Seitz Guido, Warmann Steven W, Vokuhl Christian O, Heitmann Heike, Treuner Claudia, Leuschner Ivo, Fuchs Jörg
Department of Pediatric Surgery, University Children's Hospital, Hoppe-Seyler-Strasse 1, 72076 Tuebingen, Germany.
Pediatr Surg Int. 2007 May;23(5):431-9. doi: 10.1007/s00383-006-1852-z.
The prognosis of rhabdomyosarcoma (RMS) in advanced stages is still sobering. Therapy is limited due to local tumor recurrence, development of metastases and multidrug resistance. The aim of this study was to investigate the development of multidrug resistance in cell lines and in xenografts of alveolar and embryonal RMS treated according to the German Soft Tissue Sarcoma Study (CWS). Alveolar and embryonal RMS cell lines were treated with Vincristine, Topotecan, Carboplatin, Actinomycin D, or Ifosfamide. Expression levels of resistance-associated genes were assessed using Real time-PCR. Nude mice (NMRI nu/nu, n = 10 per group) underwent xenotransplantation of human embryonal or alveolar RMS. Animals were treated with standard chemotherapeutic drugs Vincristine, Topotecan, Carboplatin, Actinomycin D, or Ifosfamide according to treatment schedules of the CWS-study. Tumor sizes were measured and relative tumor volumes were calculated. Animals were sacrificed after 20 days and standard histology, Real-time-PCR for MDR1-, MRP-, LRP- and MDM2-gene as well as immunohistochemistry for MDR1-, LRP-, and MRP-protein were performed. In the cell lines, an up-regulation of MDR-1 gene was found in alveolar rhabdomyosarcoma. In embryonal rhabdomyosarcoma, an up-regulation of LRP and MRP was found. Standard chemotherapy of alveolar rhabdomyosarcoma resulted in a significant reduction of tumor growth (P < 0.05) in all groups. In embryonal rhabdomyosarcoma strongest effects were found after treatment with Ifosfamide, Vincristine and Carboplatin (P < 0.05). RT-PCR revealed a MDR1-dependent mechanism in alveolar rhabdomyosarcoma. In embryonal rhabdomyosarcoma, MDR1 occurred to a lower degree. Immunohistochemistry revealed correlating expression levels of multidrug resistance-associated proteins. The use of established chemotherapy on human RMS in vivo had strong effects on xenografts compared to their controls. In all cases, there was only a reduction of tumor growth, but not a complete eradication of the tumors. Chemotherapy seemed to upregulate the expression of resistance-associated genes in vitro and in vivo. The mechanism of multidrug resistance depends on the tumor subtype. Therefore, further investigations will be required to evaluate multidrug resistance in patients and to investigate new modalities for a reversal of multidrug resistance.
晚期横纹肌肉瘤（RMS）的预后仍然不容乐观。由于局部肿瘤复发、转移的发生以及多药耐药性，治疗受到限制。本研究的目的是调查根据德国软组织肉瘤研究（CWS）进行治疗的肺泡型和胚胎型RMS细胞系及异种移植模型中多药耐药性的发展情况。肺泡型和胚胎型RMS细胞系分别用长春新碱、拓扑替康、卡铂、放线菌素D或异环磷酰胺进行处理。使用实时定量聚合酶链反应（Real time-PCR）评估耐药相关基因的表达水平。将裸鼠（NMRI nu/nu，每组n = 10只）进行人胚胎型或肺泡型RMS的异种移植。根据CWS研究的治疗方案，用标准化疗药物长春新碱、拓扑替康、卡铂、放线菌素D或异环磷酰胺对动物进行治疗。测量肿瘤大小并计算相对肿瘤体积。20天后处死动物，进行标准组织学检查、针对多药耐药蛋白1（MDR1）、多药耐药相关蛋白（MRP）、肺耐药蛋白（LRP）和小鼠双微体2（MDM2）基因的实时定量聚合酶链反应以及针对MDR1、LRP和MRP蛋白的免疫组织化学检查。在细胞系中，肺泡型横纹肌肉瘤中发现多药耐药蛋白1（MDR-1）基因上调。在胚胎型横纹肌肉瘤中，发现LRP和MRP上调。肺泡型横纹肌肉瘤的标准化疗导致所有组的肿瘤生长显著减少（P < 0.05）。在胚胎型横纹肌肉瘤中，用异环磷酰胺、长春新碱和卡铂治疗后效果最为显著（P < 0.05）。逆转录-聚合酶链反应（RT-PCR）显示肺泡型横纹肌肉瘤存在依赖MDR1的机制。在胚胎型横纹肌肉瘤中，MDR1的程度较低。免疫组织化学显示多药耐药相关蛋白的表达水平具有相关性。与对照组相比，对人RMS体内使用既定化疗方案对异种移植瘤有显著效果。在所有情况下，仅肿瘤生长有所减少，但并未完全根除肿瘤。化疗似乎在体外和体内均可上调耐药相关基因的表达。多药耐药机制取决于肿瘤亚型。因此，需要进一步研究以评估患者的多药耐药性，并研究逆转多药耐药性的新方法。
