, but not IgG antibodies to SARS-CoV-2 receptor binding domain were significantly higher in the MIS-C cohort than the COVID+ cohort. The serum levels of certain type-2 cytokines (interleukin (IL)-4, IL-5, IL-6, IL-8, IL-10, IL-13, and IL-33) were significantly higher in children with MIS-C compared to the COVID+ and SARS-CoV-2-negative cohorts. IgG autoantibodies to brain antigens and pentraxin were higher in children with MIS-C compared to SARS-CoV-19-negative controls, and children with MIS-C had higher levels of IgG anti-contactin-associated protein-like 2 (caspr2) compared to the COVID+ and SARS-CoV-19-negative controls. We speculate that autoimmune responses in certain COVID-19 patients may induce pathophysiological changes that lead to MIS-C. The triggers of autoimmunity and factors accounting for type-2 inflammation require further investigation.

Single-cell topographical profiling of the immune synapse reveals a biomechanical signature of cytotoxicity.

Immune cells have intensely physical lifestyles characterized by structural plasticity and force exertion. To investigate whether specific immune functions require stereotyped mechanical outputs, we used super-resolution traction force microscopy to compare the immune synapses formed by cytotoxic T cells with contacts formed by other T cell subsets and by macrophages. T cell synapses were globally compressive, which was fundamentally different from the pulling and pinching associated with macrophage phagocytosis. Spectral decomposition of force exertion patterns from each cell type linked cytotoxicity to compressive strength, local protrusiveness, and the induction of complex, asymmetric topography. These features were validated as cytotoxic drivers by genetic disruption of cytoskeletal regulators, live imaging of synaptic secretion, and in silico analysis of interfacial distortion. Synapse architecture and force exertion were sensitive to target stiffness and size, suggesting that the mechanical potentiation of killing is biophysically adaptive. We conclude that cellular cytotoxicity and, by implication, other effector responses are supported by specialized patterns of efferent force.

Pre-Transplant Frequencies of FoxP3+CD25+ in CD3+CD8+ T Cells as Potential Predictors for CMV in CMV-Intermediate Risk Kidney Transplant Recipients.

Cytomegalovirus (CMV) infection detrimentally influences graft survival in kidney transplant recipients, with the risk primarily determined by recipient and donor serostatus. However, recipient CD8+ T cells play a crucial role in CMV control. The optimal preventive strategy (prophylaxis vs. pre-emptive treatment), particularly for seropositive (intermediate risk) recipients, remains uncertain. We investigated CD8+ T cell subpopulation dynamics and CMV occurrence (DNAemia ≥ 100 IU/mL) in 65 kidney transplant recipients, collecting peripheral blood mononuclear cells before (T1) and 1 year after transplantation (T2). Comparing the two timepoints, we found an increase in granulocyte, monocyte and CD3+CD8+ T cells numbers, while FoxP3+CD25+, LAG-3+ and PD-1+ frequencies were reduced at T2. CMV DNAemia occurred in 33 recipients (55.8%) during the first year. Intermediate risk patients were disproportionally affected by posttransplant CMV (N = 29/45, 64.4%). Intermediate risk recipients developing CMV after transplantation exhibited lower leukocyte, monocyte, and granulocyte counts and higher FoxP3+CD25+ frequencies in CD3+CD8+ T cells pre-transplantation compared to patients staying CMV negative. Pre-transplant FoxP3+CD25+ in CD3+CD8+ T cells had the best discriminatory potential for CMV infection prediction within the first year after transplantation (AUC: 0.746). The FoxP3+CD25+ CD3+CD8+ T cell subset may aid in selecting intermediate risk kidney transplant recipients for CMV prophylaxis.

Synthesis of ß-Pyridyl α-Amino Acids: Conformationally Sensitive Charge Transfer-Based Fluorophores.

