不同处理因素对结直肠癌类器官奥沙利铂敏感性测试结果的影响研究

doi:10.3877/cma.j.issn.1674-0793.2021.06.004

目的

选用结直肠癌临床常用药物奥沙利铂，探究不同处理因素对结直肠癌类器官药物敏感性测试结果的影响，为肿瘤患者个体化精准治疗方案的筛选提供测试条件参考。

方法

采用不同细胞密度、给药前不同细胞培养时间、不同药物作用时间对2例结直肠癌类器官CRC51、CRC67进行处理，设计100、10、1、0.1、0.01 μmol/L 5个剂量的奥沙利铂对2例结直肠癌类器官进行作用，CCK-8检测细胞增殖活力，根据细胞存活率（T/C）比较分析不同处理因素对结直肠癌类器官药物敏感性测试结果的影响。

结果

①在细胞密度的选择上，每孔接种8 000个和4 000个细胞时，100 μmol/L的奥沙利铂均对类器官CRC51、CRC67的生长产生显著的抑制作用，两者的抑制作用差异无统计学意义。而每孔接种2 000个细胞时，100 μmol/L的奥沙利铂对类器官CRC51、CRC67的生长抑制作用均显著变弱（与每孔接种8 000个和4 000个细胞相比，P<0.05），药效评价结果变为体外无效。②在给药前细胞培养时间的选择上，没有获得规律性的研究结果，但均T/C>50%。③在药物作用时间的选择上，100 μmol/L的奥沙利铂对类器官CRC51、CRC67作用7 d比作用3 d获得更加显著的生长抑制作用（P<0.05）。

结论

不同细胞密度、给药前不同细胞培养时间、不同药物作用时间对结直肠癌类器官药物敏感性测试结果均会产生影响。选择的实验条件不同，可能会得出相反的药效评价结果。

关键词: 结直肠肿瘤, 类器官, 处理因素, 奥沙利铂, 药物敏感性

Objective

To explore the effects of different processing factors on oxaliplatin sensitivity test results of colorectal cancer organoids and provide reference for the individualized precision treatment for tumor patients.

Methods

Two cases of colorectal cancer organoids were processed with different cell density, different cell culture time prior to administration and different drug action time. Five doses of oxaliplatin were designed to act on the two cases of colorectal cancer organoids. CCK-8 was used to detect cell proliferation activity. The effects of different processing factors on drug sensitivity test results of colorectal cancer organoids were analyzed according to cell survival rate.

Results

In the selection of cell density, 100 μmol/L oxaliplatin could significantly inhibit the growth of colorectal cancer organoids in both cases when inoculated with 8 000 and 4 000 cells per well, and there was no significant difference between the two groups. When inoculated with 2 000 cells per well, the growth inhibitions of 100 μmol/L oxaliplatin on colorectal cancer organoids in both cases were significantly weakened (compared with 8 000 and 4 000 cells per well, P<0.05), and the results of efficacy evaluation became invalid in vitro. No regular results were obtained on the time of cell culture prior to administration. In the choice of time of action, 100 μmol/L oxaliplatin showed more significant growth inhibition effect on the two colorectal cancer organoids after being treated for 7 days than 3 days (P<0.05).

Conclusions

Different cell density, different cell culture time prior to administration and different drug action time show influence on the drug sensitivity test results of colorectal cancer organoids. Different experimental conditions may lead to opposite efficacy evaluation results.

Key words: Colorectal neoplasms, Organoid, Processing factors, Oxaliplatin, Drug sensitivity

图1 不同细胞密度（8 000、4 000、2 000个/孔）对结直肠癌类器官药物敏感性测试结果的影响　A、B分别为CRC51各组T/C百分比柱状图、百分比-对数浓度曲线图；C、F分别为CRC51、CRC67各组结直肠癌类器官处理后形态代表图（×100）；D、E分别为CRC67各组T/C百分比柱状图、百分比-对数浓度曲线图；^***P<0.001，^** P<0.01，^* P<0.05

图2 在相同细胞密度、奥沙利铂作用3 d条件下，给药前不同细胞培养时间（4 h、1 d）对结直肠癌类器官药物敏感性测试结果的影响　A、B分别为CRC51各组T/C百分比柱状图、百分比-对数浓度曲线图；C、F分别为CRC51、CRC67各组结直肠癌类器官处理后形态代表图（×100）；D、E分别为CRC67各组T/C百分比柱状图、百分比-对数浓度曲线图；^* P<0.05

