scholarly journals Efficient reprogramming of the heavy-chain CDR3 regions of a human antibody repertoire

2021 ◽  
Author(s):  
Tiangling Ou ◽  
Wenhui He ◽  
Brian D Quinlan ◽  
Yan Guo ◽  
Pabalu Karunadharma ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Heavy Chain ◽  
Human Antibody ◽  
Cell Repertoire ◽  
Viral Control ◽  
Human B Cells ◽  
B Cell Repertoire ◽  
Hiv 1

B cells have been engineered ex vivo to express an HIV-1 broadly neutralizing antibody (bNAb). B-cell reprograming may be scientifically and therapeutically useful, but current approaches limit B-cell repertoire diversity and disrupt the organization of the heavy-chain locus. A more diverse and physiologic B-cell repertoire targeting a key HIV-1 epitope could facilitate evaluation of vaccines designed to elicit bNAbs, help identity more potent and bioavailable bNAb variants, or directly enhance viral control in vivo. Here we address the challenges of generating such a repertoire by replacing the heavy-chain CDR3 (HCDR3) regions of primary human B cells. To do so, we identified and utilized an uncharacterized Cas12a ortholog that recognizes PAM motifs present in human and murine JH genes. We also optimized the design of 200 nucleotide homology-directed repair templates (HDRT) by minimizing the required 3'-5' resection of the HDRT-complementary strand. Using these techniques, we edited primary human B cells to express a hemagglutinin epitope tag and the HCDR3 regions of the bNAbs PG9 and CH01. Those edited with bNAb HCDR3 efficiently bound trimeric HIV-1 antigens, implying they could affinity mature in vivo in response to the same antigens. This approach generates diverse B-cell repertoires recognizing a key HIV-1 neutralizing epitope.

scholarly journals The pre-existing human antibody repertoire to computationally optimized influenza H1 hemagglutinin vaccines

2021 ◽  
Author(s):  
Kaito Nagashima ◽  
John V Dzimianski ◽  
Julianna Han ◽  
Nada Abbadi ◽  
Aaron D Gingerich ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cross Reactivity ◽  
Influenza Viruses ◽  
Human Antibody ◽  
Protein Vaccine ◽  
Cell Repertoire ◽  
Neutralizing Activity ◽  
Human B Cells ◽  
Repertoire Sequencing

The computationally optimized broadly reactive antigen (COBRA) approach has previously been used to generate hemagglutinin (HA) immunogens for several influenza subtypes that expand vaccine-elicited antibody breadth. As nearly all individuals have pre-existing immunity to influenza viruses, influenza-specific memory B cells will likely be recalled upon COBRA HA vaccination. We determined the epitope specificity and repertoire characteristics of pre-existing human B cells to H1 COBRA HA antigens. Cross-reactivity between wild type HA and H1 COBRA HA proteins were observed at both the oligoclonal B cell level and for a subset of isolated monoclonal antibodies (mAbs). The mAbs bound five distinct epitopes on the pandemic A/California/04/2009 head and stem domains, and the majority of the mAbs had HAI and neutralizing activity against pandemic H1 strains. Two head-directed mAbs, CA09-26 and CA09-45, had HAI and neutralizing activity against a pre-pandemic H1 strain. One mAb, P1-05, targets the stem region of H1 HA proteins, but does not compete with known stem-targeting H1 mAbs. We determined that mAb P1-05 recognizes a recently discovered membrane proximal epitope on HA, the anchor epitope, and we identified similar mAbs using B cell repertoire sequencing. In addition, the trimerization domain distance from HA was critical to recognition of this epitope by P1-05. Overall, these data indicate that seasonally vaccinated individuals possess a population of functional H1 COBRA HA-reactive B cells that target head, central stalk, and anchor epitopes, and demonstrate the importance of structure-based assessment of subunit protein vaccine candidates to ensure accessibility of optimal protein epitopes.

