切换至 "中华医学电子期刊资源库"

中华肺部疾病杂志(电子版) ›› 2024, Vol. 17 ›› Issue (06) : 901 -906. doi: 10.3877/cma.j.issn.1674-6902.2024.06.009

论著

新型OX40 抗体激动剂对NSCLC 人源化小鼠模型的抑制作用及机制分析
刘岩1, 赵沛妍1, 田琳1, 崔贺然2, 狄娜2, 李慧1,(), 程颖1,3,()   
  1. 1.130012 长春,吉林省肿瘤医院肿瘤转化医学实验室
    2.130012 长春,吉林省肿瘤医院生物样本库
    3.130012 长春,吉林省肿瘤医院胸部肿瘤内科
  • 收稿日期:2024-06-17 出版日期:2024-12-25
  • 通信作者: 李慧, 程颖
  • 基金资助:
    吉林省科技厅基础研究专项(202002063JC)吉林省卫健委项目(2021JC094)

Anti-tumor activity and mechanism of the novel OX40 antibody agonist in humanized mouse NSCLC model

Yan Liu1, Peiyan Zhao1, Lin Tian1, Heran Cui2, Na Di2, Hui Li1,(), Ying Cheng1,3,()   

  1. 1.Medical Oncology Translational research lab,Jilin Cancer Hospital,Changchun 130012,China
    2.Biobank,Jilin Cancer Hospital, Changchun 130012, China
    3.Department of Medical Thoracic Oncology, Jilin Cancer Hospital,Changchun 130012, China
  • Received:2024-06-17 Published:2024-12-25
  • Corresponding author: Hui Li, Ying Cheng
引用本文:

刘岩, 赵沛妍, 田琳, 崔贺然, 狄娜, 李慧, 程颖. 新型OX40 抗体激动剂对NSCLC 人源化小鼠模型的抑制作用及机制分析[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(06): 901-906.

Yan Liu, Peiyan Zhao, Lin Tian, Heran Cui, Na Di, Hui Li, Ying Cheng. Anti-tumor activity and mechanism of the novel OX40 antibody agonist in humanized mouse NSCLC model[J/OL]. Chinese Journal of Lung Diseases(Electronic Edition), 2024, 17(06): 901-906.

目的

分析肿瘤坏死因子受体超家族4(tumor necrosis factor receptor superfamily member 4)(OX40)抗体激动剂(ES102)对小细胞肺癌(non-small cell lung cancer,NSCLC)人源化OX40 小鼠模型的抑制作用和机制。

方法

使用流式细胞术检测小鼠Lewis 肺癌细胞(Lewis lung cancer cells,LLC)中CD44 表达;分别构建OX40 人源化小鼠NSCLC 荷瘤模型,溶媒对照组、不同剂量ES102 组和结肠癌荷瘤模型,阳性溶媒对照组和阳性对照组,判断每组中ES102 的抑瘤水平;采用ELISA 检测血清中干扰素-γ(interferon-γ,IFN-γ)和白介素-17(interleukin-17,IL-17);蛋白质印迹法(Western blot) 检测PI3K、NFAT、CD44、NapsinA 和SYN 蛋白表达;RNA 转录组测序检测免疫浸润细胞表达。

