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PURIFY-OBS-1 is an observational study evaluating the safety and efficacy of Seraph 100® Microbind Affinity Blood Filter (Seraph 100) use for COVID-19 patients with respiratory failure admitted to the intensive care unit (ICU). The Seraph 100 is a hemoperfusion device containing heparin-coated beads that can bind to, and reduce levels of, some circulating pathogens and inflammatory molecules. This study evaluated whether treatment with the Seraph 100 affected circulating and mucosal antibody levels in critically ill COVID-19 subjects. SARS-CoV-2 anti-spike and anti-nucleocapsid IgG and IgA levels in serum were evaluated at enrollment and on days 1, 4, 7, and 28 after Seraph 100 application, while anti-spike and nucleocapsid IgG, IgA, and secretory IgA levels in tracheal aspirates were evaluated at enrollment and on days 1, 2, 3, 7, and 28. Serum samples were also collected from the pre- and post-filter lines at 1 and 4 h following Seraph 100 application to evaluate the direct impact of the filter on circulating antibody levels. Treatment with the Seraph 100 did not alter the levels of circulating or mucosal antibodies in critically ill COVID-19 subjects admitted to the ICU. [ABSTRACT FROM AUTHOR]

Introduction: Novel hemoperfusion systems are emerging for the treatment of sepsis. These devices can directly remove pathogens, pathogen-associated molecular patterns, cytokines, and other inflammatory markers from circulation. However, significant safety concerns such as potential antibiotic clearance need to be addressed prior to these devices being used in large clinical studies. Methods: Prospective, observational study of 34 participants undergoing treatment with the Seraph 100® Microbind Affinity Blood Filter (Seraph 100) device at 6 participating sites in the USA. Patients were included for analysis if they had a record of receiving an antibiotic concurrent with Seraph 100 treatment. Patients were excluded if there was missing information for blood flow rate. Blood samples were drawn pre- and post-filter at 1 h and 4 h after treatment initiation. These average pre- and post-filter time-concentration observations were then used to estimate antibiotic clearance in L/h (CLSeraph) due to the Seraph 100 device. Results: Of the 34 participants in the study, 17 met inclusion and exclusion criteria for the antibiotic analysis. Data were obtained for 7 antibiotics (azithromycin, cefazolin, cefepime, ceftriaxone, linezolid, piperacillin, and vancomycin) and one beta-lactamase inhibitor. Mean CLSeraph for the antibiotics investigated ranged from −0.57 to 0.47 L/h. No antibiotic had a CLSeraph statistically significant from 0. Discussion/Conclusion: The Seraph 100 did not significantly clear any measured antibiotic in clinical samples. These data give further evidence to suggest that these therapies may be safely administered to critically ill patients and will not impact concentrations of administered antibiotics. [ABSTRACT FROM AUTHOR]

Background: Almost half of severe hemophilia A (HA) is caused by an intron 22 inversion mutation (Int22Inv), which disrupts the 26-exon F8 gene. Inverted F8 mRNA exons 1-22 are transcribed, while F8B mRNA, containing F8 exons 23-26, is transcribed from a promoter within intron 22. Neither FVIII activity nor FVIII antigen (cross-reacting material, CRM) are detectable in plasma of patients with an intron-22 inversion. Objectives: To test the hypothesis that (putative) intracellular synthesis of FVIII proteins encoded by inverted F8 and F8B mRNAs confers T-cell tolerance to almost the entire FVIII sequence, and to evaluate the immunogenicity of the region encoded by the F8 exon 22-23 junction sequence. Patients/Methods: Peripheral blood mononuclear cells (PBMCs) from 30 severe or moderate HA subjects (17 with an Int22Inv mutation) were tested by ELISPOT assays to detect cytokine secretion in response to FVIII proteins and peptides and to map immunodominant T-cell epitopes. Potential immunogenicity of FVIII sequences encoded by the F8 exon 22-23 junction region was also tested using peptide-MHCII binding assays. Results: Eight of the Int22Inv subjects showed robust cytokine secretion from PBMCs stimulated with FVIII proteins and/or peptides, consistent with earlier publications from the Conti-Fine group. Peptide ELISPOT assays identified immunogenic regions of FVIII. Specificity for sequences encoded within F8 mRNA exons 1-22 and F8B mRNA was confirmed by staining Int22Inv CD4+ T cells with peptide-loaded HLA-Class II tetramers. FVIII peptides spanning the F8 exon 22-23 junction (encoding M2124-V2125) showed limited binding to MHCII proteins and low immunogenicity, with cytokine secretion from only one Int22Inv subject. Conclusions: PBMCs from multiple subjects with an Int22Inv mutation, with and without a current FVIII inhibitor, responded to FVIII epitopes. Furthermore, the FVIII region encoded by the exon 22-23 junction sequence was not remarkably [ABSTRACT FROM AUTHOR]

