Why Do We Still Lack a COVID-19 Vaccine? Searching for the Missing Pieces

According to a recent newspaper report, two Israeli companies, namely MigVax Corp. and the Israel Institute for Biological Research in Ness Ziona, have made progress on their own coronavirus vaccines (28 July, 2020; https://www.jpost.com/health-science/coronavirus-vaccine-whatprotection- might-be-available-to-israelis-soon-636639). They are on the front line in this field because they had already developed an in-house vaccine to fight a coronavirus infection that appeared 4 years ago, causing tracheobronchitis in poultry (www.migal.org.il/7010). As the chicken coronavirus exhibits a high homology (80%) with severe acute respiratory syndrome" (SARS) coronavirus-2 (SARS-CoV-2) https://www.migal.org.il/en/coronavirus-vaccine-project, responsible for the current pandemic in humans, the chicken vaccine might also work in humans with a few modifications (www.jpost.com/HEALTH-SCIENCE

CD20: A target antigen for immunotherapy of autoimmune diseases

This article reviews the role of CD20 antigen in B cell function and the effectiveness and limits of passive immunotherapy with anti-CD20 monoclonal antibody (Rituximab) in the treatment of autoimmune (or immune-mediated) diseases. Active immunotherapy is a more feasible way to control these chronic diseases. A peptide that mimics the CD20 epitope recognized by Rituximab is employed to stimulate the host immune response against CD20. (c) 2005 Elsevier B.V. All rights reserved

Clinical correlates of a subset of anti-CENP-A antibodies cross-reacting with FOXE3p53-62 in systemic sclerosis

INTRODUCTION: In a subset of patients with limited cutaneous (lc) systemic sclerosis (SSc), anti-CENP-A antibodies (Ab) cross-react with a peptide (FOXE3p53-62) that presents striking homology with one of the two immunodominant epitopes of CENP-A (Ap17-30). We searched for clinical correlates of anti-FOXE3p53-62 Ab by measuring their levels along with those of Ab to Ap17-30 and to the second immunodominant epitope of CENP-A, namely Ap1-17. METHODS: Serum samples were obtained from 121 patients with SSc, 46 patients with systemic lupus erythematosus (SLE) and 25 healthy blood donors (HBD). The reactivity of serum IgG to Ap1-17, Ap17-30 and FOXE3p53-62 was measured by ELISA. The corresponding anti-peptide Ab were affinity-purified from pooled SSc sera and used to establish standard curves for quantifying these Ab in patients and HBD. Receiver operating characteristics (ROC) analysis, comparing SSc patients who were positive for anti-CENP Ab (ACA+) to those who were negative, was used to find cut-off points for dichotomizing the anti-peptide Ab levels into positive and negative. Clinical records were reviewed to extract demographic data and information about organ involvement and disease activity. RESULTS: Of 121 SSc sera, 75 were ACA+; 88.0% of these samples reacted with Ap1-17, 82.6% with Ap17-30 and 53.3% with FOXE3p53-62. Among the 46 ACA- SSc sera, 2.2% reacted with Ap1-17, 4.3% with Ap17-30 and 11% with FOXE3p53-62. The levels of these Ab were low in ACA-, SLE and HBD groups and not significantly different among them. When ACA+ SSc patients were divided into subgroups positive or negative for anti-FOXE3p53-62 Ab, the only variables that were significantly different between groups were the levels of anti-Ap17-30 Ab and disease activity index (DAI). There was a significant association between negativity for anti-FOXE3p53-62 Ab and active disease defined as either DAI ≥3 (Fisher exact test, P = 0.045) or less restrictive DAI≥2.5 (P = 0.009). CONCLUSIONS: ACA+-Anti-FOXE3p53-62+Ab identifies a subgroup of patients with lcSSc who are less likely to develop active disease. In lc SSc patients at presentation, anti-FOXE3p53-62+ can be a marker with prognostic significance