Unnatural α-amino acids with charge transfer-based poly aromatic side chains have been designed as conformationally sensitive fluorophores. These were prepared using a hetero-Diels-Alder reaction and a Knoevenagel-Stobbe process to generate a biaryl pyridyl unit, followed by iron-catalyzed bromination and a Suzuki-Miyaura cross-coupling reaction to complete the triaryl system. A photophysical study led to the discovery of a p-methoxy analogue which exhibited viscosity-sensitive fluorescence in which emission could be controlled between twisted and planar conformations.

Chemokines and Cytokines in Immunotherapy of Melanoma and Other Tumors: From Biomarkers to Therapeutic Targets.

Chemokines and cytokines represent an emerging field of immunotherapy research. They are responsible for the crosstalk and chemoattraction of immune cells and tumor cells. For instance, CXCL9/10/11 chemoattract effector CD8+ T cells to the tumor microenvironment, making an argument for their promising role as biomarkers for a favorable outcome. The cytokine Interleukin-15 (IL-15) can promote the chemokine expression of CXCR3 ligands but also XCL1, contributing to an important DC-T cell interaction. Recruited cytotoxic T cells can be clonally expanded by IL-2. Delivering or inducing these chemokines and cytokines can result in tumor shrinkage and might synergize with immune checkpoint inhibition. In addition, blocking specific chemokine and cytokine receptors such as CCR2, CCR4 or Il-6R can reduce the recruitment of tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs) or regulatory T cells (Tregs). Efforts to target these chemokines and cytokines have the potential to personalize cancer immunotherapy further and address patients that are not yet responsive because of immune cell exclusion. Targeting cytokines such as IL-6 and IL-15 is currently being evaluated in clinical trials in combination with immune checkpoint-blocking antibodies for the treatment of metastatic melanoma. The improved overall survival of melanoma patients might outweigh potential risks such as autoimmunity. However, off-target toxicity needs to be elucidated.

Submitting Novel Full-Length HLA, MIC, and KIR Alleles with TypeLoader2.

The Immuno Polymorphism Database (IPD) plays a pivotal role for immunogenetics. Due to technical limitations, genotyping often focuses on specific key regions like the antigen recognition domain (ARD) for HLA genotyping, and the databases are populated accordingly. More recently, though, modern next generation sequencing (NGS) assays allow using larger gene segments or even complete genes for genotyping. It is therefore essential that the databases are updated with complete genetic reference sequences to fully serve current and future applications. However, the process of manually annotating and submitting full-length allele sequences to IPD is time-consuming and error-prone, which may discourage HLA-genotyping laboratories or researchers from submitting full-length sequences of novel alleles.Here, we detail the process of preparing and submitting novel HLA, MIC, and KIR alleles to ENA and IPD using TypeLoader2, a convenient software tool developed to streamline this process by automating the sequence annotation, the creation of all necessary files, as well as parts of the submission process itself. The software is freely available from GitHub ( https://github.com/DKMS-LSL/typeloader ).

Full-Length Characterization of Novel HLA-DRB1 Alleles for Reference Database Submission.

The prerequisite for successful HLA genotyping is the integrity of the large allele reference database IPD-IMGT/HLA. Consequently, it is in the laboratories' best interest that the data quality of submitted novel sequences is high. However, due to its long and variable length, the gene HLA-DRB1 presents the biggest challenge and as of today only 16% of the HLA-DRB1 alleles in the database are characterized in full length. To improve this situation, we developed a protocol for long-range PCR amplification of targeted HLA-DRB1 alleles. By subsequently combining both long-read and short-read sequencing technologies, our protocol ensures phased and error-corrected sequences of reference grade quality. This dual redundant reference sequencing (DR2S) approach is of particular importance for correctly resolving the challenging repeat regions of DRB1 intron 1. Until today, we used this protocol to characterize and submit 384 full-length HLA-DRB1 sequences to IPD-IMGT/HLA.

Differences in drug removal between standard high-flux and medium cut-off dialyzers in a case of severe vancomycin toxicity.