结果

LLC 细胞中CD44 阳性率99.58%,NapsinA 蛋白低表达,提示LLC 鼠源肺癌细胞株具有NSCLC 特性。 人源化OX40小鼠NSCLC 模型中,ES102 对NSCLC 抑瘤能力无显著差异,MC38 阳性对照组中注射ES102 的第11 天、第14 天、第18 天和第21 天,ES102 抑瘤能力与阳性溶媒对照组相比增强(P=0.0004,t=5.712;P<0.0001,t=9.368;P<0.0001,t=10.64;P<0.0001,t=13.37)。 人源化小鼠血浆IL-17 的含量在不同样本中表达差异显著(F=7.703,P=0.0004);阳性对照组中IL-17 分泌含量(58.24±18.11)pg/ml 分别高于ES102(2 mg/ml)组(21.87±11.74)pg/ml,ES102(5 mg/ml)组(18.49±10.17)pg/ml 和ES102(10 mg/ml)组(20.47±10.52)pg/ml(P<0.05);NSCLC ES102(10 mg/ml)组中IFN-γ 含量高于对照组、ES102(2 mg/ml)组和ES102(5 mg/ml)组(P<0.05)。 PI3K 在阳性对照组中未见表达,在未处理组和ES102 实验组(2 μg/kg~10 μg/kg)中高表达(P<0.05)。 NFAT 作为活化型T 细胞的核因子,表达趋势与PI3K 一致,组间差异无统计学意义(P>0.05)。 转录组测序检测NSCLC 和阳性对照组小鼠肿瘤组织,转录组测序和反卷积分析显示,Tregs 细胞浸润比例为(1.95±0.02)%,低于阳性对照组中Tregs 细胞浸润比例(7.2±0.03)%(P<0.05)。

结论

ES102 对NSCLC 人源化小鼠模型疗效低于结肠癌,疗效差异与微环境中IL-17分泌降低及调节性T 细胞浸润有关。

Objective

To explore the efficacy and mechanism of OX40 antibody agonist (ES102) in humanized OX40 mouse NSCLC model.

Methods

Mouse Lewis lung cancer cells were detected using flow cytometry (Lewis lung cancer cells,CD44 expression in LLC); OX40 humanized mouse NSCLC tumor-bearing model (vehicle control group,ES102 treatment group at different doses) and colon cancer tumor-bearing model(positive vehicle control group and positive control group) were constructed respectively,To evaluate the tumor suppressive level of ES102 in different groups; Serum interferon- γ was measured by ELISA (interferon- γ,IFN- γ) and interleukin-17 (interleukin-17,IL-17) secretory level; Expression of PI3K,NFAT,CD44,NapsinA and SYN by western blot (Western blot); The expression of the immune-infiltrating cells was determined by RNA transcriptome sequencing.

Results

The CD44 positive rate in LLC cells showed 99.58%and low expression of NapsinA protein,suggesting that LLC murine-derived lung cancer cell lines had NSCLC properties.In the humanized OX40 mouse NSCLC model,ES102 inhibited NSCLC,but on Day 11,Day 14,Day 18,and Day 21 in the positive vehicle control (P=0.0004, t=5.712; P<0.0001, t=9.368; P<0.0001,t=10.364; P<0.0001, t=13.37).The content of plasma IL-17 in humanized mice was significantly differently expressed between different samples (F=7.703, P=0.0004); The secretion content of IL-17 in the positive control group (58.24±18.11)pg/ml than the ES102 (2 mg/ml) group (21.87±11.74)pg/ml,The ES102(5 mg/ml) group (18.49±10.17)pg/ml and the ES102 (10 mg/ml) group (20.47±10.52)pg/ml(P<0.05);Higher IFN- γ in NSCLC ES102 (10 mg/ml) than the control,ES102 (2 mg/ml) and ES102 (5 mg/ml)(P<0.05).PI3K expression was not seen in the positive control group and was highly expressed in the untreated group and the ES102 experimental group (2 μg/kg to 10 μg/kg)(P<0.05).As a nuclear factor in activated T cells,NFAT was expressed in a trend consistent with PI3K,without a statistically significant difference between the groups(P>0.05).After transcriptome sequencing of NSCLC and positive control mice,further transcriptome sequencing and deconvolution analysis showed that the proportion of Tregs cells infiltrated was (1.95±0.02)%,which was lower than that of Tregs cells infiltrated in the positive control group (7.2±0.03)%(P<0.05).

Conclusion

The efficacy of ES102 in humanized NSCLC mice is lower than colon cancer,and the difference may be related to the decreased secretion of IL-17 and the infiltration of Tregs cells in the microenvironment.

