Das Samayita
Innere Klinik (Tumorforschung), West German Cancer Center (WTZ), University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
Adv Radiat Oncol. 2019 Nov 27;5(2):250-259. doi: 10.1016/j.adro.2019.11.004. eCollection 2020 Mar-Apr.
Inhibition of p53 by amplification of MDM2 is one of the key contributors in the oncogenesis of a subset of soft tissue sarcoma (STS) known as well-/dedifferentiated liposarcoma. A small molecule MDM2 antagonist, nutlin-3, induces the p53 pathway by disrupting the interaction between MDM2 and p53. Radiation therapy is an integral component for treating liposarcoma and induces p53. Based on wild-type status in liposarcoma, it was investigated whether MDM2 inhibition with irradiation led to enhanced reactivation of p53 and subsequent p53-mediated effects in liposarcoma.
Clonogenic assays, immunoblotting, flow cytometry/fluorescence-activated cell sorting, and senescence assays were employed in liposarcoma cell lines after co-treatment with nutlin-3 and radiation.
Upon treatment with nutlin-3, 2 of the well-/dedifferentiated liposarcoma (MDM2/TP53) cell lines displayed radiosensitivity with sensitization enhancement ratio values of >1. In contrast, the cell line with mutant showed sensitization enhancement ratio values of ∼1. Immunoblotting revealed induced reactivation of the p53-MDM2-p21 signaling axis in response to combination therapy in all cell lines with wild-type Removal of MDM2 inhibitor (with or without radiation therapy) led to the emergence of ploidy-based heterogeneous subpopulations (4N and >4N) in wild-type cells and not in mutant cells. Immunoblotting of cell cycle markers (G1, G2/M) revealed the generation of 4N G1 cells. Sorting and long-term fate analysis of different populations (2N, 4N, and >4N) by colony assay displayed attenuated colony-forming potential and augmented senescence of the 4N and >4N cells contributing to the radiosensitization effect.
Nutlin-3 increases the vulnerability of liposarcoma cell lines to radiation by augmented activation of p53. The cells underwent senescence. Presence and activation of p53 are required for exertion of the radiosensitizing effect by nutlin-3, but this is not the sole determinant of the effect. This study opens avenues for the clinical translation in a stratified group of patients with liposarcoma.
MDM2扩增对p53的抑制作用是一种称为高分化/去分化脂肪肉瘤的软组织肉瘤（STS）亚群肿瘤发生的关键因素之一。小分子MDM2拮抗剂nutlin-3通过破坏MDM2与p53之间的相互作用来诱导p53通路。放射治疗是治疗脂肪肉瘤不可或缺的组成部分，且能诱导p53。基于脂肪肉瘤中的野生型状态，研究了用辐射抑制MDM2是否会导致p53的再激活增强以及随后在脂肪肉瘤中p53介导的效应。
在脂肪肉瘤细胞系中，将nutlin-3与辐射联合处理后，采用克隆形成试验、免疫印迹、流式细胞术/荧光激活细胞分选和衰老试验。
用nutlin-3处理后，2株高分化/去分化脂肪肉瘤（MDM2/TP53）细胞系表现出放射敏感性，增敏增强比值>1。相比之下，具有突变体的细胞系增敏增强比值约为1。免疫印迹显示，在所有具有野生型的细胞系中，联合治疗可诱导p53-MDM2-p21信号轴的再激活。去除MDM2抑制剂（无论有无放射治疗）会导致野生型细胞中出现基于倍性的异质性亚群（4N和>4N），而突变体细胞中则不会出现。细胞周期标志物（G1、G2/M）的免疫印迹显示产生了4N G1细胞。通过集落试验对不同群体（2N、4N和>4N）进行分选和长期命运分析，结果显示4N和>4N细胞的集落形成潜力减弱，衰老增加，从而产生放射增敏效应。
Nutlin-3通过增强p53的激活增加了脂肪肉瘤细胞系对辐射的敏感性。细胞发生了衰老。p53的存在和激活是nutlin-3发挥放射增敏作用所必需的，但这不是该效应的唯一决定因素。本研究为分层的脂肪肉瘤患者群体的临床转化开辟了道路。
