Engineering of Cytolethal Distending Toxin B by Its Reducing Immunogenicity and Maintaining Stability as a New Drug Candidate for Tumor Therapy; an In Silico Study

The cytolethal distending toxin (CDT), Haemophilus ducreyi, is one of the bacterial toxins that have recently been considered for targeted therapies, especially in cancer therapies. CDT is an A-B2 exotoxin. Its catalytic subunit (CdtB) is capable of inducing DNA double strand breaks, cell cycle arrest and apoptosis in host eukaryotic cells. The sequence alignment indicates that the CdtB is structurally homologyr to phosphatases and deoxyribonucleases I (DNase I). Recently, it has been found that CdtB toxicity is mainly related to its nuclease activity. The immunogenicity of CDT can reduce its effectiveness in targeted therapies. However, the toxin can be very useful if its immunogenicity is significantly reduced. Detecting hotspot ectopic residues by computational servers and then mutating them to eliminate B-cell epitopes is a promising approach to reduce the immunogenicity of foreign protein-based therapeutics. By the mentioned method, in this study, we try to reduce the immunogenicity of the CdtB- protein sequence. This study initially screened residue of the CdtB is B-cell epitopes both linearly and conformationally. By overlapping the B-cell epitopes with the excluded conserve residues, and active and enzymatic sites, four residues were allowed to be mutated. There were two mutein options that show reduced antigenicity probability. Option one was N19F, G74I, and S161F with a VaxiJen score of 0.45 and the immune epitope database (IEDB) score of 1.80, and option two was N19F, G74I, and S161W with a VaxiJen score of 0.45 and IEDB score of 1.88. The 3D structure of the proposed sequences was evaluated and refined. The structural stability of native and mutant proteins was accessed through molecular dynamic simulation. The results showed that the mutations in the mutants caused no considerable changes in their structural stability. However, mutant 1 reveals more thermodynamic stability during the simulation. The applied approaches in this study can be used as rough guidelines for finding hot spot immunogen regions in the therapeutic proteins. Our results provide a new version of CdtB that, due to reduced immunogenicity and increased stability, can be used in toxin-based drugs such as immunotoxins.

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SARS-CoV-2-induced humoral immunity through B cell epitope analysis and neutralizing activity in COVID-19 infected individuals in Japan

10.1101/2020.07.22.212761 ◽

2020 ◽

Cited By ~ 1

Author(s):

Shota Yoshida ◽

Chikako Ono ◽

Hiroki Hayashi ◽

Satoshi Shiraishi ◽

Kazunori Tomono ◽

...

Keyword(s):

B Cell ◽

Antibody Production ◽

Vaccine Development ◽

Hot Spot ◽

Cell Epitope ◽

Large Individual ◽

B Cell Epitopes ◽

S Protein ◽

Neutralizing Activity ◽

B Cell Epitope

AbstractThe aim of this study is to understand adaptive immunity to SARS-CoV-2 through the analysis of B cell epitope and neutralizing activity in coronavirus disease 2019 (COVID-19) patients. We obtained serum from thirteen COVID-19 patients. Most individuals revealed neutralizing activity against SARS-CoV-2 assessed by a pseudotype virus-neutralizing assay. The antibody production against the spike glycoprotein (S protein) or receptor-binding domain (RBD) of SARS-CoV-2 was elevated, with large individual differences, as assessed by ELISA. In the analysis of the predicted the linear B cell epitopes, two regions (671-690 aa. and 1146-1164 aa.), which were located in S1 and S2 but not in the RBD, were highly reactive with the sera from patients. In the further analysis of the B cell epitope within the S protein by utilizing a B cell epitope array, a hot spot in the N-terminal domain of the S protein but not the RBD was observed in individuals with neutralizing activity. Overall, the analysis of antibody production and B cell epitopes of the S protein from patient serum may provide a novel target for the vaccine development against SARS-CoV-2.

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Removal of B cell epitopes as a practical approach for reducing the immunogenicity of foreign protein-based therapeutics☆

Advanced Drug Delivery Reviews ◽

10.1016/j.addr.2009.07.014 ◽

2009 ◽

Vol 61 (11) ◽

pp. 977-985 ◽

Cited By ~ 54

Author(s):

Satoshi Nagata ◽

Ira Pastan

Keyword(s):

B Cell ◽

Practical Approach ◽

Foreign Protein ◽

B Cell Epitopes

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Construction of A Synthetic Gene Encoding the Multi-Epitope of Toxoplasma gondii and Demonstration of the Relevant Recombinant Protein Production: A Vaccine Candidate

Galen Medical Journal ◽

10.31661/gmj.v9i0.1708 ◽

2020 ◽

Vol 9 ◽

pp. e1708

Author(s):

Maryam Karimi ◽

Seyyed Javad Seyyed Tabaei ◽

Mohammad Mehdi Ranjbar ◽

Fardin Fathi ◽

Ali Jalili ◽

...