图1 CD44、SYN、NapsinA 在LLC 中的表达。 注:A:阴性对照组中CD44 含量占比;B:CD44 在LLC 细胞中的含量占比;C:CD44、SYN、NapsinA 抗体在LLC 和SBC-5 细胞中的表达水平
图2 OX40 人源化小鼠NSCLC 和结肠癌移植瘤模型对ES102 敏感性。 注:A:ES102 处理后NSCLC 移植瘤模型平均肿瘤体积;B:ES102 处理后结肠癌移植瘤模型平均肿瘤体积;C:ES102 处理后NSCLC 移植瘤模型体质量变化;D:ES102 处理后结肠癌移植瘤模型体质量变化;***:P<0.001
图3 western blot 检测不同PI3K 及NFAT 蛋白表达。 注:A:Western blot 检测泳道图;B、C:柱状图分析各蛋白的相对表达量,**: P<0.01
图4 转录组测序分析NSCLC 和阳性对照组肿瘤组织。 注:A:反卷积热图;B:柱状图分析免疫细胞比例;*:P<0.05
1
郑荣寿,陈 茹,韩冰峰,等.2022 年中国恶性肿瘤流行情况分析[J].中华肿瘤杂志,2024,46(3):221-231.
2
吴国明,钱桂生.非小细胞肺癌靶向治疗研究进展及新理念[J/CD].中华肺部疾病杂志(电子版),2019,12(4):405-408.
3
孟芸畅,许 可,宋 勇.新辅助免疫治疗在可切除非小细胞肺癌中的研究进展[J/CD].中华肺部疾病杂志(电子版),2023,16(5):734-738.
4
Sathish G,Monavarshini LK,Sundaram K,et al.Immunotherapy for lung cancer[J].Pathol Res Pract,2024,254:155104.
5
Lahiri A,Maji A,Potdar PD,et al.Lung cancer immunotherapy:progress,pitfalls,and promises[J].Mol Cancer,2023,22(1):40.
6
Swart M,Verbrugge I,Beltman JB.Combination approaches with immune-checkpoint blockade in cancer therapy[J].Front Oncol,2016,6:233.
7
Sharma P,Allison JP.The future of immune checkpoint therapy[J].Science,2015,348(6230):56-61.
8
Chen DS,Mellman I.Elements of cancer immunity and the cancerimmune set point[J].Nature,2017,541(7637):321-330.
9
Dostert C,Grusdat M,Letellier E,et al.The TNF family of ligands and receptors:Communication modules in the immune system and beyond[J].Physiol Rev,2019,99(1):115-160.
10
Lu X.OX40 and OX40L interaction in Cancer[J].Curr Med Chem,2021,28(28):5659-5673.
11
Buchan SL,Rogel A,Al-Shamkhani A.The immunobiology of CD27 and OX40 and their potential as targets for cancer immunotherapy[J].Blood,2018,131(1):39-48.
12
Thapa B,Kato S,Nishizaki D,et al.OX40/OX40 ligand and its role in precision immune oncology[J].Cancer Metastasis Rev,2024,43(3):1001-1013.
13
Xu Y,Yu Q.E-cadherin negatively regulates CD44-hyaluronan interaction and CD44-mediated tumor invasion and branching morphogenesis[J].J Biol Chem,2003,278(10):8661-8668.
14
Diab A,Hamid O,Thompson JA,et al.A phase I,open-label,dose-escalation study of the OX40 agonist ivuxolimab in patients with locally advanced or metastatic cancers[J].Clin Cancer Res,2022,28(1):71-83.
15
Davis EJ,Martin-Liberal J,Kristeleit R,et al.First-in-human phaseⅠ/Ⅱ,open-label study of the anti-OX40 agonist INCAGN01949 in patients with advanced solid tumors[J].J Immunother Cancer,2022,10(10):e004235.
16
Kuang Z,Jing H,Wu Z,et al.Development and characterization of a novel anti-OX40 antibody for potent immune activation[J].Cancer Immunol Immunother,2020,69(6):939-950.
