Cloning and recombinant expression of the Immunoglobulin-like variable domains of humans PD-1 and PD-L1 immunoreceptors

Authors

Keywords:

Cloning, Recombinant protein, PD-1, PD-L1, Immunoreceptor

Abstract

In cancer immunotherapy, the use of monoclonal antibodies (MAbs) that block the interaction of PD-1/PD-L1 immunoreceptors improves patient treatment. For MAb production, the availability of the protein antigen is a critical aspect. In this scenario, biomedical biotechnology allows the production of almost inexhaustible quantities of recombinant versions of these proteins. In this context, our objective was to clone and express the immunoglobulin-like variable domains (IgV) of PD-1 and PD-L1, because the interaction between them occurs among their IgV extracellular domains, specific primers were designed to clone them. Once cloned, they were individually ligated to pcDNA6 and introduced into Escherichia coli, and positive transformants were defined by PCR and sequencing. With the respective transformants, pilot expressions induced by 1 mM IPTG were performed, and the optimal time of expression was defined by electrophoresis in 15 % polyacrylamide gels. The cloning results were products of 375 bp for PD-1 and 357 bp for PD-L1, whose sequences showed 99% identity for PD-1 (GenBank OM363223) and 100% for PD-L1 (GenBank OM363224). Both proteins showed an optimal expression time of 4 h post-induction and ~14 kDa consistent with estimates. In conclusion, the cloning and recombinant expression of the IgV extracellular domains of PD-1 and PD-L1 was achieved, which will allow the production of a MAb or other immunotherapeutic tool against cancer, something that has not yet been achieved in Mexico.

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Author Biographies

  • González P, Laboratorio de Biomedicina Molecular, CAC Biotecnología Biomédica UAS-264, Posgrado en Ciencias Biomédicas y Posgrado en Biotecnología en Salud, UAS, México..

    Priscilla González is a researcher in training, born in Moreno Valley, Riverside, California, United States. Priscilla completed her Bachelor's degree in Genomic Biotechnology at the FCQB-UAS, graduating with Thesis in 2019, assigned to the Lab. Molecular Biomedicine (LBM), her work led to an article indexed in JCR doi.org/10.1016/j.ram. 2022.05.008. Later he studied the Master's Degree in Biomedical Sciences at our faculty, where he graduated with Honors in 2021, he also developed his Thesis at the LBM, and from his experimental work, he derived this manuscript. Currently, she has been accepted to the Doctorate in Biomedical Sciences of the San Diego Graduate Division of the University of California, which begins next fall

  • Beltrán-López E., Laboratorio de Biomedicina Molecular, UAS, México.

    Evangelina Beltrán, MSc, is a Pharmaceutical Chemist (QFB) from the Facultad de Ciencias Químico Biológicas (FCQB) of the UNiversidad Autónoma de Sinaloa (UAS), and holds a Master's degree in Forensic Sciences from the Marist University of Guadalajara, Jalisco. She has been a professor at the FCQB since May 2009 and has seven publications, six of which are indexed in Journal Citation Reports (JCR) journals. Currently, Evangelina is a third-semester doctoral student in Biomedical Sciences at the FCQB, affiliated with the Molecular Biomedicine Laboratory.

  • López-Moreno H. M., Facultad de Ciencias Químico Biológicas-Universidad Autónoma de Sinaloa, México.

    Ph.D. López-Moreno has a degree in Chemist, Pharmaceutical Biology from the Faculty of Chemical-Biological Sciences (FCQB) of the Autonomous University of Sinaloa, and a Doctor of Science from the Direct Doctorate in Biomedical Sciences, UNAM. He is a “C” Senior Research Professor assigned to the FCQB-UAS since 2004. Member of the National System of Researchers (Level I); Name of QFB Generation 2012-2017; Evaluator of the Mexican Council for the Accreditation of Pharmaceutical Education (COMAEF) AC, from October 2010 to date; International Evaluator of the National Council for University Evaluation and Accreditation of Panama (CONEAUPA), from 2012 to date; PRODEP evaluator, SEP; CONAHCYT Project Evaluator; PROFAPI Project Evaluator, UAS; Technical Manager of 17 research projects (9 with external financing to the UAS); Peer reviewer of the journal “Vector Borne and Zoonotic Diseases”, the journal “Frontiers in Immunology” and the journal Biomedica of Colombia; Leader of the CAC Biomedical Biotechnology UAS-264; He cultivates two Lines of research: 1. Biomedical Biotechnology and 2. Apoptosis of Giardia lamblia; University Counselor of the UAS, from 2009-2010; Career Manager at QFB in 2008 and 2011; Honorary Member of the Sinaloense System of Researchers and Technologists; PRODEP Profile, SEP. He has supervised 55 Theses (4 Doctorates, 17 Master's, and 34 Bachelor's). He has published 38 Scientific Articles indexed in JCR, 18 Memoirs in Extenso, 13 Dissemination Publications, and 60 Presentations at National and International Congresses. He has 330 scientific citations.

     

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Cloning and recombinant expression of the Immunoglobulin-like variable domains of humans PD-1 and PD-L1 immunoreceptors

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Published

2024-07-09

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No. 2 (2024): QUIBIOUAS Revista de Ciencias Químico Biológicas

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González, P., Beltrán López , E. ., & López Moreno , H. S. . (2024). Cloning and recombinant expression of the Immunoglobulin-like variable domains of humans PD-1 and PD-L1 immunoreceptors. QUIBIOUAS Revista De Ciencias Químico Biológicas, 2, 1-10. https://revistas.uas.edu.mx/index.php/QBU/article/view/598