A study on optimized lentiviral transduction conditions in CAR-T cells targeting CD30

Gaohui TIAN; Qinxing ZHANG; Jiangzhou SHI; Fenfang ZHAO; Ning WANG; Jiaxuan ZHAO; Yulin LU; Yao XU

doi:10.19401/j.cnki.1007-3639.2023.07.002

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A study on optimized lentiviral transduction conditions in CAR-T cells targeting CD30

Article | 更新时间：2025-12-31

- A study on optimized lentiviral transduction conditions in CAR-T cells targeting CD30
- China Oncology Vol. 33, Issue 7, Pages: 646-654(2023)
- 作者机构：
  
  1. 武汉科技大学生命科学与健康学院生物医学研究院，湖北武汉 430081
  2. 天津科技大学生物工程学院，天津 300457
- 作者简介：
  
  XU Yao.
- 基金信息：
- DOI：10.19401/j.cnki.1007-3639.2023.07.002
  CLC： R730.54
- Received：09 December 2022，
  
  Revised：2023-05-04，
  
  Published：30 July 2023
- 稿件说明：
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Gaohui TIAN, Qinxing ZHANG, Jiangzhou SHI, et al. A study on optimized lentiviral transduction conditions in CAR-T cells targeting CD30[J]. China Oncology, 2023, 33(7): 646-654.
DOI：

Gaohui TIAN, Qinxing ZHANG, Jiangzhou SHI, et al. A study on optimized lentiviral transduction conditions in CAR-T cells targeting CD30[J]. China Oncology, 2023, 33(7): 646-654. DOI： 10.19401/j.cnki.1007-3639.2023.07.002.

摘要

背景与目的：

嵌合抗原受体T（chimeric antigen receptor T，CAR-T）细胞技术在血液肿瘤治疗领域已被广泛应用，慢病毒转导是CAR-T细胞制备的关键环节，与CAR-T细胞的质量密切相关，因此慢病毒转导过程涉及的各项参数仍需进一步优化。本研究旨在探讨携带CD30抗体序列的慢病毒转导T细胞的感染复数（multiplicity of infection，MOI）、温育密度、转导前活化时间和转导体系高度对抗CD30 CAR-T细胞的影响，优化CAR-T细胞的转导条件，提高转导效率和CAR-T细胞功能。

方法：

采用不同的MOI、温育密度、转导前活化时间和转导体系高度等对人外周血来源的T细胞进行转导优化，转导后分别检测抗CD30 CAR-T细胞的增殖能力、转导效率、细胞存活率和体外杀伤效率等，以确定最优的T细胞转导条件。

结果：

MOI为1.00、1.50和3.00时转导效率和有效细胞数显著高于0.00、0.25和0.50组。在温育密度大于1.0×10

个/mL时，温育密度对T细胞转导效率无影响。活化时间为72 h组细胞存活率低于80%，显著低于其他组；24、48 h的转导效率显著高于0、8、16 h组；48 h组CAR-T细胞的增殖速率显著高于24 h组。转导体系高度为0.16 mm时转导效率和增殖倍数都显著高于0.53 mm组，但对CAR-T细胞的体外杀伤效率无影响。

结论：

通过对CAR-T细胞功能的综合评估，确定慢病毒的最佳转导条件为MOI=1、温育密度为1.0×10

个/mL、转导前活化48 h、转导体系高度为0.16 mm。

Abstract

Background and purpose:

Chimeric antigen receptor T (CAR-T) cell technology has been widely used in the field of blood tumor treatment. Lentivirus transduction is a key link in the preparation of CAR-T cells

which is closely related to the quality of CAR-T cells. Therefore

the parameters involved in the lentivirus transduction process still need to be further optimized. This study aimed t

o investigate the effects of multiplicity of infection (MOI)

incubation density

activation time before transduction and transduction system height on anti-CD30 CAR-T cells in lentiviral transduction T cells carrying CD30 antibody sequences

optimize the transduction conditions of CAR-T cells

and improve transduction efficiency and CAR-T cells function.

Methods:

Different MOI

incubation density

activation time before transduction and transduction system height were used to optimize the transduction of human peripheral blood-derived T cells

then proliferation capacity

transduction efficiency

cell viability and killing efficiency

in vitro

of anti-CD30 CAR-T cells were detected.

Results:

When the MOI was 1.00

1.50 and 3.00

the transduction efficiency and effective cell number were significantly higher than those in the 0.00

0.25 and 0.50 groups. When the incubation density was greater than 0.5×10

cells/mL

the incubation density had no significant effect on the transduction efficiency of T cells. The cell viability rate of the 72 h group was lower than 80%

which was lower compared with other groups significantly

the transduction efficiency of 24 and 48 h was significantly higher compared with the 0 h

8 h and 16 h groups

and the expansion rate of CAR-T cells in the 48 h group was significantly higher compared with the 24 h group. The transduction efficiency and proliferation capacity of 0.16 mm in height of transduction system were significantly higher compared with the 0.53 mm group

however

had no significant effect on killing efficiency of CAR-T cells

in vitro

Conclusion:

Through the comprehensive evaluation of CAR-T cells function

the optimal transduction conditions for lentivirus include MOI of 1.00

incubation density of 1.0×10

cells/mL

activation before transduction for 48 h and transduction system height of 0.16 mm.

关键词

Keywords

references

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