背景:黑色素瘤,一种高度恶性的皮肤癌,是肿瘤治疗研究的热门话题。如今,肿瘤免疫治疗,特别是免疫疗法与其他疗法的结合,引起了越来越多的关注。吲哚胺2,3-双加氧酶2(IDO2),免疫抑制犬尿液中色氨酸代谢途径的一种酶,在黑色素瘤组织中高度表达。此外,IDO2显著抑制机体抗肿瘤免疫,已成为黑色素瘤治疗的新靶点。硝呋嗪,作为一种肠道抗菌剂,发现能够抑制Stat3表达并发挥抗肿瘤作用。因此,本研究旨在检查自行设计的IDO2-小干扰RNA(siRNA)通过减毒沙门氏菌联合硝呋嗪对黑素瘤小鼠的治疗效果,以及确定其潜在机制。
方法:用流式细胞术检测硝呋嗪对黑色素瘤的影响,CCK-8和集落形成能力测定,分别,在体外。构建siRNA-IDO2质粒,建立小鼠黑素瘤模型。治疗后,监测肿瘤生长和存活率,HE染色检测肿瘤组织的形态学变化。Westernblotting检测相关蛋白的表达,IHC和IF检测肿瘤组织中CD4和CD8阳性T细胞的表达,流式细胞术检测脾脏中CD4和CD8阳性T细胞的比例。
结果:结果表明,联合治疗可有效抑制黑色素瘤细胞中Stat3的磷酸化和IDO2的表达水平,能有效抑制肿瘤生长,延长荷瘤小鼠的生存时间。机械研究表明,与对照组和单药治疗组相比,联合治疗组减少了肿瘤细胞的异型性,增加了凋亡率,增强肿瘤组织中T淋巴细胞的浸润,增加脾脏中的CD4+和CD8+T淋巴细胞,提示其机制可能与抑制肿瘤细胞增殖有关,细胞凋亡的增加和细胞免疫的增强。
结论:结论:IDO2-siRNA联合nifuroxazide治疗可在黑素瘤小鼠的治疗中发挥重要作用。提高肿瘤免疫力,为临床上寻找治疗黑色素瘤的新组合方法提供实验依据。
Melanoma, a highly malignant skin cancer, is a hot topic in oncology treatment research. Nowadays, tumor immunotherapy, especially immunotherapy combined with other therapies, has attracted more and more attention. Indoleamine 2,3-dioxygenase 2 (IDO2), a ratelimiting enzyme of the tryptophan metabolism pathway in the urine of dogs with immunosuppression, is highly expressed in melanoma tissue. Additionally, IDO2 significantly inhibits the anti-tumor immunity of the body and has become a novel target of melanoma treatment.
Nifuroxazide, as an intestinal antibacterial agent, was found to be able to inhibit Stat3 expression and exert an anti-tumor effect. Therefore, the present study aimed to examine the therapeutic effect of a self-designed IDO2-small interfering RNA (siRNA) delivered by attenuated Salmonella combined with
nifuroxazide on melanoma- bearing mice, as well as determine its underlying mechanism.
The effect of
nifuroxazide on melanoma was detected by flow cytometry, CCK-8 and colony- forming ability assays, respectively, in vitro. The plasmid of siRNA-IDO2 was constructed, and the mice-bearing melanoma model was established. After the treatment, the tumor growth and survival rate were monitored, and the morphological changes of tumor tissue were detected by HE staining. The expression of related proteins was detected by Western blotting, and the expression of CD4 and CD8 positive T cells in tumor tissue was detected by IHC and IF, and the proportion of CD4 and CD8 positive T cells in spleen was detected by flow cytometry.
The results demonstrated that the combination therapy effectively inhibited the phosphorylation of Stat3 and the expression level of IDO2 in melanoma cells, which effectively inhibited tumor growth and prolonged the survival time of tumor-bearing mice. The mechanistic study revealed that, compared with control groups and monotherapy groups, the combination treatment group reduced the atypia of tumor cells, increased the apoptotic rate, enhanced the infiltration of T lymphocytes in tumor tissue and increased the CD4+ and CD8+ T lymphocytes in the spleen, suggesting that the mechanism may be associated with the inhibition of tumor cell proliferation, the increase of apoptosis and the enhancement of the cellular immunity.
In conclusion, IDO2-siRNA combined with
nifuroxazide therapy could serve a significant role in the treatment of melanoma-bearing mice, enhance the tumor immunity and provide an experimental basis for identifying a novel combination method for the treatment of melanoma clinically.