丝裂原活化蛋白激酶（MAPK）途径中的激酶抑制是具有激活BRAF突变的癌症患者的标准疗法。然而,抗肿瘤作用和临床益处仅是短暂的，肿瘤容易出现治疗抵抗和复发。为了阐明耐药性的机理见解，我们建立了恶性黑色素瘤MAPK抑制剂耐药的体外细胞模型。
细胞模型是根据BRAF抑制剂的临床剂量而进化的，vemurafenib,PLX4032.我们使用RNA-Seq和RT-qPCR阵列进行转录组表达谱分析。黑素生成的途径,MAPK信号，细胞周期,与对照组相比，维罗非尼耐药细胞的一组差异表达基因之间的代谢显着富集。发现治疗抗性和通路重新布线的潜在机制是基于非基因组适应，并在两个不同的黑色素瘤模型中得到验证。SK-MEL-28和A375。两种细胞系都具有激活的BRAF突变并显示转移潜力。
双特异性磷酸酶的下调，肿瘤抑制剂，负MAPK调节剂重新接合有丝分裂信号。生长因子的上调，细胞因子,和同源受体触发信号通路绕过BRAF阻断。Further,尽管MAPK抑制剂治疗,但氨基酸和一碳代谢的变化支持细胞增殖.此外,治疗抗性细胞上调色素沉着和黑素生成，与MAPK信号部分重叠的通路。上游调节因子分析发现，致癌叉头盒和缺氧诱导因子家族转录因子存在显著扰动。
建立的细胞模型提供了对恶性黑色素瘤中抑制剂抗性下的细胞变化和治疗靶标的机制见解。在系统生物学层面，MAPK通路在获得抑制剂抗性的同时经历了重大的重新连接。这种转录可塑性的结果是选择一组转录主调节因子，绕过上游靶向激酶，并提供促有丝分裂激活的替代途径。冗余信号的精细编织网络维持相似的效应基因，从而允许肿瘤细胞存活和治疗抗性癌症的恶性进展。
Kinase inhibition in the mitogen activated protein kinase (MAPK) pathway is a standard therapy for cancer patients with activating BRAF mutations. However, the anti-tumorigenic effect and clinical benefit are only transient, and tumors are prone to treatment resistance and relapse. To elucidate mechanistic insights into drug resistance, we have established an in vitro cellular model of MAPK inhibitor resistance in malignant melanoma.
The cellular model evolved in response to clinical dosage of the BRAF inhibitor, vemurafenib, PLX4032. We conducted transcriptomic expression profiling using RNA-Seq and RT-qPCR arrays. Pathways of melanogenesis, MAPK signaling, cell cycle, and metabolism were significantly enriched among the set of differentially expressed genes of vemurafenib-resistant cells vs control. The underlying mechanism of treatment resistance and pathway rewiring was uncovered to be based on non-genomic adaptation and validated in two distinct melanoma models, SK-MEL-28 and A375. Both cell lines have activating BRAF mutations and display metastatic potential.
Downregulation of dual specific phosphatases, tumor suppressors, and negative MAPK regulators reengages mitogenic signaling. Upregulation of growth factors, cytokines, and cognate receptors triggers signaling pathways circumventing BRAF blockage. Further, changes in amino acid and one-carbon metabolism support cellular proliferation despite MAPK inhibitor treatment. In addition, treatment-resistant cells upregulate pigmentation and melanogenesis, pathways which partially overlap with MAPK signaling. Upstream regulator analysis discovered significant perturbation in oncogenic forkhead box and hypoxia inducible factor family transcription factors.
The established cellular models offer mechanistic insight into cellular changes and therapeutic targets under inhibitor resistance in malignant melanoma. At a systems biology level, the MAPK pathway undergoes major rewiring while acquiring inhibitor resistance. The outcome of this transcriptional plasticity is selection for a set of transcriptional master regulators, which circumvent upstream targeted kinases and provide alternative routes of mitogenic activation. A fine-woven network of redundant signals maintains similar effector genes allowing for tumor cell survival and malignant progression in therapy-resistant cancer.
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