Production of a granulysin-based, tn-targeted cytolytic immunotoxin using pulsed electric field technology

Guerrero-Ochoa, Patricia (Universidad de Zaragoza) ; Aguilar-Machado, D. ; Ibáñez-Pérez, Raquel (Universidad de Zaragoza) ; Macías-León, Javier (Universidad de Zaragoza) ; Hurtado-Guerrero, Ramón (Universidad de Zaragoza) ; Raso, Javier (Universidad de Zaragoza) ; Anel, Alberto (Universidad de Zaragoza)

Resumen: Granulysin is a protein present in the granules of human cytotoxic T lymphocytes (CTL) and natural killer (NK) cells, with cytolytic activity against microbes and tumors. Previous work demonstrated the therapeutic effect of the intratumoral injection of recombinant granulysin and of the systemic injection of an immunotoxin between granulysin and the anti-carcinoembryonic antigen single-chain Fv antibody fragment MFE23, which were produced in the yeast Pichia pastoris. In the present work, we developed a second immunotoxin combining granulysin and the anti-Tn antigen single-chain Fv antibody fragment SM3, that could have a broader application in tumor treatment than our previous immunotoxin. In addition, we optimized a method based on electroporation by pulsed electric field (PEF) to extract the remaining intracellular protein from yeast, augmenting the production and purificiation yield. The immunotoxin specifically recognized the Tn antigen on the cell surface. We also compared the thermal stability and the cytotoxic potential of the extracellular and intracellular immunotoxins on Tn-expressing human cell lines, showing that they were similar. Moreover, the bioactivity of both immunotoxins against several Tn+ cell lines was higher than that of granulysin alone.

Idioma: Inglés

DOI: 10.3390/ijms21176165

Año: 2020

Publicado en: International Journal of Molecular Sciences 21, 17 (2020), 6165 1-13
ISSN: 1661-6596

Factor impacto JCR: 5.923 (2020)

Categ. JCR: BIOCHEMISTRY & MOLECULAR BIOLOGY rank: 67 / 297 = 0.226 (2020) - Q1 - T1
Categ. JCR: CHEMISTRY, MULTIDISCIPLINARY rank: 49 / 178 = 0.275 (2020) - Q2 - T1

Factor impacto SCIMAGO: 1.455 - Catalysis (Q1) - Computer Science Applications (Q1) - Inorganic Chemistry (Q1) - Spectroscopy (Q1) - Molecular Biology (Q1) - Organic Chemistry (Q1) - Physical and Theoretical Chemistry (Q1) - Medicine (miscellaneous) (Q1)

Financiación: info:eu-repo/grantAgreement/ES/DGA/B31-17R
Financiación: info:eu-repo/grantAgreement/ES/DGA/E34-R17
Financiación: info:eu-repo/grantAgreement/ES/MINECO/BFU2016-75633-P
Financiación: info:eu-repo/grantAgreement/ES/MINECO/CTQ2013-44367-C2-2-P
Financiación: info:eu-repo/grantAgreement/ES/MINECO/PID2019-105451GB-I00
Financiación: info:eu-repo/grantAgreement/ES/MINECO/SAF2016-76338-R

Tipo y forma: Article (Published version)

Área (Departamento): Área Bioquímica y Biolog.Mole. (Dpto. Bioq.Biolog.Mol. Celular)
Área (Departamento): Área Biología Celular (Dpto. Bioq.Biolog.Mol. Celular)
Área (Departamento): Área Tecnología de Alimentos (Dpto. Produc.Animal Cienc.Ali.)