In vivo suppression of perinatal multispecific B cells results in a distortion of the adult B cell repertoire

1986 ◽  
Vol 16 (9) ◽  
pp. 1159-1165 ◽  
Author(s):  
Meenal Vakil ◽  
Helmut Sauter ◽  
Christopher Paige ◽  
John F. Kearney
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Repertoire ◽  
B Cell Repertoire

scholarly journals A role for gut-associated lymphoid tissue in shaping the human B cell repertoire

2013 ◽  
Vol 210 (9) ◽  
pp. 1665-1674 ◽  
Author(s):  
Anna Vossenkämper ◽  
Paul A. Blair ◽  
Niloufar Safinia ◽  
Louise D. Fraser ◽  
Lisa Das ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Lymphoid Tissue ◽  
Plasma Cells ◽  
Cell Subset ◽  
Cell Repertoire ◽  
Human B Cells ◽  
B Cell Repertoire ◽  
Immature B Cells

We have tracked the fate of immature human B cells at a critical stage in their development when the mature B cell repertoire is shaped. We show that a major subset of bone marrow emigrant immature human B cells, the transitional 2 (T2) B cells, homes to gut-associated lymphoid tissue (GALT) and that most T2 B cells isolated from human GALT are activated. Activation in GALT is a previously unknown potential fate for immature human B cells. The process of maturation from immature transitional B cell through to mature naive B cell includes the removal of autoreactive cells from the developing repertoire, a process which is known to fail in systemic lupus erythematosus (SLE). We observe that immature B cells in SLE are poorly equipped to access the gut and that gut immune compartments are depleted in SLE. Thus, activation of immature B cells in GALT may function as a checkpoint that protects against autoimmunity. In healthy individuals, this pathway may be involved in generating the vast population of IgA plasma cells and also the enigmatic marginal zone B cell subset that is poorly understood in humans.

scholarly journals Anti-idiotypic antibodies elicit anti-HIV-1–specific B cell responses

2019 ◽  
Vol 216 (10) ◽  
pp. 2316-2330 ◽  
Author(s):  
Pia Dosenovic ◽  
Anna-Klara Pettersson ◽  
Abigail Wall ◽  
Eddy S. Thientosapol ◽  
Junli Feng ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibodies ◽  
Target Cells ◽  
Human B Cells ◽  
Cd4 Binding Site ◽  
Idiotypic Antibody ◽  
Anti Hiv ◽  
Hiv 1

Human anti-HIV-1 broadly neutralizing antibodies (bNAbs) protect against infection in animal models. However, bNAbs have not been elicited by vaccination in diverse wild-type animals or humans, in part because B cells expressing the precursors of these antibodies do not recognize most HIV-1 envelopes (Envs). Immunogens have been designed that activate these B cell precursors in vivo, but they also activate competing off-target responses. Here we report on a complementary approach to expand specific B cells using an anti-idiotypic antibody, iv8, that selects for naive human B cells expressing immunoglobulin light chains with 5–amino acid complementarity determining region 3s, a key feature of anti-CD4 binding site (CD4bs)–specific VRC01-class antibodies. In mice, iv8 induced target cells to expand and mature in the context of a polyclonal immune system and produced serologic responses targeting the CD4bs on Env. In summary, the results demonstrate that an anti-idiotypic antibody can specifically recognize and expand rare B cells that express VRC01-class antibodies against HIV-1.

The normal IGHV1-69–derived B-cell repertoire contains stereotypic patterns characteristic of unmutated CLL

Blood ◽  
2010 ◽  
Vol 115 (1) ◽  
pp. 71-77 ◽  
Author(s):  
Francesco Forconi ◽  
Kathleen N. Potter ◽  
Isla Wheatley ◽  
Nikos Darzentas ◽  
Elisa Sozzi ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Heavy Chain ◽  
Immunoglobulin Heavy Chain ◽  
Immunoglobulin M ◽  
Lymphocytic Leukemia ◽  
Immunoglobulin Gene ◽  
Cell Of Origin ◽  
Cell Repertoire ◽  
B Cell Repertoire

