The ongoing COVID-19 pandemic calls for more effective diagnostic tools. T cell response assessment serves as an independent indicator of prior COVID-19 exposure while also contributing to a more comprehensive characterization of SARS-CoV-2 immunity. In this study, we systematically assessed the immunogenicity of 118 epitopes with immune cells collected from multiple cohorts of vaccinated, convalescent, healthy unexposed, and SARS-CoV-2 exposed donors. We identified 75 immunogenic epitopes, 24 of which were immunodominant. We further confirmed HLA restriction for 49 epitopes, and described association with more than one HLA allele for 14 of these. Exclusion of two cross-reactive epitopes that generated a response in pre-pandemic samples, left us with a 73-epitope set that offered excellent diagnostic specificity without losing sensitivity compared to full-length antigens, which evoked a robust cross-reactive response. We subsequently incorporated this set of epitopes into an in vitro diagnostic ‘Corona-T-test’ which achieved a diagnostic accuracy of 95% in a clinical trial. In a cohort of asymptomatic seronegative individuals with a history of prolonged SARS-CoV-2 exposure, we observed a complete absence of T cell response to our epitope panel. In combination with strong reactivity to full-length antigens, this suggests that a cross-reactive response might protect these individuals.
Aleksei Titov, Regina Shaykhutdinova, Olga V. Shcherbakova, Yana V. Serdyuk, Savely A. Sheetikov, Ksenia V. Zornikova, Alexandra V. Maleeva, Alexandra Khmelevskaya, Dmitry V. Dianov, Naina T. Shakirova, Dmitry B. Malko, Maxim Shkurnikov, Stepan Nersisyan, Alexander Tonevitsky, Ekaterina Khamaganova, Anton V. Ershov, Elena Y. Osipova, Ruslan V. Nikolaev, Dmitry E. Pershin, Viktoria A. Vedmedskia, Mikhail Maschan, Victoria R. Ginanova, Grigory A. Efimov
BACKGROUND. Potent synthetic opioids, such as fentanyl, are increasingly abused, resulting in unprecedented numbers of fatalities from respiratory depression. Treatment with the high-affinity mu-opioid receptor partial agonist buprenorphine may prevent fatalities by reducing binding of potent opioids to the opioid receptor, limiting respiratory depression. METHODS. To characterize buprenorphine-fentanyl interaction at the level of the mu-opioid receptor in two populations (opioid-naïve individuals and chronic users of high-dose opioids), the effects of escalating intravenous fentanyl doses with range 0.075-0.35 mg/70kg (opioid-naïve) and 0.25-0.70 mg/70 kg (chronic opioid users) on iso-hypercapnic ventilation at 2-3 background doses of buprenorphine (target plasma concentrations range: 0.2-5 ng/mL) were quantified using receptor association/dissociation models combined with biophase distribution models. RESULTS. Buprenorphine produced mild respiratory depression, while high doses of fentanyl caused pronounced respiratory depression and apnea in both populations. When combined with fentanyl, buprenorphine produced a receptor binding-dependent reduction of fentanyl-induced respiratory depression in both populations. In chronic opioid users, at buprenorphine plasma concentrations ≥2 ng/mL, a protective effect against high-dose fentanyl was observed. CONCLUSION. Overall, the results indicate that when buprenorphine mu-opioid receptor occupancy is sufficiently high, fentanyl is unable to activate the mu-opioid receptor and consequently will not cause further respiratory depression in addition to the mild respiratory effects of buprenorphine. TRIAL REGISTRATION. Trialregister.nl, number NL7028 (https://www.trialregister.nl/trial/7028) FUNDING. Indivior Inc., North Chesterfield, VA.
