Weidemann Heidrun, Feger Daniel, Ehlert Jan E, Menger Marcus M, Krempien Robert C
Clinic for Radiotherapy, HELIOS Hospital Berlin-Buch, Schwanebecker Chaussee 50, 13125, Berlin, Germany.
Reaction Biology Europe GmbH, Engesserstr.4, 79108, Freiburg, Germany.
Discov Oncol. 2023 Feb 25;14(1):27. doi: 10.1007/s12672-023-00633-2.
Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor prognosis. GMB are highly recurrent mainly because of radio- and chemoresistance. Radiotherapy with Temozolomide (TMZ) is until today the golden standard adjuvant therapy, however, the optimal treatment of recurrent glioblastoma remains controversial. Ouabain belongs to the Cardiotonic Steroids (CTS) the natural ligands of the Na/K-ATPase (NKA). It is established that the NKA represents a signal transducer with either stimulating or inhibiting cell growth, apoptosis, migration and angiogenesis. Over the last decade evidence grew that CTS have anti-tumor properties especially in GBM.
Proceeding from recent studies we wanted to further demonstrate a divergent effect of Ouabain on a TMZ-resistant (T98G) as compared to a TMZ-sensitive (LN229) GBM cell line.
We analyzed the effect of Ouabain on cell migration and plasma cell membrane potential (PCMP) in the LN229 and T98G GBM cell line as well as underlying mechanisms (Bcl-2 and p-Akt/pan-Akt expression). Moreover, we analyzed the anti-angiogenic effect of Ouabain on human umbilical vein endothelial cells (HUVECs).
T98G cells showed a significant inhibition of cell migration and a significant depolarization of the PCMP at similar Ouabain concentrations (IC50 = 1.67 × 10 M) resp. (IC50 = 2.72 × 10 M) with a strong inverse correlation (R = 0.95). In contrast, LN229 cells did not respond to Ouabain in these assays at all. Similarly, only T98G but not LN229 cells revealed Bcl-2 down-regulation at nanomolar Ouabain concentrations. This unique response to Ouabain is associated with a down-regulation of pan-Akt in T98G cells 24 h after Ouabain (1.0 × 10 M) treatment. For the first time, the anti-angiogenic effect of Ouabain on HUVEC cells (IC50 = 5.49 × 10 M) was demonstrated which correlated strongly with the anti-migratory effect (R = 0.85).
The TMZ-resistant T98G cell line as compared to the TMZ-sensitive LN229 cell line shows a high sensitivity towards Ouabain. We consider it as a promising new compound especially in recurrent GBM to overcome the resistance to TMZ and irradiation.
多形性胶质母细胞瘤（GBM）是最具侵袭性的原发性脑肿瘤，预后较差。GBM具有高度复发性，主要原因是对放疗和化疗耐药。迄今为止，替莫唑胺（TMZ）放疗是金标准辅助治疗，但复发性胶质母细胞瘤的最佳治疗方法仍存在争议。哇巴因属于强心甾类（CTS），是钠钾ATP酶（NKA）的天然配体。已证实NKA是一种信号转导分子，可刺激或抑制细胞生长、凋亡、迁移和血管生成。在过去十年中，越来越多的证据表明CTS具有抗肿瘤特性，尤其是在GBM中。
基于最近的研究，我们想进一步证明与TMZ敏感（LN229）GBM细胞系相比，哇巴因对TMZ耐药（T98G）细胞系具有不同的作用。
我们分析了哇巴因对LN229和T98G GBM细胞系中细胞迁移和质膜电位（PCMP）的影响以及潜在机制（Bcl-2和p-Akt/pan-Akt表达）。此外，我们分析了哇巴因对人脐静脉内皮细胞（HUVECs）的抗血管生成作用。
在相似的哇巴因浓度（IC50 = 1.67×10⁻⁶ M）下，T98G细胞的细胞迁移受到显著抑制，PCMP出现显著去极化（IC50 = 2.72×10⁻⁶ M），两者呈强负相关（R = 0.95）。相比之下，LN229细胞在这些实验中对哇巴因完全没有反应。同样，只有T98G细胞而非LN229细胞在纳摩尔浓度的哇巴因作用下显示Bcl-2下调。对哇巴因的这种独特反应与哇巴因（1.0×10⁻⁶ M）处理24小时后T98G细胞中pan-Akt的下调有关。首次证明了哇巴因对HUVEC细胞具有抗血管生成作用（IC50 = 5.49×10⁻⁶ M），且与抗迁移作用密切相关（R = 0.85）。
与TMZ敏感的LN229细胞系相比，TMZ耐药的T98G细胞系对哇巴因表现出高敏感性。我们认为它是一种有前景的新化合物，特别是在复发性GBM中，可克服对TMZ和放疗的耐药性。
