Structural Biology Department, Institute of Natural and Biological Sciences, Federal University of Triângulo Mineiro, Uberaba, Brazil.
Medical Clinical Department, Institute of Health Sciences, Federal University of Triângulo Mineiro, Uberaba, Brazil.
J Oral Pathol Med. 2021 Apr;50(4):394-402. doi: 10.1111/jop.13143. Epub 2020 Dec 7.
This study evaluated the effect of treatment with TKI-258 on apoptosis, involving Rho GTPases and their effectors in SCC-4 cells of oral squamous cell carcinoma.
Markers of cell death and apoptosis were analyzed in control and TKI-258-treated SCC-4 cells by flow cytometry. The involvement of Rho GTPases and effectors in the induction of apoptosis by TKI-258 was evaluated by quantification of cleaved PARP. Also, gene expression analysis of those proteins was performed.
The treatment with TKI-258 led to a significant increase in cell death (7-AAD) and apoptosis (annexin V and cleaved PARP). When Rho GTPases were stimulated with LPA and inhibited with toxin A Clostridium difficile, the percentage of apoptotic cells increased and decreased, respectively. A similar effect was found when the treatment was with TKI-258 combined with LPA and toxin A. Treatment with TKI-258 significantly increased RhoA gene expression, while RhoB, RhoC, Rac1, and Cdc42 decreased significantly. ROCKs inhibitors (Y-27632 and HA-1077) reduced apoptosis compared with control. TKI-258 combined with Y-27632 or HA-1077 led to an increase in apoptosis compared with inhibitors only. Treatment with TKI-258 led to an increase in ROCK1 and ROCK2 gene expression, and a decrease in PAK1 and PAK2 gene expression.
TKI-258 stimulates apoptosis in SCC-4 cells of oral squamous cell carcinoma. Possibly, RhoA GTPase and their effectors ROCKs participate in the signaling pathway inhibited by TKI-258.
Therapies with multi-target inhibitors, such as TKI-258, may be promising alternatives for the clinical treatment of oral squamous cell carcinoma.
本研究评估了 TKI-258 对口腔鳞状细胞癌细胞 SCC-4 中 Rho GTPases 及其效应物凋亡的影响。
通过流式细胞术分析对照和 TKI-258 处理的 SCC-4 细胞中的细胞死亡和凋亡标志物。通过定量分析 cleaved PARP 评估 TKI-258 诱导凋亡过程中 Rho GTPases 和效应物的参与。还进行了这些蛋白质的基因表达分析。
TKI-258 治疗导致细胞死亡（7-AAD）和凋亡（Annexin V 和 cleaved PARP）显著增加。当 Rho GTPases 被 LPA 刺激，被 C. difficile 毒素 A 抑制时，凋亡细胞的百分比分别增加和减少。当用 TKI-258 联合 LPA 和毒素 A 治疗时，也发现了类似的效果。TKI-258 治疗显著增加 RhoA 基因表达，而 RhoB、RhoC、Rac1 和 Cdc42 则显著降低。ROCKs 抑制剂（Y-27632 和 HA-1077）与对照相比减少了凋亡。与单独抑制剂相比，TKI-258 联合 Y-27632 或 HA-1077 导致凋亡增加。TKI-258 治疗导致 ROCK1 和 ROCK2 基因表达增加，而 PAK1 和 PAK2 基因表达减少。
TKI-258 刺激口腔鳞状细胞癌细胞 SCC-4 凋亡。可能，RhoA GTPase 及其效应物 ROCKs 参与了 TKI-258 抑制的信号通路。
多靶点抑制剂如 TKI-258 的治疗可能是口腔鳞状细胞癌临床治疗的有前途的选择。
