叶酸-聚乙二醇-阿霉素连接物对肝癌裸鼠皮下移植瘤生长及心脏毒性影响的研究

doi:10.3877/cma.j.issn.2095-2015.2026.02.002

目的

观察叶酸-聚乙二醇-阿霉素（FA-PEG-hyd-DOX）连接物对肝癌裸鼠皮下移植瘤生长及心脏毒性的影响。

方法

选择4~6周龄裸鼠60只，随机分为5组，包括对照组、DOX组、连接物1组、连接物2组、连接物3组，每组12只。对照组注射生理盐水；DOX组注射DOX（剂量5.0 mg/kg），连接物1组、2组及3组注射FA-PEG-hyd-DOX连接物（剂量分别为2.5、5.0、10.0 mg/kg）。比较裸鼠给药不同时间的肿瘤体积和体质量、肿瘤组织的形态变化；比较裸鼠给药后B淋巴细胞瘤-2（Bcl-2）、Bcl-2相关X蛋白（Bax）、髓系细胞白血病-1（Mcl-1）、裂解型半胱氨酸天冬氨酸特异性蛋白酶3（cleaved caspase-3）、Ki-67的相对表达量，DOX在裸鼠心脏、肝脏以及肾脏的分布情况以及DOX给药后的血液学指标。

结果

给药20 d，对照组、DOX组、连接物1组的肿瘤体积均明显高于连接物2组和连接物3组（P<0.05）。给药20 d，DOX组和连接物3组的体质量明显低于连接物2组、连接物1组以及对照组（P<0.05）。连接物2组和连接物3组Bcl-2、Mcl-1、Ki-67表达量均明显低于DOX组和连接物1组，Bax、cleaved-caspase-3的表达明显高于对照组和连接物1组（P<0.05）。与DOX组相比，连接物2组DOX在心脏的分布减少。连接物1组、连接物2组、连接物3组CK水平均明显低于DOX组（P<0.05）。

结论

与DOX相比，FA-PEG-hyd-DOX连接物能够有效抑制肝癌裸鼠皮下移植瘤体积，具有较好的抗肿瘤效果，且能够更为有效地减少肿瘤细胞增殖，促进肿瘤细胞凋亡，减轻对心脏的损害。

关键词: 肝癌, 叶酸-聚乙二醇-阿霉素连接物, 皮下移植瘤, 裸鼠, 细胞凋亡

Objective

To investigate the effects of the folate-polyethylene glycol-doxorubicin (FA-PEG-hyd-DOX) linker on the growth and cardiac toxicity of subcutaneous transplanted tumors of nude mice with liver cancer.

Methods

A total of 60 4-6 weeks nude mice were randomly divided into 5 groups, which were control group, DOX group, linker 1 group, linker 2 group and linker 3 group, with 12 mice in each group. The control group was injected with physiological saline solution, the DOX group was injected with DOX (dose 5.0 mg/kg), while the linker 1 group, linker 2 group and linker 3 group were injected with FA-PEG-hyd-DOX linker (dose 2.5, 5.0 and 10.0 mg/kg) was injected in groups 1, 2 and 3, respectively. The tumor volume and body mass were compared in nude mice at different time points, and the morphological changes of tumor tissue in nude mice after administration were compared. The relative expression levels of B-cell lymphoma-2 (Bcl-2), Bcl2-associated X (Bax)、myeloid cell leukemia-1 (Mcl-1), cleaved cysteine-dependent aspartate-specific protease-3 (cleaved caspase-3) and Ki-67 in nude mice after administration were compared. The distribution of DOX in the hearts, livers, and kidneys of nude mice and the hematological indexes were compared.

Results

After 20 days of administration, the tumor volume of control group, DOX group and linker 1 group were significantly higher than that of linker 2 group and linker 3 group (P<0.05). After 20 days of administration, the body weight of DOX group and linker 3 group was significantly lower than that of linker 2 group, linker 1 group and control group (P<0.05). The expressions of Bcl-2, Mcl-1 and Ki-67 in linker 2 and linker 3 groups were significantly lower than those in DOX group and linker 1 group, and the expressions of Bax and cleaved caspase-3 were significantly higher than those in control group and linker 1 group (P<0.05). Compared with the DOX group, the distribution of DOX in the heart was reduced in the linker 2 group. The levels of CK in linker 1 group, linker 2 group and linker 3 group were significantly lower than those in DOX group (P<0.05).

Conclusion

Compared with DOX, the FA-PEG-hyd-DOX linker can effectively inhibit the volume of subcutaneous transplanted tumors in nude mice with liver cancer, having good anti-tumor effects, and can more effectively promote tumor cell apoptosis, reducing damage to the heart.