Erik Olofsen, Marijke Hyke Algera, Laurence Moss, Robert L. Dobbins, Geert J. Groeneveld, Monique van Velzen, Marieke Niesters, Albert Dahan, Celine M. Laffont
BACKGROUND. Accumulation of advanced glycation endproducts (AGEs) may contribute to the pathophysiology of type 2 diabetes and its vascular complications. AGEs are widely present in food, but whether restricting AGE intake improves risk factors for type 2 diabetes and vascular dysfunction is controversial. RESEARCH DESIGN AND METHODS. Abdominally obese but otherwise healthy individuals were randomly assigned to a specifically designed 4-week diet low or high in AGEs in a double blind parallel-design. Insulin sensitivity, secretion, and clearance were assessed by a combined hyperinsulinemic-euglycemic and hyperglycemic clamp. Micro- and macrovascular function, inflammation, and lipid profile were assessed by state-of-art in vivo measurements and biomarkers. Specific urinary and plasma AGEs Nε-(carboxymethyl)lysine (CML), Nε-(1-carboxyethyl)lysine (CEL), and Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1) were assessed by mass spectrometry. RESULTS. In 73 individuals (22 males, mean ± SD age and BMI 52 y ± 14, 30.6 kg/m2 ± 4.0), intake of CML, CEL, and MG-H1 differed 2.7, 5.3, and 3.7-fold between the low and high AGE diets, which led to corresponding changes of these AGEs in urine and plasma. Despite this, there was no difference in insulin sensitivity, secretion, or clearance, micro- and macrovascular function, overall inflammation, or lipid profile between the low and high dietary AGE groups (all p for treatment effects > 0.05). CONCLUSIONS. This comprehensive RCT demonstrates very limited biological consequences of a 4-week diet low or high in AGEs in abdominally obese individuals. TRIAL REGISTRATION. clinicaltrials.gov: NCT03866343, trialregister.nl: NTR7594. FUNDING. Diabetesfonds and ZonMw.
Armand M A Linkens, Alfons J. Houben, Petra M Niessen, Nicole Wijckmans, Erica de Goei, Mathias D.G. Van den Eynde, Jean L. J. M. Scheijen, Marjo Waarenburg, Andrea Mari, Tos T.J.M. Berendschot, Lukas Streese, Henner Hanssen, Martien C.J.M. van Dongen, Christel van Gool, Coen D.A. Stehouwer, Simone JPM Eussen, Casper Schalkwijk.
BACKGROUND. Recessive dystrophic epidermolysis bullosa (RDEB) is a rare, devastating, and life-threatening inherited skin fragility disorder due to a lack of functional type VII collagen, for which no effective therapy exists. ABCB5-positive dermal mesenchymal stem cells (ABCB5+ MSCs) possess immunomodulatory capacities, a favorable skin homing potential and the ability to secrete type VII collagen. In a COL7A1–/– mouse model of RDEB, treatment with ABCB5+ MSCs markedly extended the animals’ lifespans. METHODS. In this international, multicentric, single-arm, phase I/IIa clinical trial, 16 patients (aged 4–36 years) enrolled into four age cohorts received three intravenous infusions of 2×106 ABCB5+ MSCs/kg on days 0, 17 and 35. Patients were followed up for 12 weeks regarding efficacy and 12 months regarding safety. RESULTS. At 12 weeks, statistically significant median (IQR) reductions in the Epidermolysis Bullosa Disease Activity and Scarring Index activity (EBDASI activity) score of 13.0% (2.9%-30%; P = 0.049) and the Instrument for Scoring Clinical Outcome of Research for Epidermolysis Bullosa clinician (iscorEB c) score of 18.2% (4.1%-41.7%; P = 0.037) were observed. Reductions in itch and pain numerical rating scale scores were greatest on day 35, amounting to 37.5% (0.0%-42.9%; P = 0.033) and 25.0% (-8.4%-46.4%; P = 0.168), respectively. Three adverse events were considered related to the cell product, one mild lymphadenopathy and two hypersensitivity reactions. The latter two were serious but resolved without sequelae shortly after withdrawal of treatment. CONCLUSION. This trial demonstrates good tolerability, manageable safety and potential efficacy of intravenous ABCB5+ MSCs as a readily available disease-modifying therapy for RDEB and provides a rationale for further clinical evaluation. TRIAL REGISTRATION. clinicaltrials.gov NCT03529877; EudraCT 2018-001009-98 FUNDING. The trial was sponsored by RHEACELL GmbH & Co. KG, Heidelberg, Germany. Contributions by NY Frank and MH Frank to this work were supported by the National Institutes of Health (NIH)/National Eye Institute (NEI) grants RO1EY025794 and R24EY028767.