Dietary cholesterol supplementation and inhibitory factor 1 serum levels in two dizygotic Smith-Lemli-Opitz syndrome twins: a case report

Smith-Lemli-Opitz syndrome (SLOS) is a rare genetic neurodevelopmental disorder caused by the defect in the 7-dehydrocholesterol reductase. This defect leads to the deficiency of cholesterol biosynthesis with accumulation of 7-dehydrocholesterol. Inhibitory factor 1 (IF1) is a well-known mitochondrial protein. Recently, it has been discovered in the human serum where it is reported to be involved in the HDL-cholesterol intake. Here we report the IF1 presence in the serum of two paediatric SLOS dizygotic twins treated with dietary cholesterol supplementation

Identification of an antigenic and immunogenic motif expressed by two 7-Mer Rituximab-specific cyclic peptides. Journal of Immunology

Two 7-mer cyclic peptides, bearing the antigenic motif recognized by the anti-CD20 mAb Rituximab and differing because of motif-surrounding amino acids, inhibit the binding of Rituximab to raft-associated CD20 and Rituximab-induced membrane ceramide on human lymphoid DAUDI cells. These peptides displayed different immunogenic profiles, in that antibodies recognizing CD20 were induced in two and five out of five BALB/c mice immunized with Rp-15-C and Rp13-C respectively. Analysis of immunogenic motif, performed by panning a 7-mer phage display peptide library with purified anti-peptide IgGs, showed that the motif defined by anti-Rp15-C mostly included amino acids surrounding the Rituximab-specific antigenic motif "ANPS", whereas that defined by anti-Rp13-C was "NPS". These data showed that the motif-surrounding amino acids can markedly influence the specificity of antibodies, even when elicited with a short 7-mer peptide. Since anti-peptides antibodies analyzed are IgG, their specificity is likely to reflect how peptides are processed at the T cell level and suggests that, within a short peptide, the motif defined by T cells during the recognition phase may be different from that recognized by these cells upon their stimulation. Our findings can explain the failure of most peptide-based immunotherapy in cancer and autoimmune diseases and suggest strategies to implement the specificity of peptides-induced antibodies against the target antige

EXPRESSION OF THE TRANSCRIPTION FACTOR FORKHEAD BOX E3 (FOXE3) IN MONOCYTES FROM PATIENTS WITH SYSTEMIC SCLEROSIS AND CORRELATION WITH THEIR SEROLOGICAL PROFILE

