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[Killing effect of Robo1 targeted Chimeric Antigen Receptor modified NK92 cells against glioma and neuroblastoma cells].
Qu, Y; Bi, J Z.
Affiliation
  • Qu Y; Department of Neural Medicine, Second Hospital of Shandong University, Jinan 250033, China.
Zhonghua Yi Xue Za Zhi ; 98(11): 860-866, 2018 Mar 20.
Article in Zh | MEDLINE | ID: mdl-29609271
ABSTRACT

Objective:

To study the cytotoxicity of Robo1-CAR-NK92 cells against U87-MG and SH-SY5Y cells, to explore the effects of IL-15, IL-21 and dexamethasone on the proliferation, survival and cytotoxicity of Robo1-CAR-NK92 cells and to optimize the culture protocol.

Methods:

Robo1-CAR-NK92 cells were constructed by lentivirus transfection.The Robo1 car positive cells were sorted, expanded and detected by flow cytometry.The levels of Robo1 expression in SH-SY5Y and U87-MG cells were examined by flow cytometry.The cytotoxicity of Robo1-CAR-NK92 or NK92 cells against target cells was tested by CCK-8 and live cell imaging. The levels of cytokines in the supernatant of cultured cells during the cytotoxicity assay were quantified by the multiplex bead-array assay.NK92 and Robo1-CAR-NK92 cells (4×10(4)/ml) were treated with 25 ng/ml of IL-15, 25 ng/ml of IL-21 and/or 50 nmol/L dexamethasone for 3 days and were stained with trypan blue to acquire the viable cell numbers and survival rates.

Results:

Robo1-CAR-NK92 cells were constructed and tested 98.89% positive after sorting and expansion. While 88.14% of U87-MG cells were Robo1 positive, there were 99.75% of Robo1 positive SH-SY5Y cells.The specific lysis of Robo1-CAR-NK92 cells against target cells was significantly higher than that of NK92 cells (P<0.05). Robo1-CAR-NK92 cells obviously secreted more cytokines including IL-6, IL-10, TNF-α and IFN-γ than parental NK92 cells during cytotoxic activity against U87-MG cells (P<0.05). IL-15 significantly increased the proliferation and survival of Robo1-CAR-NK92 cells, but IL-21 played the opposite role.Remarkably, IL-21 and IL-15+ IL-21 enhanced the cytotoxicity of NK92 and Robo1-CAR-NK92 cells.The combination of dexamethasone and interleukins dramatically promoted the proliferation and survival but obviously impaired the cytotoxicity of NK92 and Robo1-CAR-NK92 cells (except that IL+ 15 and dexamethasone have no effect on the cytotoxicity of Robo1-CAR-NK92 cells).

Conclusions:

Compared to parental NK92 cells, Robo1-CAR-NK92 cells exhibited more potent targeted killing against glioma and neuroblastoma cells.Collectively, treatment of IL-15 and dexamethasone was the optimized protocol for culture of Robo1 CAR NK cells during our experimental time.
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Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Glioma / Neuroblastoma Type of study: Guideline Limits: Humans Language: Zh Journal: Zhonghua Yi Xue Za Zhi Year: 2018 Document type: Article Affiliation country: China

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Glioma / Neuroblastoma Type of study: Guideline Limits: Humans Language: Zh Journal: Zhonghua Yi Xue Za Zhi Year: 2018 Document type: Article Affiliation country: China