Vancomycin is a widely used glycopeptide antibiotic with the need for therapeutic drug monitoring to avoid renal toxicity. We report a case of severe vancomycin-associated anuric acute kidney injury managed with successful drug-removal by hemodialysis (HD) using different types of dialyzers. Medium cut-off (MCO) and high-flux dialyzers were effective in drug removal. Higher vancomycin elimination rate and lower plasma half-life were achieved with MCO dialyzer despite low-flow vascular access and intolerance to ultrafiltration. MCO dialyzers may be reasonable for drug removal in patients with intolerance of ultrafiltration, low-flow vascular access or impracticality of hemodiafiltration. Future studies should explore the use of MCO dialyzers in comparison with high-flux HD and hemodiafiltration in both the acute and chronic setting.

Successful Treatment of Severe Pemphigus Vulgaris with Reduced Side Effects Using a Novel IVIg Preparation.

Pemphigus vulgaris (PV) is a rare autoimmune bullous dermatosis (AIBD) characterized by painful blistering of the skin and mucosa caused by autoantibodies that lead to loss of adhesion in the epidermis. Standard therapy for PV is corticosteroids, either alone or in combination with steroid-sparing immunosuppressants or infusions with rituximab. According to the published European guideline, high-dose intravenous immunoglobulin (IVIg) therapy with a dosage of 2 g per kg body weight distributed over 2-5 days every 4 weeks is a promising treatment option, especially for severe or refractory disease. This report describes a 73-year-old female patient with severe and recurrent disease who achieved stabilization with IVIg treatment. However, the patient experienced side effects such as headaches, nausea, and vomiting, which affected daily life. Hence, she was transitioned to a new IVIg preparation with a new manufacturing process, resulting in fewer side effects and an improved quality of life. Further follow-up is necessary to fully evaluate the effectiveness and tolerability of this new IVIg product.

Unraveling the neuroimmune interface in chronic pain-the association between cytokines in the cerebrospinal fluid and pain in patients with lumbar disk herniation or degenerative disk disease.

ABSTRACT: Recent evidence highlights the importance of the neuroimmune interface, including periphery-to-central nervous system (CNS) neuroimmune crosstalk, in chronic pain. Although neuroinflammatory processes have been implicated in central sensitization for a long time, their potential neuroprotective and analgesic effects remain relatively elusive. We have explored the relationships between cytokine expression and symptom severity, and candidates for periphery-to-CNS crosstalk. Patients with degenerative disk disease (DDD) (nociceptive pain) or patients with lumbar disk herniation (LDH) with radiculopathy (predominantly neuropathic pain) completed questionnaires regarding pain and functional disability, underwent quantitative sensory testing, and provided blood and cerebrospinal fluid (CSF) samples. Proximity extension assay (PEA) was used to measure the levels of 92 inflammatory proteins in the CSF and serum from a total of 160 patients and controls, and CSF/serum albumin quotients was calculated for patients with DDD and patients with LDH. We found signs of neuroimmune activation, in the absence of systemic inflammation. Regarding periphery-to-CNS neuroimmune crosstalk, there were significant associations between several cytokines and albumin quotient, despite the latter being primarily at subclinical levels. The cytokines CCL11, CD5, IL8, and MMP-10 were elevated in the CSF, had positive correlations between CSF and serum levels, and associated in a nonlinear manner with back, but not leg, pain intensity in the LDH, but not the DDD, group. In conclusion, we found evidence for neuroimmune activation in the CNS of both patient groups in the absence of systemic inflammation and signs of a communication between CSF and serum. Complex and disease-specific associations were found between cytokines in CSF and back pain intensity.

Plasma membrane abundance dictates phagocytic capacity and functional cross-talk in myeloid cells.

