格尔达霉素是一种苯醌类的氨基糖苷，从水浸链霉菌中分离得到。它可以抑制酪氨酸激酶和热休克蛋白90（HSP90）。将格尔达霉素和其11个衍生物与HSP90进行分子对接，发现17-去甲氧基-17-N,N-二甲氨基-格尔达霉素（17-DMAG）是与之结合亲和力最高（-7.73±0.12 kcal/mol）且抑制常数最低（2.16±0.49 μM）的化合物。因此，与格尔达霉素相比，选择17-DMAG作为进一步实验的对象。多药耐药现象是癌症治疗成功的一个主要问题。我们测试了格尔达霉素和17-DMAG对不同耐药性癌细胞系的作用。虽然格尔达霉素和17-DMAG抑制了所有被测细胞系的增殖，但多药耐药的P-糖蛋白高表达CEM/ADR5000细胞显示了耐药性；过表达ΔEGFR的肿瘤细胞和p53基因敲除细胞对这两种化合物显示了敏感性。我们进行了COMPARE和层次聚类分析，并鉴定出60个基因，可以预测59个NCI肿瘤细胞系对格尔达霉素和17-DMAG的敏感性或耐药性。根据细胞系的mRNA表达谱进行的分布分析显示，对这两种化合物的敏感性或耐药性具有统计学意义。此外，还运用生物信息学工具研究了格尔达霉素和17-DMAG的潜在作用机制。利用Galaxy Cistrome分析在候选基因的启动子区域预测转录因子结合位点，发现NF-κB DNA结合位点（Rel）是最主要的转录因子。此外，对这60个基因进行了Ingenuity Pathway Analysis（IPA）以研究与这些基因相关的信号通路相互作用。有趣的是，IPA还揭示NF-κB信号通路是这些基因中的主要网络。最后，NF-κB报告基因试验验证了生物信息学的预测，格尔达霉素和17-DMAG在接触后24小时显著抑制了NF-κB活性。总之，格尔达霉素和17-DMAG对不同的肿瘤细胞系具有细胞毒性活性。它们的活性不仅限于HSP90，还涉及到NF-κB途径的参与。Copyright © 2023. Published by Elsevier B.V.

Geldanamycin is an ansamycin-derivative of a benzoquinone isolated from Streptomyces hygroscopicus. It inhibits tyrosine kinases and heat shock protein 90 (HSP90). Geldanamycin and 11 derivatives were subjected to molecular docking to HSP90, and 17-desmethoxy-17-N,N-dimethylamino-geldanamycin (17-DMAG) was the compound with the highest binding affinity (-7.73 ± 0.12 kcal/mol) and the lowest inhibition constant (2.16 ± 0.49 μM). Therefore, 17-DMAG was selected for further experiments in comparison to geldanamycin. Multidrug resistance (MDR) represents a major problem for successful cancer therapy. We tested geldanamycin and 17-DMAG against various drug-resistant cancer cell lines. Although geldanamycin and 17-DMAG inhibited the proliferation in all cell lines tested, multidrug-resistant P-glycoprotein-overexpressing CEM/ADR5000 cells were cross-resistant, ΔEGFR-overexpressing tumor cells and p53 knockout cells were sensitive to these two compounds. COMPARE and hierarchical cluster analyses were performed, and 60 genes were identified to predict the sensitivity or resistance of 59 NCI tumor cell lines towards geldanamycin and 17-DMAG. The distribution of cell lines according to their mRNA expression profiles indicated sensitivity or resistance to both compounds with statistical significance. Moreover, bioinformatic tools were used to study possible mechanisms of action of geldanamycin and 17-DMAG. Galaxy Cistrome analyses were carried out to predict transcription factor binding motifs in the promoter regions of the candidate genes. Interestingly, the NF-ĸB DNA binding motif (Rel) was identified as the top transcription factor. Furthermore, these 60 genes were subjected to Ingenuity Pathway Analysis (IPA) to study the signaling pathway interactions of these genes. Interestingly, IPA also revealed the NF-ĸB pathway as the top network among these genes. Finally, NF-ĸB reporter assays confirmed the bioinformatic prediction, and both geldanamycin and 17-DMAG significantly inhibited NF-κB activity after exposure for 24 h. In conclusion, geldanamycin and 17-DMAG exhibited cytotoxic activity against different tumor cell lines. Their activity was not restricted to HSP90 but indicated an involvement of the NF-KB pathway.Copyright © 2023. Published by Elsevier B.V.