17
Buchan SL,Rogel A,Alshamkhani A.The immunobiology of CD27 and OX40 and their potential as targets for cancer immunotherapy[J].Blood,2017,131(1):39-48.
18
Aspeslagh S,Postel-Vinay S,Rusakiewicz S,et al.Rationale for anti-OX40 cancer immunotherapy[J].Eur J Cancer,2016,52:50-66.
19
Webb GJ,Hirschfield GM,Lane PJ.OX40,OX40L and autoimmunity:A comprehensive review[J].Clin Rev Allergy Immunol,2016,50(3):312-332.
20
Lin Y,Song Y,Zhang Y,et al.NFAT signaling dysregulation in cancer:Emerging roles in cancer stem cells [ J].Biomed Pharmacother,2023,165:115167.
21
Lv YW,Chen Y,Lv HT,et al.Kawasaki disease OX40-OX40L axis acts as an upstream regulator of NFAT signaling pathway[J].Pediatr Res,2019,85(6):835-840.
22
Zhou Z,Lin L,An Y,et al.The combination immunotherapy of TLR9 agonist and OX40 agonist via intratumoural injection for hepatocellular carcinoma[J].J Hepatocell Carcinoma,2021,8:529-543.
23
Iriki H,Takahashi H,Amagai M.Diverse role of OX40 on T cells as a therapeutic target for skin diseases[J].J Invest Dermatol,2023,143(4):545-553.
24
Yan LH,Liu XL,Mo SS,et al.OX40 as a novel target for the reversal of immune escape in colorectal cancer[J].Am J Transl Res,2021,13(3):923-934.
25
Alvim RG,Georgala P,Nogueira L,et al.Combined OX40 agonist and PD-1 inhibitor immunotherapy improves the efficacy of vascular targeted photodynamic therapy in a urothelial tumor model[J].Molecules,2021,26(12):3744.
26
Ruby CE,Yates MA,Hirschhorn-Cymerman D,et al.Cutting Edge:OX40 agonists can drive regulatory T cell expansion if the cytokine milieu is right[J].J Immunol,2009,183(8):4853-4857.
27
Imianowski CJ ,Kuo P,Whiteside SK,et al.IFNγ production by functionally reprogrammed tregs promotes antitumor efficacy of OX40/CD137 bispecific agonist therapy[J].Cancer Res Commun,2024,4(8):2045-2057.
28
Kitamura N,Murata S,Ueki T,et al.OX40 costimulation can abrogate Foxp3+ regulatory T cell-mediated suppression of antitumor immunity[J].Int J Cancer,2009,125(3):630-638.
29
Weinberg AD,Morris NP,Kovacsovics-Bankowski M,et al.Science gone translational:the OX40 agonist story[J].Immunol Rev,2011,244(1):218-231.
30
Polesso F,Sarker M,Weinberg AD,et al.OX40 agonist tumor immunotherapy does not impact regulatory T cell suppressive function[J].J Immunol,2019,203(7):2011-2019.
31
Curti BD,Kovacsovics-Bankowski M,Morris N,et al.OX40 is a potent immune-stimulating target in late-stage cancer patients[J].Cancer Res,2013,73(24):7189-7198.