Professional phagocytes like neutrophils and macrophages tightly control what they consume, how much they consume, and when they move after cargo uptake. We show that plasma membrane abundance is a key arbiter of these cellular behaviors. Neutrophils and macrophages lacking the G protein subunit Gß4 exhibited profound plasma membrane expansion, accompanied by marked reduction in plasma membrane tension. These biophysical changes promoted the phagocytosis of bacteria, fungus, apoptotic corpses, and cancer cells. We also found that Gß4-deficient neutrophils are defective in the normal inhibition of migration following cargo uptake. Sphingolipid synthesis played a central role in these phenotypes by driving plasma membrane accumulation in cells lacking Gß4. In Gß4 knockout mice, neutrophils not only exhibited enhanced phagocytosis of inhaled fungal conidia in the lung but also increased trafficking of engulfed pathogens to other organs. Together, these results reveal an unexpected, biophysical control mechanism central to myeloid functional decision-making.

Size matters: Total testicular volume predicts sperm count in Tanner V varicocele patients.

INTRODUCTION: It is known the prevalence of varicoceles in adolescent men is 14-29% but there is debate surrounding implications on fertility. As obtaining a semen analysis (SA) may be challenging, there is need for objective tests as measures of fecundity. Our aim was to investigate the relationship between testicular volume differential (TVD), varicocele grade, and total testicular volume (TTV) on seminal parameters including total motile sperm count (TMSC). MATERIALS AND METHODS: We conducted a retrospective single-center chart review over 14 years of 486 Tanner V adolescent males. Three hundred and four met inclusion of palpable, non-operated left-sided varicocele who underwent at least one SA and ultrasound. Abnormal TMSC was defined by World Health Organization 2010 criteria for minimal reference ranges. Multivariate logistic regression, receiver operating characteristic analysis with Youden J-statistic and descriptive statistics were performed. RESULTS: Three hundred and four Tanner V adolescents with median age of 18.0 years (18.0-19.0), median TTV of 34.5 cc (28.9, 40.2) and median TMSC of 62.5 million/ejaculate (25.4, 123.4) were evaluated. TTV cutoff of 29.5 cc was found to predict TMSC of <9 million/ejaculate with negative predictive value of 96.2% and odds ratio of 6.08 ([2.13-17.42], p < 0.001). TVD greater than 20% did not reach statistical significance with an odds ratio of 1.66 ([0.41-6.62], p = 0.50). DISCUSSION: In clinical practice, each patient will need to have an individualized plan. Based on our data, for older adolescents (17 or 18 years) with varicocele and an abnormal TTV, clinicians may have a lower threshold for advising SA, and if unable to obtain, surgical intervention and/or closer surveillance should be stressed. Patients should be informed of their six-fold increase in abnormal SA. Patients with normal TTV should be advised they are at lower risk of having abnormal SA. Younger patients with varicocele and an initial TVD>20%, should be followed closely but intervention delayed until 17 or 18 to better assess TTV. The importance of trending patient data should be emphasized as a single measurement has low predictive value for developing adolescents. Limitations of our study include a retrospective design and the lack of uniform correlation between adolescent SA and paternity. CONCLUSIONS: Total testicular volume less than 29.5 cc increased odds of abnormal semen analysis by over six times and had a negative predictive value of 96.2%. Ultrasound results may be useful for risk stratification and counselling on appropriateness of surgical intervention.

Transcriptome sequencing of seven deep marine invertebrates.

We present 4k video and whole transcriptome data for seven deep-sea invertebrate animals collected in the Eastern Pacific Ocean during a research expedition onboard the Schmidt Ocean Institute's R/V Falkor in August of 2021. The animals include one jellyfish (Atolla sp.), three siphonophores (Apolemia sp., Praya sp., and Halistemma sp.), one larvacean (Bathochordaeus mcnutti), one tunicate (Pyrosomatidae sp.), and one ctenophore (Lampocteis sp.). Four of the animals were sequenced with long-read RNA sequencing technology, such that the reads themselves define a reference assembly for those animals. The larvacean tissues were successfully preserved in situ and has paired long-read reference data and short read quantitative transcriptomic data for within-specimen analyses of gene expression. Additionally, for three animals we provide quantitative image data, and a 3D model for one siphonophore. The paired image and transcriptomic data can be used for species identification, species description, and reference genetic data for these deep-sea animals.