Dimitra Kiritsi, Kathrin Dieter, Elke Niebergall-Roth, Silvia Fluhr, Cristina Daniele, Jasmina Esterlechner, Samar Sadeghi, Seda Ballikaya, Leoni Erdinger, Franziska Schauer, Stella Gewert, Martin Laimer, Johann W. Bauer, Alain Hovnanian, Giovanna Zambruno, May El Hachem, Emmanuelle Bourrat, Maria Papanikolaou, Gabriela Petrof, Sophie Kitzmüller, Christen L. Ebens, Markus H. Frank, Natasha Y. Frank, Christoph Ganss, Anna E. Martinez, John A. McGrath, Jakub Tolar, Mark A. Kluth
BACKGROUND. A previous Phase I study showed that the infusion of autologous Treg cells expanded ex-vivo into recent onset Type 1 Diabetes (T1D) patients had an excellent safety profile, however, the majority of the infused Tregs could no longer be detected in the peripheral blood three months post-infusion (NCT01210664-Treg-T1D Trial). Interleukin-2 (IL-2) has been shown to enhance human Treg cell survival and expansion at low doses in vivo. Therefore, we conducted a Phase 1 study (NCT02772679-TILT trial) combining polyclonal Treg and low-dose IL-2 and assessed the impact over time on Tregs populations and other immune cells. METHODS. T1D Patients were treated with a single infusion of autologous polyclonal Tregs, expanded ex vivo, followed by one or two 5-day courses of recombinant human low dose IL-2 (ld-IL-2) at 0.3 x 106 to 1 x 106 units/dose. Fluorescence-based flow cytometry, Cytometry by Time Of Flight (CyTOF) and 10X Genomics single cell RNAseq were used to follow the distinct immune cell populations phenotypes over time following immunotherapy. RESULTS. Multiparametric analysis revealed that the combination therapy led to an increase in the number of infused and endogenous Tregs but also resulted in a significant increase from baseline in a subset of activated NK, Mucosal-Associated Invariant T (MAIT) cells and clonal CD8+ T populations. CONCLUSIONS. These data support the hypothesis that Id-IL-2 expands exogenously administered Tregs but also can expand cytotoxic cells. These results have important implications for the use of a combination of ld-IL-2 and Tregs for the treatment of autoimmune diseases with pre-existing active immunity. TRIAL REGISTRATION. ClinicalTrials.gov NCT01210664 (Treg-T1D Trial), NCT02772679 (TILTtrial). FUNDING. Sean N. Parker Autoimmunity Research Laboratory Fund, National Center for Research Resources.
Shen Dong, Kamir J. Hiam-Galvez, Cody T. Mowery, Kevan C. Herold, Stephen E. Gitelman, Jonathan H. Esensten, Weihong Liu, Angela P. Lares, Ashley S. Leinbach, Michael Lee, Vinh Nguyen, Stanley J. Tamaki, Whitney Tamaki, Courtney M. Tamaki, Morvarid Mehdizadeh, Amy L. Putnam, Matthew H. Spitzer, C. Jimmie Ye, Qizhi Tang, Jeffrey A. Bluestone
BACKGROUND. Naturally acquired immunity to malaria is incompletely understood. We used controlled human malaria infection (CHMI) to study the impact of past exposure to malaria in Kenyan adults in relation to infection with a non-Kenyan parasite strain. METHODS. We administered 3.2x103 aseptic, purified, cryopreserved Plasmodium falciparum (Pf) sporozoites (SPZ) [Sanaria® PfSPZ Challenge, NF54 West African strain] by direct venous inoculation and undertook clinical monitoring and serial quantitative PCR (qPCR) of the 18S ribosomal RNA gene. The study endpoint was met when either: parasitaemia reached ≥500 parasites/μl blood; clinically significant symptoms were seen; or at 21 days after inoculation. All volunteers received antimalarial drug treatment on meeting the endpoint. RESULTS. One hundred and sixty-one (161) volunteers underwent CHMI between Aug 4, 2016, and Feb 14, 2018. CHMI was well tolerated with no severe or serious adverse events. Nineteen volunteers (11.8%) were excluded from the analysis based on detection of antimalarial drugs above the minimal inhibitory concentration or parasites genotyped as non-NF54. Of the 142 volunteers who were eligible for analysis: 26 (18.3%) had febrile symptoms and were treated; 30 (21.1%) reached ≥500 parasites/μl and were treated; 53 (37.3%) had parasitaemia without meeting thresholds for treatment and; 33 (23.2%) remained qPCR negative. CONCLUSION. We find that past exposure to malaria, as evidenced by location of residence, in some Kenyan adults can completely suppress in vivo growth of a parasite strain originating from outside Kenya. TRAIL REGISTRATION. The study was registered on ClinicalTrials.gov (NCT02739763). FUNDING. Wellcome Trust
Melissa C. Kapulu, Patricia Njuguna, Mainga Hamaluba, Domtila Kimani, Joyce M. Ngoi, Janet Musembi, Omar Ngoto, Edward Otieno, Peter F. Billingsley
BACKGROUND. The aberrant activation of the PI3K/mTOR signaling circuitry is one of the most frequently dysregulated signaling events in head and neck squamous cell carcinoma (HNSCC). Here, we conducted a single-arm, open label phase IIa clinical trial (NCT02581137) in individuals with oral premalignant lesions (OPL) to explore the potential of metformin to target PI3K/mTOR signaling for HNSCC prevention. METHODS. Subjects with OPL, otherwise healthy and without diabetes, underwent pre- and post-treatment clinical exam and biopsy. Participants received metformin for 12 weeks (week 1, 500 mg; week 2, 1,000 mg; week 3-12, 2,000 mg daily). Pre- and post-treatment biopsies, saliva, and blood were obtained for biomarker analysis, including immunohistochemical (IHC) assessment of mTOR signaling and exome sequencing. RESULTS. Twenty-three participants were evaluable for response. The clinical response rate (defined as ≥50% reduction in lesion size) was 17%. While lower than the proposed threshold for favorable clinical response, the histologic response rate (improvement in histologic grade) was 60%, including 17% complete responses and 43% partial responses. Logistic regression analysis revealed that when compared to never smokers, current and former smokers had statistically significantly increased histologic responses (p=0.016). Remarkably, a significant correlation existed between decreased mTOR activity (pS6 IHC staining) in the basal epithelial layer of OPL and the histological (p=0.04) and clinical (p=0.01) responses. CONCLUSIONS. This is the first phase II trial of metformin in individuals with OPL, providing evidence that metformin administration results in encouraging histological responses and mTOR pathway modulation, thus supporting its further investigation as a chemopreventive agent.
