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Construction, expression and purification of a mammalian secretory recombinant fusion protein rPC / 生物工程学报
Chinese Journal of Biotechnology ; (12): 969-978, 2020.
Article in Chinese | WPRIM | ID: wpr-826879
Responsible library: WPRO
ABSTRACT
Drugs targeting immune checkpoint are used for cancer treatment, but resistance to single drug may occur. Combination therapy blocking multiple checkpoints simultaneously can improve clinical outcome. Therefore, we designed a recombinant protein rPC to block multiple targets, which consists of extracellular domains of programmed cell death protein 1 (PD-1) and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4). The coding sequence was inserted into expression vector and stably transfected into HEK293 cells. The culture supernatant was collected and rPC was affinity-purified. Real-time quantitative PCR was used to evaluate the expression levels of ligands for PD-1 and CTLA-4 in several human cancer cell lines. The binding of rPC with cancer cells was examined by immunofluorescence cell staining, the influence of rPC on cancer cell growth was assayed by CCK-8. The results showed that rPC could be expressed and secreted by stably transfected HEK293 cells, the purified rPC could bind to lung cancer NCI-H226 cells which have high levels of ligands for PD-1 and CTLA-4, no direct impact on cancer cell growth could be observed by rPC treatment. The recombinant protein rPC can be functionally assayed further for developing novel immunotherapeutic drugs for cancer.
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Full text: Available Index: WPRIM (Western Pacific) Main subject: Protein Binding / Recombinant Fusion Proteins / Humans / Cell Proliferation / HEK293 Cells / CTLA-4 Antigen / Programmed Cell Death 1 Receptor / Protein Domains / Genetics / Lung Neoplasms Language: Chinese Journal: Chinese Journal of Biotechnology Year: 2020 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Main subject: Protein Binding / Recombinant Fusion Proteins / Humans / Cell Proliferation / HEK293 Cells / CTLA-4 Antigen / Programmed Cell Death 1 Receptor / Protein Domains / Genetics / Lung Neoplasms Language: Chinese Journal: Chinese Journal of Biotechnology Year: 2020 Type: Article