Background The generation of interferon-gamma (IFN-γ) by MHC class II activated CD4+ T helper cells play a substantial contribution in the control of infections such as caused by Mycobacterium tuberculosis. In the past, numerous methods have been developed for predicting MHC class II binders that can activate T-helper cells. Best of author's knowledge, no method has been developed so far that can predict the type of cytokine will be secreted by these MHC Class II binders or T-helper epitopes. In this study, an attempt has been made to predict the IFN-γ inducing peptides. The main dataset used in this study contains 3705 IFN-γ inducing and 6728 non-IFN-γ inducing MHC class II binders. Another dataset called IFNgOnly contains 4483 IFN-γ inducing epitopes and 2160 epitopes that induce other cytokine except IFN-γ. In addition we have alternate dataset that contains IFN-γ inducing and equal number of random peptides. Results It was observed that the peptide length, positional conservation of residues and amino acid composition affects IFN-γ inducing capabilities of these peptides. We identified the motifs in IFN-γ inducing binders/peptides using MERCI software. Our analysis indicates that IFN-γ inducing and non-inducing peptides can be discriminated using above features. We developed models for predicting IFN-γ inducing peptides using various approaches like machine learning technique, motifs-based search, and hybrid approach. Our best model based on the hybrid approach achieved maximum prediction accuracy of 82.10% with MCC of 0.62 on main dataset. We also developed hybrid model on IFNgOnly dataset and achieved maximum accuracy of 81.39% with 0.57 MCC. Conclusion Based on this study, we have developed a webserver for predicting i) IFN-γ inducing peptides, ii) virtual screening of peptide libraries and iii) identification of IFN-γ inducing regions in antigen (http://crdd.osdd.net/raghava/ifnepitope/). Reviewers This article was reviewed by Prof Kurt Blaser, Prof Laurence Eisenlohr and Dr Manabu Sugai. [ABSTRACT FROM AUTHOR]

Background: Pulmonary tuberculosis (TB) is caused by Mycobacterium tuberculosis (M. tb). In lungs, alveolar macrophages and type II alveolar epithelial cells serve as a replicative niche for this pathogen. Secretory proteins released by actively replicating tubercle bacilli are known to interact with host cells at the initial stages of infection. To understand the role of these cells in TB pathogenesis, it is important to identify the mycobacterial components involved in interaction with alveolar epithelial cells. Materials and Methods: We fractionated the whole secretory proteome of M. tb H37Rv into 10 narrow molecular mass fractions (A1-A10; <20 kDa to >90 kDa) that were studied for their binding potential with A549; type II alveolar epithelial cell line. We also studied the consequences of this interaction in terms of change in epithelial cell viability by MTT assay and cytokine release by ELISA. Results: Our results show that several mycobacterial proteins bind and confer cytolysis in epithelial cells. Amongst all the fractions, proteins ranging from 35- 45 kDa (A5) exhibited highest binding to A549 cells with a consequence of cytolysis of these cells. This fraction (A5) also led to release of various cytokines important in anti-mycobacterial immunity. Conclusion: Fraction A5 (35-45 kDa) of mycobacterial secretory proteome play an important role in mediating M. tb interaction with type II alveolar epithelial cells with the consequences detrimental for the TB pathogenesis. Further studies are being carried out to identify the candidate proteins from this region. [ABSTRACT FROM AUTHOR]

Background: Almost half of severe hemophilia A (HA) is caused by an intron 22 inversion mutation (Int22Inv), which disrupts the 26-exon F8 gene. Inverted F8 mRNA exons 1-22 are transcribed, while F8B mRNA, containing F8 exons 23-26, is transcribed from a promoter within intron 22. Neither FVIII activity nor FVIII antigen (cross-reacting material, CRM) are detectable in plasma of patients with an intron-22 inversion., Objectives: To test the hypothesis that (putative) intracellular synthesis of FVIII proteins encoded by inverted F8 and F8B mRNAs confers T-cell tolerance to almost the entire FVIII sequence, and to evaluate the immunogenicity of the region encoded by the F8 exon 22-23 junction sequence., Patients/methods: Peripheral blood mononuclear cells (PBMCs) from 30 severe or moderate HA subjects (17 with an Int22Inv mutation) were tested by ELISPOT assays to detect cytokine secretion in response to FVIII proteins and peptides and to map immunodominant T-cell epitopes. Potential immunogenicity of FVIII sequences encoded by the F8 exon 22-23 junction region was also tested using peptide-MHCII binding assays., Results: Eight of the Int22Inv subjects showed robust cytokine secretion from PBMCs stimulated with FVIII proteins and/or peptides, consistent with earlier publications from the Conti-Fine group. Peptide ELISPOT assays identified immunogenic regions of FVIII. Specificity for sequences encoded within F8 mRNA exons 1-22 and F8B mRNA was confirmed by staining Int22Inv CD4 + T cells with peptide-loaded HLA-Class II tetramers. FVIII peptides spanning the F8 exon 22-23 junction (encoding M2124-V2125) showed limited binding to MHCII proteins and low immunogenicity, with cytokine secretion from only one Int22Inv subject., Conclusions: PBMCs from multiple subjects with an Int22Inv mutation, with and without a current FVIII inhibitor, responded to FVIII epitopes. Furthermore, the FVIII region encoded by the exon 22-23 junction sequence was not remarkably immunoreactive and is therefore unlikely to contain an immunodominant, promiscuous CD4 + T-cell epitope. Our results indicate that putative intracellular expression of partial FVIII proteins does not confer T-cell tolerance to FVIII regions encoded by inverted F8 mRNA or F8B mRNA., Competing Interests: Author KP is an inventor on patents related to FVIII immunogenicity. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gunasekera, Vir, Karim, Ragni and Pratt.)