Key words: Liver cancer, Folate-polyethylene glycol-doxorubicin linker, Subcutaneous transplanted tumors, Nude mice, Apoptosis

图1 裸鼠肝癌模型的建立

图2 裸鼠肝癌模型不同给药时间的肿瘤体积变化注：与对照组相比，^aP<0.05；与DOX（阿霉素）组相比，^bP<0.05；与连接物1组相比，^cP<0.05

图3 裸鼠肝癌模型不同给药时间的体质量比较注：与对照组相比，^aP<0.05；与连接物1组相比，^bP<0.05；与连接物2组相比，^cP<0.05。DOX阿霉素

图4 裸鼠肝癌模型给药后肿瘤组织的变化比较注：HE×40。4A为对照组；4B为DOX（阿霉素）组；4C为连接物2组

表1 裸鼠肝癌模型给药后肿瘤组织凋亡和增殖相关蛋白表达量比较（

±s）

组别	例数	Bcl-2	Bax	Mcl-1	Ki-67	cleaved-caspase-3
对照组	12	1.24±0.11	0.30±0.03	0.60±0.02	0.88±0.06	0.79±0.02
DOX组	12	0.80±0.07^a	0.71±0.02^a	0.41±0.03^a	0.67±0.05^a	0.98±0.02^a
连接物1组	12	0.97±0.06^ab	0.56±0.02^ab	0.52±0.01^ab	0.71±0.04^ab	0.87±0.04^ab
连接物2组	12	0.41±0.04^abc	1.34±0.05^abc	0.30±0.05^abc	0.57±0.05^abc	1.13±0.05^abc
连接物3组	12	0.37±0.03^abc	1.37±0.06^abc	0.27±0.03^abc	0.53±0.06^abc	1.16±0.02^abc
F		349.076	2570.576	460.211	98.697	628.356
P		<0.001	<0.001	<0.001	<0.001	<0.001

表1 裸鼠肝癌模型给药后肿瘤组织凋亡和增殖相关蛋白表达量比较（

±s）

组别	例数	Bcl-2	Bax	Mcl-1	Ki-67	cleaved-caspase-3
对照组	12	1.24±0.11	0.30±0.03	0.60±0.02	0.88±0.06	0.79±0.02
DOX组	12	0.80±0.07^a	0.71±0.02^a	0.41±0.03^a	0.67±0.05^a	0.98±0.02^a
连接物1组	12	0.97±0.06^ab	0.56±0.02^ab	0.52±0.01^ab	0.71±0.04^ab	0.87±0.04^ab
连接物2组	12	0.41±0.04^abc	1.34±0.05^abc	0.30±0.05^abc	0.57±0.05^abc	1.13±0.05^abc
连接物3组	12	0.37±0.03^abc	1.37±0.06^abc	0.27±0.03^abc	0.53±0.06^abc	1.16±0.02^abc
F		349.076	2570.576	460.211	98.697	628.356
P		<0.001	<0.001	<0.001	<0.001	<0.001

图5 裸鼠肝癌模型给药后各器官DOX的分布比较注：红色表示DOX（阿霉素）。5A为心脏；5B为肝脏；5C为肾脏

表2 裸鼠肝癌模型给药后血液学指标的变化（

±s）

组别	鼠数	谷丙转氨酶(U/L)	谷草转氨酶(U/L)	肌酐(μmol/L)	尿素氮(mmol/L)	肌酸激酶(IU/L)
对照组	12	44.61±1.73	120.03±4.97	17.48±2.21	7.86±2.48	70.37±16.15
DOX组	12	52.06±6.02^a	135.58±16.47^a	24.96±6.03^a	13.12±3.98^a	178.32±37.11^a
连接物1组	12	49.21±2.46^a	132.87±7.48^a	22.69±1.60^a	11.41±2.38^a	90.51±15.51^ab
连接物2组	12	49.79±4.20^a	133.40±6.85^a	23.52±2.49^a	11.95±1.48^a	91.21±14.02^ab
连接物3组	12	50.63±3.72^a	134.65±5.94^a	24.42±2.20^a	12.42±2.00^a	92.75±12.17^ab
F		4.577	3.397	9.822	7.496	48.071
P		0.003	0.012	<0.001	<0.001	<0.001

表2 裸鼠肝癌模型给药后血液学指标的变化（

±s）

组别	鼠数	谷丙转氨酶(U/L)	谷草转氨酶(U/L)	肌酐(μmol/L)	尿素氮(mmol/L)	肌酸激酶(IU/L)
对照组	12	44.61±1.73	120.03±4.97	17.48±2.21	7.86±2.48	70.37±16.15
DOX组	12	52.06±6.02^a	135.58±16.47^a	24.96±6.03^a	13.12±3.98^a	178.32±37.11^a
连接物1组	12	49.21±2.46^a	132.87±7.48^a	22.69±1.60^a	11.41±2.38^a	90.51±15.51^ab
连接物2组	12	49.79±4.20^a	133.40±6.85^a	23.52±2.49^a	11.95±1.48^a	91.21±14.02^ab
连接物3组	12	50.63±3.72^a	134.65±5.94^a	24.42±2.20^a	12.42±2.00^a	92.75±12.17^ab
F		4.577	3.397	9.822	7.496	48.071
P		0.003	0.012	<0.001	<0.001	<0.001

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Effects of FA-PEG-hyd-DOX linker on the growth and cardiac toxicity of subcutaneous transplanted tumors of nude mice with liver cancer

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Effects of FA-PEG-hyd-DOX linker on the growth and cardiac toxicity of subcutaneous transplanted tumors of nude mice with liver cancer