32
Jensen SM,Maston LD,Gough MJ,et al.Signaling through OX40 enhances antitumor immunity[J].Semin Oncol,2010,37(5):524-532.
33
Nuebling T,Schumacher CE,Hofmann M,et al.The immune checkpoint modulator OX40 and its ligand OX40L in NK-cell immunosurveillance and acute myeloid leukemia [ J].Cancer Immunol Res,2018,6(2):209-221.
34
Shibahara I,Saito R,Zhang R,et al.OX40 ligand expressed in glioblastoma modulates adaptive immunity depending on the microenvironment:A clue for successful immunotherapy[J].Mol Cancer,2015,14:41.
35
Weinberg AD,Morris NP,Kovacsovics-Bankowski M,et al.Science gone translational:the OX40 agonist story[J].Immunol Rev,2011,244(1):218-231.
[1] 张礼江, 沈玲佳, 施我大. 倾向性评分匹配分析奥希替尼对晚期NSCLC 预后的影响[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(05): 820-822.
[2] 杨慧, 郭丽娟, 冯晓丹, 李静, 黄成谋, 蔡兴锐, 覃英娇, 王远礼. 非小细胞肺癌铂类药物耐药mi RNA表达特征及预测分析[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(05): 719-724.
[3] 赖淼, 景鑫, 李桂珍, 李怡. 非小细胞肺癌EGFR 突变亚型的临床病理和预后意义[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(05): 731-737.
[4] 梁丽斯, 李洁, 贺帅, 来艳君, 刘铭, 张琳. MMP-9、MMP-2 及TLR4、HE4对非小细胞肺癌早期诊断意义[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(05): 756-761.
[5] 赵蒙蒙, 黄洁, 余荣环, 王葆青. 过表达小GTP酶Rab32抑制非小细胞肺癌细胞侵袭性生长[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(04): 512-518.
[6] 张桂萍, 丘勇林, 湛绮婷, 孙乐栋. 晚期非小细胞肺癌血清Ape1/Ref-1对放射性肺损伤发生的预测意义[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(04): 519-523.
[7] 韩晓宇, 李柯育, 赵志菲, 高建平. SNHG17过表达对非小细胞肺癌切除术预后的意义[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(04): 543-547.
[8] 刘松, 张进召, 贾艳云. 帕博利珠单抗治疗晚期非小细胞肺癌反应降低与抗生素预处理的关系[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(04): 553-557.
[9] 李多, 郝昭昭, 陈延伟, 南岩东. 血清PTX3表达与非小细胞肺癌骨转移的相关性分析[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(04): 558-562.
[10] 崔伟, 邓屹, 叶苏意, 李静, 陈晓明, 张靖, 许荣德. 载药微球支气管动脉化疗栓塞术治疗罕见非小细胞肺癌的临床疗效和安全性分析[J/OL]. 中华介入放射学电子杂志, 2024, 12(04): 303-310.
[11] 崔伟, 叶苏意, 邓屹, 陈晓明, 张靖, 李静, 许荣德. 载药微球支气管动脉化疗栓塞术治疗难治性非小细胞肺癌的临床疗效及安全性[J/OL]. 中华介入放射学电子杂志, 2024, 12(04): 311-316.
[12] 崔伟, 李静, 陈晓明, 张靖, 邓屹, 许荣德. 载药微球支气管动脉化疗栓塞术治疗非小细胞肺癌的研究进展[J/OL]. 中华介入放射学电子杂志, 2024, 12(04): 289-295.
[13] 蔡剑桥, 蒋雷. 单孔胸腔镜与开胸双袖式肺叶切除治疗非小细胞肺癌对比[J/OL]. 中华胸部外科电子杂志, 2024, 11(04): 225-230.
[14] 张迅夫, 马金山, 蒋云龙, 加纳提·托勒恒, 侯昌剑, 萨伍提·斯拉吉丁. GATA3在非小细胞肺癌组织中的表达及临床病理意义[J/OL]. 中华胸部外科电子杂志, 2024, 11(03): 175-179.
[15] 李子健, 王锐, 钟云鹏, 张迪轩, 梁韵娟, 杨超, 何建行, 李树本. 自体肺移植术在胸部恶性肿瘤中的临床应用[J/OL]. 中华胸部外科电子杂志, 2024, 11(03): 193-200.
阅读次数
全文


摘要