p53, the major tumor suppressor, is frequently mutated in many cancers, and up to 84% of human melanomas harbor wild-type p53, which is considered to be an ideal target for melanoma therapy. Here, we evaluated the antitumor activity of a carbazole derivative, 9-ethyl-9H-carbazole-3-carbaldehyde (ECCA), on melanoma cells. ECCA had a selectively strong inhibitory activity against the growth of BRAF-mutated and BRAF-wild-type melanoma cells but had little effect on normal human primary melanocytes. ECCA inhibited melanoma cell growth by increasing cell apoptosis, which was associated with the upregulation of caspase activities and was significantly abrogated by the addition of a caspase inhibitor. In vivo assays confirmed that ECCA suppressed melanoma growth by enhancing cell apoptosis and reducing cell proliferation, and importantly ECCA did not have any evident toxic effects on normal tissues. RNA-Seq analysis identified several pathways related to cell apoptosis that were affected by ECCA, notably, activation of the p53 signaling pathway. Biochemical assays demonstrated that ECCA enhanced the phosphorylation of p53 at Ser15 in melanoma cells harboring wild-type p53, and importantly, the knockdown or deletion of p53 in those cells counteracted the ECCA-induced apoptosis, as well as senescence. Further investigations revealed that ECCA enhanced the phosphorylation of p38-MAPK and c-Jun N-terminal kinase (JNK), and treatment with either a p38-MAPK or a JNK inhibitor rescued the cell growth inhibition elicited by ECCA, which depended on the expression of the p53 gene. Finally, the combination of ECCA with a BRAF inhibitor significantly enhanced the growth inhibition of melanoma cells. In summary, our study demonstrates that the carbazole derivative, ECCA, induces melanoma cell apoptosis and senescence through the activation of p53 to significantly and selectively suppress the growth of melanoma cells without affecting normal human melanocytes, suggesting its potential to develop a new drug for melanoma therapy.

p53 是主要的肿瘤抑制因子，在许多癌症中经常发生突变，多达 84% 的人类黑色素瘤含有野生型 p53，它被认为是黑色素瘤治疗的理想靶点。在这里，我们评估了咔唑衍生物 9-乙基-9H-咔唑-3-甲醛 (ECCA) 对黑色素瘤细胞的抗肿瘤活性。ECCA 对 BRAF 突变型和 BRAF 野生型黑色素瘤细胞的生长具有选择性强抑制活性，但对正常人原代黑色素细胞几乎没有影响。ECCA 通过增加细胞凋亡来抑制黑色素瘤细胞的生长，这与半胱天冬酶活性的上调有关，并通过添加半胱天冬酶抑制剂显着消除。体内试验证实，ECCA 通过增强细胞凋亡和减少细胞增殖来抑制黑色素瘤的生长，重要的是，ECCA 对正常组织没有任何明显的毒性作用。RNA-Seq 分析确定了几种与受 ECCA 影响的细胞凋亡相关的通路，特别是 p53 信号通路的激活。生化分析表明，ECCA 增强了携带野生型 p53 的黑色素瘤细胞中 p53 Ser15 位点的磷酸化，重要的是，这些细胞中 p53 的敲低或缺失抵消了 ECCA 诱导的细胞凋亡和衰老。进一步的研究表明，ECCA 增强了 p38-MAPK 和 c-Jun N-末端激酶 (JNK) 的磷酸化，并且用 p38-MAPK 或 JNK 抑制剂治疗挽救了由 ECCA 引起的细胞生长抑制，这取决于表达p53 基因。最后，ECCA 与 BRAF 抑制剂的组合显着增强了黑色素瘤细胞的生长抑制。总之，我们的研究表明，咔唑衍生物 ECCA 通过激活 p53 诱导黑色素瘤细胞凋亡和衰老，显着并选择性地抑制黑色素瘤细胞的生长，而不影响正常人黑色素细胞，表明其具有开发黑色素瘤新药的潜力治疗。