J. Silvio Gutkind, Alfredo Molinolo, Xingyu Wu, Zhiyong Wang, Daniela Nachmanson, Olivier Harismendy, Ludmil B. Alexandrov, Beverly R. Wuertz, Frank G. Ondrey, Denise M. Laronde, Leigha D. Rock, Miriam P. Rosin, Charles S. Coffey, Valerie D. Butler, Lisa Bengtson, Chiu-Hsieh Hsu, Julie E. Bauman, Stephen M. Hewitt, Ezra E.W. Cohen, H.H. Sherry Chow, Scott M. Lippman, Eva Szabo
BACKGROUND. High circulating levels of ceramides (Cer) and sphingomyelins (SM) have been associated with cardiometabolic diseases. The consumption of whole-fat dairy products, which naturally contain such polar lipids (PL), is associated with health benefits, but the impact on sphingolipidome remains unknown. We investigated how milk PL supplementation impacts circulating and intestinal SM and Cer composition in association with improvement of cardiovascular markers. METHODS. In a 4 week-randomized double-blind controlled study, 58 postmenopausal women consumed daily a cream cheese containing 0, 3 or 5 g of milk PL. Postprandial metabolic explorations were performed before and after the supplementation. SM and Cer species were analyzed in serum, intestine-derived chylomicrons and feces. The ileal content of 4 ileostomy patients was also explored after milk PL intake in a crossover double-blind study. RESULTS. Milk PL consumption decreased serum atherogenic C24:1 Cer (Pgroup = 0.033), C16:1 (Pgroup = 0.007) and C18:1 (Pgroup = 0.003) SM species. Changes in serum C16+18 SM species were positively correlated with the reduction of total cholesterol (r = 0.706, P < 0.001), LDL-C (r = 0.666, P < 0.001) and ApoB (r = 0.705, P < 0.001). Milk PL decreased the concentration in chylomicrons of total SM (Pgroup < 0.0001) and of C24:1 Cer (Pgroup = 0.001). Saturated SM and Cer species, which are also the major species found in milk PL-enriched cheeses, increased in ileal efflux and feces. There was a marked increase in total fecal Cer after milk PL supplementation (Pgroup = 0.0002). Milk PL also modulated the abundance of some specific SM and Cer species in ileal efflux and feces, suggesting differential absorption and metabolization processes in the gut. CONCLUSION. These data demonstrate that milk PL supplementation decreases atherogenic SM and Cer species associated with an improvement of cardiovascular risk markers. Our findings bring new insights on sphingolipid metabolism in the gastrointestinal tract, especially Cer as such signaling molecules potentially participating in the beneficial effect of milk PL. ClinicalTrials.gov, NCT02099032, NCT02146339. FUNDINGS. Agence Nationale de la Recherche, ANR-11-ALID-007-01; Regional Hospital Clinical Research Program (PHRCI-2014: VALOBAB, n°14-007); French Dairy Interbranch Organization (CNIEL); Groupe Lipides et Nutrition (GLN 2018-11-07), Hospices Civils de Lyon as sponsor.