Background: The process of epithelial_mesenchymal transition (EMT) has been regarded in systemic sclerosis (SSc) as one of the possible mechanisms favouring tissue accumulation of monocyte_derived fibrocytes or myofibroblasts, which contribute to tissue fibrosis [1]. Forkhead box E3 (FOXE3) is a transcription factor involved in EMT of lens epithelial cells (LEC). Its expression progressively decreases with the migration of LEC from the anterior to the equatorial region. FOXE3 expression cessation marks initiation of fiber differentiation, suggesting that the loss of FOXE3 expression favors a pro_fibrotic phenotype [2]. No data are available on mRNA FOXE3 expression in sites other than LEC. Objectives: In this study, we investigated the FOXE3 mRNA expression in unstimulated and TGF_ß_ or IL_4_stimulated monocytes from SSc patients and healthy blood donors (HBD), to established whether i) FOXE3 is constitutively expressed in human monocytes; ii) FOXE3 expression can be modulated in vitro by cytokines involved in SSc profibrotic process; iii) there is any association between FOXE3 expression and a particular SSc serological profile. Methods: PBMC were isolated from heparinized peripheral blood of 9 patients with SSc (5 Scl70 + ; 4 Scl70 – ), and 3 HBD by Ficoll_Hypaque density gradient centrifugation. Monocytes (CD14+) were isolated by positive selection using microbeads. Cells (1x10 6 cells/ml) were stimulated TGF_ß (10 ng/ml) and IL_4 (40 ng/ml) for 14 days. mRNA was extracted and semi_quantitative PCR was performed to assess FOXE3 expression. GM_CSF stimulation (50ng/ml) was used as positive control. The levels of FOXE3 mRNA were quantified by normalizing its expression against that of GAPDH. Expression was measured as mean relative expression level (MREL). Variation of expression was measured as mean fold change (MFC). Results: Similar baseline levels of FOXE3 mRNA was observed in unstimulated CD14 + cells from SSc patients and HBD (MREL SSc=0.32; HBD=0.26). As expected, GM_CSF stimulation of CD14 + cells from SSc patients and HBD markedly up_regulated FOXE3 expression (SSc: MFC=3.24; HBD: MFC=1.84). TGF_ß and IL_4 behaved similarly to GM_CSF in enhancing FOXE3 expression in CD14 + cells from all HBD (MFC TGF_ß =1.35; MFC IL_4 =1.59) and from 3 out 4 Scl70 – patients (MFC TGF_ß =2.36; MFC IL_4 =2.9), being the expression unchanged in the remaining Scl70 – patient. By contrast, in the 4 Scl70 + patients, CD14 + FOXE3 expression markedly decreased following these cytokines stimulation (MFC TGF_ ß =0.28; MFC IL_4 =0.31) Conclusions: This is the first study to demonstrate FOXE3 mRNA expression in monocytes from HBD and SSc patients, and its differential expression following TGF_ß and IL_4 stimulation, correlating with the serological profile of SSc patients. The data suggest that the down_regulation of FOXE3 induced by TGF_ß and IL_4 may direct monocytes toward a more profibrotic phenotype in Scl70 + as compared to Scl70 – patients. The relationship of this finding with the anti_FOXE3 antibodies recently detected in SSc sera [3], remains to be determined. References: Postlethwaite AE et al. Curr Opin Rheumatol 16:733_738, 2004. Landgren H et al. Invest Ophthalmol Vis Sci 49:4269_4277, 2008. Perosa F et al. Arthritis Res Ther 15:R72, 2013. Acknowledgements: This work was supported by a 2013 grant from the Italian Group for Systemic Sclerosis (GILS), Milan, Italy. Disclosure of Interest: None declared DOI: 10.1136/annrheumdis_2014_eular.4130 Citation: Ann Rheum Dis 2014;73(Suppl2

Generation of biologically active linear and cyclic peptides has revealed a unique fine specificity of rituximab and its possible cross-reactivity with acid sphingomyelinase-like phosphodiesterase 3b precursor

Heterogeneity of the effector functions displayed by rituximab and other anti-CD20 monoclonal antibodies (mAbs) apparently recognizing the same CD20 epitope suggests that additional mechanisms, probably related to mAb fine specificity, are responsible for B-cell depletion. To improve our understanding of rituximab's function, its fine specificity was investigated by means of phage display peptide library (PDPL)-expressing 7-mer cyclic (c7c) or 7-/12-mer linear peptides. Rituximab-specific c7c PDPL-derived clone insert sequences expressed the motif A(S)NPS overlapping the human CD20 (170)ANPS(173). P-172 was the most critical for rituximab binding, since its replacement with S-172 (of mouse CD20) abolished the reactivity. The WPXWLE motif expressed by the linear PDPL-derived clone insert sequences could only be aligned to the reverse-oriented (WPXWLE156)-W-161 of acid sphingomyelinase-like phosphodiesterase 3b precursor (ASMLPD), though linear peptides bearing WPXWLE competed with cyclic ones for rituximab-paratope binding. Anti-CD20 mAb 1F5 only displayed a reactivity profile similar to that of rituximab, which also reacted with ASMLPD-derived peptides. Peptides induced antibodies with specificity and effector functions similar to those of rituximab. Our results show a unique fine specificity of rituximab, define the molecular basis for the lack of rituximab reactivity with mouse CD20 (mCD20), and the potential of targeting CD20 in an active immunotherapy setting. A possible rituximab interaction with ASMLPD is suggested