A high-throughput assay for screening natural products that boost NK cell-mediated killing of cancer cells

Zihang Xu,Xiaowen Zhu,Lin Su,Chunpu Zou,Xiao Chen,Yifei Hou,Chenyuan Gong,Wanyi Ng,Zhongya Ni,Lixin Wang,Xuewei Yan,Yangzhuangzhuang Zhu,Xiaoning Jiao,Chao Yao,Shiguo Zhu

doi:10.1080/13880209.2020.1748661

Zihang Xu, Xiaowen Zhu + Show 13 more

Open Access

https://doi.org/10.1080/13880209.2020.1748661

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Abstract

Context Natural killer (NK) cells can eliminate malignant cells and play a vital role in immunosurveillance. Administration of natural compounds represents a promising approach for antitumor immunotherapy, which may enhance the NK cell activity via multiple mechanisms. Objective Establishing approaches to evaluate the effect of select natural products on NK cell-mediated cytotoxicity. Materials and methods We selected a natural product library containing 2880 pure compounds, which was provided by the National Centre for Drug Screening of China. 0.1% DMSO was employed as a negative control, and 100 U/mL human recombinant IL-2 was employed as a positive control. To evaluate the % of tumour cells which were killed by NK cells, expanded NK cells were co-cultured with tumour cells and then treated with natural products at the concentration of 10 μM. After 24-h co-incubation, luminescent signal was detected and percent lysis was calculated. Results We report on the results of a three-round high-throughput screening effort that identified 20-deoxyingenol 3-angelate (DI3A) and its analogue ingenol 3-angelate (I3A) as immuno enhancers which boosts NK cell-mediated killing of non-small cell lung cancer cells (NSCLCs). Biophotonic cytotoxicity assay and calcein release assay were used as two well-established NK cell cytotoxicity detection assays to validate the immuno-enhancing effects of DI3A and I3A, which was achieved by increasing degranulation and interferon-gamma secretion of NK cells. Conclusions Our newly established ATP-based method was a valuable and information-rich screening tool to investigate the biological effects of natural products on both NK cells and tumour cells.

Highlights

Natural killer (NK) cells are an important component of the innate immune system, which can eliminate transformed cells and virally infected cells (Guillerey et al 2016; Rezvani et al 2017; Glasner et al 2018)
We found that 406 natural products could increase the cytotoxicity of NK cells by more than 20%, and 222 natural products could increase the proliferation of NK cells by more than 20%
To validate whether ATP could be used as a sensor to determine NK cellmediated cytotoxicity, we evaluated the lysis of tumour cells by NK cells treated with either IL-2 or DMSO (Figure 3(C))

Summary

Introduction

Natural killer (NK) cells are an important component of the innate immune system, which can eliminate transformed cells and virally infected cells (Guillerey et al 2016; Rezvani et al 2017; Glasner et al 2018). Functions of NK cells are regulated by a variety of activating and inhibitory receptors at the cell surface. The tumoricidal activity of NK cells is mediated by (a) release of a variety of cytokines (such as IFN-c and TNF-a), (b) exocytosis of lytic granules containing granzymes a (Dotiwala et al 2016) and (c) expression of some death receptor ligands (Pesce et al 2017). NK cell-mediated targeting of solid tumour is usually not efficient, the tumour cells express high amounts of activating ligands and low levels of inhibitory ligands (Grossenbacher et al 2017). The escape of tumour cells from immunosuppressive cells leads to the failure of immunotherapy, especially for solid tumours in clinical settings. It is urgently necessary to develop new drugs with the ability to block the immunosuppressive environment in order to efficiently prevent the escape of tumour cells from immunosuppressive cells and restore NK cell-mediated antitumor response (Ohs et al 2017)

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