Mélanie Le Barz, Cécile Vors, Emmanuel Combe, Laurie Joumard-Cubizolles, Manon Lecomte, Florent Joffre, Michèle Trauchessec, Sandra Pesenti, Emmanuelle Loizon, Anne-Esther Breyton, Emmanuelle Meugnier, Karène Bertrand, Jocelyne Drai, Chloé Robert, Annie Durand, Charlotte Cuerq, Patrice Gaborit, Nadine Leconte, Annick Bernalier-Donadille, Eddy Cotte, Martine Laville, Stéphanie Lambert-Porcheron, Lemlih Ouchchane, Hubert Vidal, Corinne Malpuech-Brugère, David Cheillan, Marie-Caroline Michalski
BACKGROUND. Beige and brown adipose tissue (BAT) are associated with improved metabolic homeostasis. We recently reported that the β3AR agonist mirabegron induced beige adipose tissue in obese insulin resistant subjects, and this was accompanied by improved glucose metabolism. Here, we evaluated whether pioglitazone treatment, or the combination pioglitazone and mirabegron treatment, was more effective at inducing beige adipose or BAT than mirabegron treatment alone. Both drugs were used at FDA-approved dosages. METHODS. We measured BAT by PET CT scans, beige adipose tissue by immunohistochemistry, and comprehensively characterized glucose and lipid homeostasis and insulin sensitivity by euglycemic clamp and oral glucose tolerance tests. Subcutaneous white adipose tissue, muscle fiber type composition and capillary density, lipotoxicity, and systemic inflammation were evaluated by immunohistochemistry, gene expression profiling, mass spectroscopy, and ELISAs. RESULTS. Treatment with pioglitazone or the combination of pioglitazone and mirabegron increased beige adipose tissue protein marker expression and improved insulin sensitivity and glucose homeostasis, but neither treatment induced BAT in these obese subjects. When the magnitude of the responses to the treatments were evaluated, mirabegron was found to be the most effective at inducing beige adipose tissue. Although monotherapy with either mirabegron or pioglitazone induced adipose beiging, combination treatment resulted in less beiging than either alone. The three treatments also had different effects on muscle fiber type switching and capillary density. CONCLUSION. The addition of pioglitazone to mirabegron treatment does not enhance beiging or increase BAT in obese, insulin-resistant research participants. TRIAL REGISTRATION. Clinicaltrials.gov NCT02919176. FUNDING. NIH: DK112282, P20GM103527, and CTSA grant UL1TR001998.
Brian S. Finlin, Hasiyet Memetimin, Beibei Zhu, Amy L. Confides, Hemendra J. Vekaria, Riham H. El Khouli, Zachary R. Johnson, Philip M. Westgate, Jianzhong Chen, Andrew J. Morris, Patrick G. Sullivan, Esther E. Dupont-Versteegden, Philip A. Kern
BACKGROUND Recessive dystrophic epidermolysis bullosa (RDEB) is an incurable disease that causes severe mucocutaneous fragility due to mutations in COL7A1 (encoding type VII collagen [C7]). In this phase I/IIa trial, we evaluated the safety and possible clinical efficacy of intravenous infusion of allogeneic human umbilical cord blood–derived mesenchymal stem cells (hUCB-MSCs) in patients with RDEB.METHODS Four adult and two pediatric patients with RDEB were treated with 3 intravenous injections of hUCB-MSCs (1 × 106 to 3 × 106 cells/kg) every 2 weeks and followed up for 8–24 months after treatment. The primary endpoint was safety. Secondary endpoints related to efficacy included clinical parameters, such as disease severity score, wound assessment, itch and pain score, and quality of life. C7 expression levels and inflammatory infiltrates in the skin, as well as serum levels of inflammatory markers and neuropeptides, were also assessed.RESULTS Intravenous hUCB-MSC infusions were well tolerated, without serious adverse events. Improvements in the Birmingham Epidermolysis Bullosa Severity Score, body surface area involvement, blister counts, pain, pruritus, and quality of life were observed with maximal effects at 56–112 days after treatment. hUCB-MSC administration induced M2 macrophage polarization and reduced mast cell infiltration in RDEB skin. Serum levels of substance P were decreased after therapy. Increased C7 expression was observed at the dermoepidermal junction in 1 of 6 patients at day 56.CONCLUSION To the best of our knowledge, this is the first clinical trial of systemic administration of allogeneic hUCB-MSCs in patients with RDEB, demonstrating safety and transient clinical benefits.TRIAL REGISTRATION ClinicalTrials.gov NCT04520022.FUNDING This work was supported by Daewoong Pharmaceutical Co. Ltd. and Kangstem Biotech Co. Ltd.
Sang Eun Lee, Seung-Ju Lee, Song-Ee Kim, Kinam Kim, Boyoung Cho, Kyounghwan Roh, Soo-Chan Kim
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