Keyword(s):

T Cell ◽

Toxoplasma Gondii ◽

B Cell ◽

Western Blotting ◽

3D Structure ◽

In Silico Analysis ◽

T Cell Epitopes ◽

Gene Encoding ◽

B Cell Epitopes ◽

Immunological Responses

Background: Toxoplasma gondii is a widely-distributed parasite all over the world whose attributed severe afflicting complications in human necessitate the development of serodiagnostic tests and vaccines for it. Immunological responses to monovalent vaccines and the application of diagnostic reagents including single antigens are not optimally effective. Bioinformatics approaches were used to introduce these epitopes, predict their immunogenicity and preliminarily evaluate their potential as an effective DNA vaccine and for serodiagnostic goals. Materials and Methods: A 3D structure of proteins was predicted by I-TASSER server, and linear and conformational B cell and T cell epitopes were predicted using the online servers. Then, the predicted epitopes were constructed and called Toxoeb, and their expression in the prokaryotic and eukaryotic cells was demonstrated using SDS-PAGE. In the next step, Western blotting with pooled sera of mice infected with T. gondii was done. Results: The current in silico analysis revealed that the B cell epitopes with high immunogenicity for GRA4 protein were located in the residues 34-71, and 230-266, for GRA14 in 308-387, for SAG1 in 182-195, 261-278, and for GRA7 in residues 101-120, 160-176. The T cell epitopes were selected in overlapping regions with the B cell epitopes. The immunogenic region for GRA4 are in the residues 245-253, 50-58, and 40-54, for GRA14 in 307-315, 351-359, and 308-322, for SAG1 261-269, and 259-267, and for GRA7 in the residues 103-112, and 167-175. The results of the western blotting showed that the expressed protein had immunogenicity. Conclusion: Our constructed multi-epitope of T. gondii could be considered as a candidate for diagnostic and vaccination purposes. [GMJ.2020;9:e1708]

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Faculty Opinions recommendation of Characterization of the B cell epitopes associated with a truncated form of Pseudomonas exotoxin (PE38) used to make immunotoxins for the treatment of cancer patients.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1113121.569111 ◽

2008 ◽

Author(s):

Arthur Frankel

Keyword(s):

B Cell ◽

Cancer Patients ◽

Truncated Form ◽

Pseudomonas Exotoxin ◽

B Cell Epitopes

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Exploration of Targeted Anti-tumor Therapy: a contribution to the development of targeted therapies

10.37349/etat.2020.00001 ◽

2020 ◽

Vol 1 (1) ◽

pp. 1-2

Author(s):

Nicola Normanno ◽

Graham Packham

Keyword(s):

Targeted Therapies ◽

Tumor Therapy

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Linear B-Cell Epitopes in Human Norovirus GII.4 Capsid Protein Elicit Blockade Antibodies

Vaccines ◽

10.3390/vaccines9010052 ◽

2021 ◽

Vol 9 (1) ◽

pp. 52

Author(s):

Hassan Moeini ◽

Suliman Qadir Afridi ◽

Sainitin Donakonda ◽

Percy A. Knolle ◽

Ulrike Protzer ◽

...

Keyword(s):

B Cell ◽

Capsid Protein ◽

Immune Escape ◽

Solvent Accessibility ◽

Effective Vaccine ◽

Blood Group Antigens ◽

Human Norovirus ◽

B Cell Epitopes ◽

Blocking Rate ◽

Linear B

Human norovirus (HuNoV) is the leading cause of nonbacterial gastroenteritis worldwide with the GII.4 genotype accounting for over 80% of infections. The major capsid protein of GII.4 variants is evolving rapidly, resulting in new epidemic variants with altered antigenic potentials that must be considered for the development of an effective vaccine. In this study, we identify and characterize linear blockade B-cell epitopes in HuNoV GII.4. Five unique linear B-cell epitopes, namely P2A, P2B, P2C, P2D, and P2E, were predicted on the surface-exposed regions of the capsid protein. Evolving of the surface-exposed epitopes over time was found to correlate with the emergence of new GII.4 outbreak variants. Molecular dynamic simulation (MD) analysis and molecular docking revealed that amino acid substitutions in the putative epitopes P2B, P2C, and P2D could be associated with immune escape and the appearance of new GII.4 variants by affecting solvent accessibility and flexibility of the antigenic sites and histo-blood group antigens (HBAG) binding. Testing the synthetic peptides in wild-type mice, epitopes P2B (336–355), P2C (367–384), and P2D (390–400) were recognized as GII.4-specific linear blockade epitopes with the blocking rate of 68, 55 and 28%, respectively. Blocking rate was found to increase to 80% using the pooled serum of epitopes P2B and P2C. These data provide a strategy for expanding the broad blockade potential of vaccines for prevention of NoV infection.

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Comparison of Monoclonal Gammopathies Linked to Poliovirus or Coxsackievirus vs. Other Infectious Pathogens

Cells ◽

10.3390/cells10020438 ◽

2021 ◽

Vol 10 (2) ◽

pp. 438

Author(s):

Jean Harb ◽

Nicolas Mennesson ◽

Cassandra Lepetit ◽

Maeva Fourny ◽

Margaux Louvois ◽

...

Keyword(s):

Multiple Myeloma ◽

B Cell ◽

Monoclonal Gammopathy ◽

Coat Proteins ◽

Chronic Stimulation ◽

B Cell Epitopes ◽

Monoclonal Immunoglobulins ◽

Self Antigen ◽

Coxsackievirus B1 ◽

Undetermined Significance

Chronic stimulation by infectious pathogens or self-antigen glucosylsphingosine (GlcSph) can lead to monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM). Novel assays such as the multiplex infectious antigen microarray (MIAA) and GlcSph assays, permit identification of targets for >60% purified monoclonal immunoglobulins (Igs). Searching for additional targets, we selected 28 purified monoclonal Igs whose antigen was not represented on the MIAA and GlcSph assays; their specificity of recognition was then analyzed using microarrays consisting of 3760 B-cell epitopes from 196 pathogens. The peptide sequences PALTAVETG and PALTAAETG of the VP1 coat proteins of human poliovirus 1/3 and coxsackievirus B1/B3, respectively, were specifically recognized by 6/28 monoclonal Igs. Re-analysis of patient cohorts showed that purified monoclonal Igs from 10/155 MGUS/SM (6.5%) and 3/147 MM (2.0%) bound to the PALTAVETG or PALTAAETG epitopes. Altogether, PALTAV/AETG-initiated MGUS are not rare and few seem to evolve toward myeloma.

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