Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method

Detalhes bibliográficos
Autor(a) principal: Ribeiro, Priscila Rafaela
Data de Publicação: 2019
Outros Autores: Chicaybam, Leonardo, Pretti, Marco Antônio, Bonamino, Martín Hernán
Tipo de documento: Artigo de conferência
Idioma: eng
Título da fonte: Repositório Institucional da FIOCRUZ (ARCA)
Texto Completo: https://www.arca.fiocruz.br/handle/icict/32831
Resumo: Os autores do resumo não possuem vínculo com a Fiocruz, entretanto, ao submeterem o trabalho no evento organizado por Bio-Manguinhos, cedem automaticamente os direitos à unidade.
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spelling Ribeiro, Priscila RafaelaChicaybam, LeonardoPretti, Marco AntônioBonamino, Martín Hernán2019-04-29T14:19:53Z2019-04-29T14:19:53Z2019RIBEIRO, Priscila Rafaela et al. Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method. In: INTERNATIONAL SYMPOSIUM ON IMMUNOBIOLOGICAL, 4., 2019; SEMINÁRIO ANUAL CIENTÍFICO E TECNOLÓGICO DE BIO-MANGUINHOS, 7., 2019, Rio de Janeiro. Anais... Rio de Janeiro: Bio-Manguinhos, 2019. p. 173.https://www.arca.fiocruz.br/handle/icict/3283110.35259/isi.sact.2019_32831Os autores do resumo não possuem vínculo com a Fiocruz, entretanto, ao submeterem o trabalho no evento organizado por Bio-Manguinhos, cedem automaticamente os direitos à unidade.Os autores externos submeteram sua publicação para apresentação de trabalho no evento “International Symposium on Immunobiologicals”, que foi coordenado e organizado pelo Instituto de Tecnologia em Imunobiológicos (Bio-Manguinhos), da Fundação Oswaldo Cruz.Instituto Nacional de Câncer José Alencar Gomes da Silva. Rio de Janeiro, RJ, Brasil.Instituto Nacional de Câncer José Alencar Gomes da Silva. Rio de Janeiro, RJ, Brasil.Instituto Nacional de Câncer José Alencar Gomes da Silva. Rio de Janeiro, RJ, Brasil.Instituto Nacional de Câncer José Alencar Gomes da Silva. Rio de Janeiro, RJ, Brasil.Introduction: Inhibitory receptors such as PD-1, LAG-3 and CTLA-4 have gained special attention as potential targets for immunotherapy, since manipulation of negative signals mediated by these receptors may provide new therapeutic options for several diseases, as cancer. LAG-3 was described as a cell surface molecule interacting with MHC class II molecules. Identifying how proteins transduce the signal from these receptors has been a challenge but, once identified, these molecules can also be targets for novel therapeutics. In 2012, a method called BioID was developed based on the fusion of a protein of interest to a mutated biotin ligase (R118G), which has the ability to add biotin to molecules that are at 20 nm or less from the protein of interest. Once biotinylated, the proteins can be recovered and identified by mass spectrometry. Objective: To perform a screening of proteins interacting with LAG-3 through the BioID method. Methodology: chimeric antigen receptors (CARs) were constructed with the anti-CD20 scFv fused to the intracellular domains consisting of: Lag-3 WT, Lag-3 Kmut, Lag3 EPdel (deleted EP domain), all fused to the BirA domain, with further induction of expression in the HEK293T and Jurkat cell lines. Flow cytometry analysis will be performed to verify the CAR’s expression in the cells, and immunofluorescence assay to analyze the localization of the CARs. Results: The CAR anti-CD20/Lag3 WT-BirA was electroporated in the HEK293T cells presenting 80% of expression. CARs Ep del and Kmut showed 39% and 41% of CAR expression. By immunofluorescence staining it was possible to observe the cytoplasmatic localization of the CARs. Analysis of the biotinylation pattern by Western blotting was performed for CAR anti-CD20/Lag3WT-BirA, and the expected ladder pattern of biotinylation was observed. Conclusion: The constructed CARs were expressed in the target cell lines leading to the expected biotinilated patterns.engFiocruz/Instituto de Tecnologia em Imunobiológicoshttps://www.arca.fiocruz.br/handle/icict/32894BioIDReceptores