Abstract The cell of origin of chronic lymphocytic leukemia (CLL) has long been sought, and immunoglobulin gene analysis provides new clues. In the unmutated subset (U-CLL), there is increased usage of the 51p1-related alleles of the immunoglobulin heavy chain variable 1-69 gene, often combined with selected genes and with immunoglobulin heavy chain diversity IGHJ6. Stereotypic characteristics of the HCDR3 result and suggest antigen selection of the leukemic clones. We have now analyzed 51p1/IGHJ6 combinations in normal blood B cells from 3 healthy persons for parallel sequence patterns. A high proportion (33.3% of sequences) revealed stereotypic patterns, with several (15.0%) being similar to those described in U-CLL. Previously unreported CLL-associated stereotypes were detected in 4.8%. Stereotypes (13.6%) not detected in CLL also were found. The HCDR2-IGHJ6 sequences were essentially unmutated. Junctional amino acids in normal B cells were heterogeneous, as in cases of stereotyped CLL. Phenotypically, normal B cells expressing 51p1-derived immunoglobulin M were naive. This snapshot of the naive B-cell repertoire reveals subsets of B cells closely related to those characteristic of CLL. Conserved patterns in the 51p1-encoded immunoglobulin M of normal B cells suggest a restricted sequence repertoire shaped by evolution to recognize common pathogens. Proliferative pressure on these cells is the likely route to U-CLL.

scholarly journals Production of HIV-1 by Human B Cells Infectedin Vitro:Characterization of an EBV Genome-Negative B Cell Line Chronically Synthetizing a Low Level of HIV-1 after Infection

Virology ◽  
1998 ◽  
Vol 244 (2) ◽  
pp. 542-551 ◽  
Author(s):  
Laurent Fritsch ◽  
Vincent Marechal ◽  
Véronique Schneider ◽  
Corinne Barthet ◽  
Willy Rozenbaum ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Line ◽  
Low Level ◽  
Human B Cells ◽  
Hiv 1

scholarly journals Altered BCR and TLR signals promote enhanced positive selection of autoreactive transitional B cells in Wiskott-Aldrich syndrome

2015 ◽  
Vol 212 (10) ◽  
pp. 1663-1677 ◽  
Author(s):  
Nikita S. Kolhatkar ◽  
Archana Brahmandam ◽  
Christopher D. Thouvenel ◽  
Shirly Becker-Herman ◽  
Holly M. Jacobs ◽  
...  
Keyword(s):  
B Cells ◽  
Positive Selection ◽  
B Cell ◽  
Heavy Chain ◽  
High Throughput Sequencing ◽  
Cell Stage ◽  
Wiskott Aldrich Syndrome ◽  
Transitional B Cells ◽  
Selection Of

Wiskott-Aldrich syndrome (WAS) is an X-linked immunodeficiency disorder frequently associated with systemic autoimmunity, including autoantibody-mediated cytopenias. WAS protein (WASp)–deficient B cells have increased B cell receptor (BCR) and Toll-like receptor (TLR) signaling, suggesting that these pathways might impact establishment of the mature, naive BCR repertoire. To directly investigate this possibility, we evaluated naive B cell specificity and composition in WASp-deficient mice and WAS subjects (n = 12). High-throughput sequencing and single-cell cloning analysis of the BCR repertoire revealed altered heavy chain usage and enrichment for low-affinity self-reactive specificities in murine marginal zone and human naive B cells. Although negative selection mechanisms including deletion, anergy, and receptor editing were relatively unperturbed, WASp-deficient transitional B cells showed enhanced proliferation in vivo mediated by antigen- and Myd88-dependent signals. Finally, using both BCR sequencing and cell surface analysis with a monoclonal antibody recognizing an intrinsically autoreactive heavy chain, we show enrichment in self-reactive cells specifically at the transitional to naive mature B cell stage in WAS subjects. Our combined data support a model wherein modest alterations in B cell–intrinsic, BCR, and TLR signals in WAS, and likely other autoimmune disorders, are sufficient to alter B cell tolerance via positive selection of self-reactive transitional B cells.

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