图3 在相同细胞密度、奥沙利铂作用7 d条件下，给药前不同细胞培养时间（4 h、1 d）对结直肠癌类器官药物敏感性测试结果的影响　A、B分别为CRC51各组T/C百分比柱状图、百分比-对数浓度曲线图；C、F分别为CRC51、CRC67各组结直肠癌类器官处理后形态代表图（×100）；D、E分别为CRC67各组T/C百分比柱状图、百分比-对数浓度曲线图；^***P<0.001，^**P<0.01

图4 在相同细胞密度、给药前细胞培养4 h条件下，不同药物作用时间（3、7 d）对结直肠癌类器官药物敏感性测试结果的影响　A、B分别为CRC51各组T/C百分比柱状图、百分比-对数浓度曲线图；C、F分别为CRC51、CRC67各组结直肠癌类器官处理后形态代表图（×100）；D、E分别为CRC67各组T/C百分比柱状图、百分比-对数浓度曲线图；^***P<0.001

图5 在相同细胞密度、给药前细胞培养1 d条件下，不同药物作用时间（3、7 d）对结直肠癌类器官药物敏感性测试结果的影响　A、B分别为CRC51各组T/C百分比柱状图、百分比-对数浓度曲线图；C、F分别为CRC51、CRC67各组结直肠癌类器官处理后形态代表图（×100）；D、E分别为CRC67各组T/C百分比柱状图、百分比-对数浓度曲线图；^**P<0.01

[1]	Xinaris C, Brizi V, Remuzzi G. Organoid models and applications in biomedical research[J]. Nephron, 2015, 130(3): 191-199.
[2]	Fujii M, Shimokawa M, Date S, et al. A colorectal tumor organoid library demonstrates progressive loss of niche factor requirements during tumorigenesis[J]. Cell Stem Cell, 2016, 18(6): 827-838.
[3]	Usui T, Sakurai M, Enjoji S, et al. Establishment of a novel model for anticancer drug resistance in three-dimensional primary culture of tumor microenvironment[J]. Stem Cells Int, 2016, 2016: 7053872.
[4]	Verissimo CS, Overmeer RM, Ponsioen B, et al. Targeting mutant RAS in patient-derived colorectal cancer organoids by combinatorial drug screening[J]. Elife, 2016, 5: e18489.
[5]	Vlachogiannis G, Hedayat S, Vatsiou A, et al. Patient-derived organoids model treatment response of metastatic gastrointestinal cancers[J]. Science, 2018, 359(6378): 920-926.
[6]	Christenser S, Van Der Roest B, Besselink N, et al. 5-fluorouracil treatment induces characteristic T>G mutations in human cancer[J]. Nat Commun, 2019, 10(1): 4571.
[7]	Cho YH, Ro EJ, Yoon JS, et al. 5-FU promotes stemness of colorectal cancer via p53-mediated WNT/β-catenin pathway activation[J]. Nat Commun, 2020, 11(1): 5321.
[8]	Pauli C, Hopkins B D, Prandi D, et al. Personalized in vitro and in vivo cancer models to guide precision medicine[J]. Cancer Discov, 2017, 7(5): 462-477.
[9]	Schütte M, Risch T, Abdavi-Azar N, et al. Molecular dissection of colorectal cancer in pre-clinical models identifies biomarkers predicting sensitivity to EGFR inhibitors[J]. Nat Commun, 2017, 8: 14262.
[10]	van de Wetering M, Francies HE, Francis JM, et al. Prospective derivation of a living organoid biobank of colorectal cancer patients[J]. Cell, 2015, 161(4): 933-945.
[11]	Ng SY, Tan WJ, Pek MMX, et al. Mechanically and chemically defined hydrogel matrices for patient-derived colorectal tumor organoid culture[J]. Biomaterials, 2019, 219: 119400.
[12]	Pinho D, Santos D, Vila A, et al. Establishment of colorectal cancer organoids in microfluidic-based system[J]. Micromachines-Basel, 2021, 12(5): 497.
[13]	Siegel RL, Miller KD, Fedewa SA, et al. Colorectal cancer statistics, 2017[J]. CA Cancer J Clin, 2017, 67(3): 177-193.
[14]	Usui T, Sakurai M, Umata K, et al. Hedgehog signals mediate anti-cancer drug resistance in three-dimensional primary colorectal cancer organoid culture[J]. Int J Mol Sci, 2018, 19(4): 1098.

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Effect of different processing factors on oxaliplatin sensitivity test results of colorectal cancer organoids

模态框（Modal）标题

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Effect of different processing factors on oxaliplatin sensitivity test results of colorectal cancer organoids