antígenos quiméricosLAG-3BioIDChimeric antigens receptorsLAG-3Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID methodinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da FIOCRUZ (ARCA)instname:Fundação Oswaldo Cruz (FIOCRUZ)instacron:FIOCRUZLICENSElicense.txttext/plain1748https://www.arca.fiocruz.br/bitstream/icict/32831/1/license.txt8a4605be74aa9ea9d79846c1fba20a33MD51ORIGINALORT_18.pdfapplication/pdf47562https://www.arca.fiocruz.br/bitstream/icict/32831/2/ORT_18.pdf297b9f28440132f14ddf4fbe78e4fcc8MD52TEXTORT_18.pdf.txtORT_18.pdf.txtExtracted texttext/plain2387https://www.arca.fiocruz.br/bitstream/icict/32831/3/ORT_18.pdf.txt0d0844b513ab5e012ec7ce29a7bb9d9dMD53icict/328312023-04-26 16:59:02.569oai:www.arca.fiocruz.br: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Repositório InstitucionalPUBhttps://www.arca.fiocruz.br/oai/requestrepositorio.arca@fiocruz.bropendoar:21352023-04-26T19:59:02Repositório Institucional da FIOCRUZ (ARCA) - Fundação Oswaldo Cruz (FIOCRUZ)false
dc.title.pt_BR.fl_str_mv Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
title Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
spellingShingle Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
Ribeiro, Priscila Rafaela
BioID
Receptores antígenos quiméricos
LAG-3
BioID
Chimeric antigens receptors
LAG-3
title_short Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
title_full Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
title_fullStr Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
title_full_unstemmed Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
title_sort Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method
author Ribeiro, Priscila Rafaela
author_facet Ribeiro, Priscila Rafaela
Chicaybam, Leonardo
Pretti, Marco Antônio
Bonamino, Martín Hernán
author_role author
author2 Chicaybam, Leonardo
Pretti, Marco Antônio
Bonamino, Martín Hernán
author2_role author
author
author
dc.contributor.author.fl_str_mv Ribeiro, Priscila Rafaela
Chicaybam, Leonardo
Pretti, Marco Antônio
Bonamino, Martín Hernán
dc.subject.other.pt_BR.fl_str_mv BioID
Receptores antígenos quiméricos
LAG-3
topic BioID
Receptores antígenos quiméricos
LAG-3
BioID
Chimeric antigens receptors
LAG-3
dc.subject.en.pt_BR.fl_str_mv BioID
Chimeric antigens receptors
LAG-3
description Os autores do resumo não possuem vínculo com a Fiocruz, entretanto, ao submeterem o trabalho no evento organizado por Bio-Manguinhos, cedem automaticamente os direitos à unidade.
publishDate 2019
dc.date.accessioned.fl_str_mv 2019-04-29T14:19:53Z
dc.date.available.fl_str_mv 2019-04-29T14:19:53Z
dc.date.issued.fl_str_mv 2019
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/conferenceObject
format conferenceObject
status_str publishedVersion
dc.identifier.citation.fl_str_mv RIBEIRO, Priscila Rafaela et al. Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method. In: INTERNATIONAL SYMPOSIUM ON IMMUNOBIOLOGICAL, 4., 2019; SEMINÁRIO ANUAL CIENTÍFICO E TECNOLÓGICO DE BIO-MANGUINHOS, 7., 2019, Rio de Janeiro. Anais... Rio de Janeiro: Bio-Manguinhos, 2019. p. 173.
dc.identifier.uri.fl_str_mv https://www.arca.fiocruz.br/handle/icict/32831
dc.identifier.doi.pt_BR.fl_str_mv 10.35259/isi.sact.2019_32831
identifier_str_mv RIBEIRO, Priscila Rafaela et al. Screening of proteins related to the immunological checkpoint lymphocyte activation gene-3 (lag-3) through the BioID method. In: INTERNATIONAL SYMPOSIUM ON IMMUNOBIOLOGICAL, 4., 2019; SEMINÁRIO ANUAL CIENTÍFICO E TECNOLÓGICO DE BIO-MANGUINHOS, 7., 2019, Rio de Janeiro. Anais... Rio de Janeiro: Bio-Manguinhos, 2019. p. 173.
10.35259/isi.sact.2019_32831
url https://www.arca.fiocruz.br/handle/icict/32831
dc.language.iso.fl_str_mv eng
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dc.publisher.none.fl_str_mv Fiocruz/Instituto de Tecnologia em Imunobiológicos
publisher.none.fl_str_mv Fiocruz/Instituto de Tecnologia em Imunobiológicos
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