Olanich Mary E, Sun Wenyue, Hewitt Stephen M, Abdullaev Zied, Pack Svetlana D, Barr Frederic G
Cancer Molecular Pathology Section, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland.
Tissue Array Research Program and Applied Molecular Pathology Laboratory, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland.
Clin Cancer Res. 2015 Nov 1;21(21):4947-59. doi: 10.1158/1078-0432.CCR-14-2955. Epub 2015 Mar 25.
Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma and includes a PAX3- or PAX7-FOXO1 fusion-positive subtype. Amplification of chromosomal region 12q13-q14, which contains the CDK4 proto-oncogene, was identified in an aggressive subset of fusion-positive RMS. CDK4/6 inhibitors have antiproliferative activity in CDK4-amplified liposarcoma and neuroblastoma, suggesting CDK4/6 inhibition as a potential therapeutic strategy in fusion-positive RMS.
We examined the biologic consequences of CDK4 knockdown, CDK4 overexpression, and pharmacologic CDK4/6 inhibition by LEE011 in fusion-positive RMS cell lines and xenografts.
Knockdown of CDK4 abrogated proliferation and transformation of 12q13-14-amplified and nonamplified fusion-positive RMS cells via G1-phase cell-cycle arrest. This arrest was mediated by reduced RB phosphorylation and E2F-responsive gene expression. Significant differences in E2F target expression, cell-cycle distribution, proliferation, or transformation were not observed in RMS cells overexpressing CDK4. Treatment with LEE011 phenocopied CDK4 knockdown, decreasing viability, RB phosphorylation, and E2F-responsive gene expression and inducing G1-phase cell-cycle arrest. Although all fusion-positive cell lines showed sensitivity to CDK4/6 inhibition, there was diminished sensitivity associated with CDK4 amplification and overexpression. This variable responsiveness to LEE011 was recapitulated in xenograft models of CDK4-amplified and nonamplified fusion-positive RMS.
Our data demonstrate that CDK4 is necessary but overexpression is not sufficient for RB-E2F-mediated G1-phase cell-cycle progression, proliferation, and transformation in fusion-positive RMS. Our studies indicate that LEE011 is active in the setting of fusion-positive RMS and suggest that low CDK4-expressing fusion-positive tumors may be particularly susceptible to CDK4/6 inhibition.
横纹肌肉瘤（RMS）是最常见的儿童软组织肉瘤，包括PAX3或PAX7 - FOXO1融合阳性亚型。在侵袭性融合阳性RMS亚组中发现了包含细胞周期蛋白依赖性激酶4（CDK4）原癌基因的染色体区域12q13 - q14的扩增。CDK4/6抑制剂在CDK4扩增的脂肪肉瘤和神经母细胞瘤中具有抗增殖活性，提示CDK4/6抑制作为融合阳性RMS的一种潜在治疗策略。
我们研究了在融合阳性RMS细胞系和异种移植模型中，CDK4敲低、CDK4过表达以及LEE011对CDK4/6的药理抑制作用所产生的生物学后果。
敲低CDK4通过G1期细胞周期阻滞消除了12q13 - 14扩增和未扩增的融合阳性RMS细胞的增殖和转化。这种阻滞是由RB磷酸化减少和E2F反应性基因表达介导的。在过表达CDK4的RMS细胞中未观察到E2F靶标表达、细胞周期分布、增殖或转化的显著差异。用LEE011处理模拟了CDK4敲低的效果，降低了细胞活力、RB磷酸化和E2F反应性基因表达，并诱导G1期细胞周期阻滞。虽然所有融合阳性细胞系对CDK4/6抑制均表现出敏感性，但与CDK4扩增和过表达相关的敏感性降低。在CDK4扩增和未扩增的融合阳性RMS异种移植模型中再现了对LEE011的这种可变反应性。
我们的数据表明，在融合阳性RMS中，CDK4对于RB - E2F介导的G1期细胞周期进程、增殖和转化是必需的，但过表达并不充分。我们的研究表明LEE011在融合阳性RMS中具有活性，并提示低CDK4表达的融合阳性肿瘤可能对CDK4/6抑制特别敏感。
