Department of Anatomy and Cell Biology, Rush University Medical Center, Chicago, Illinois 60612, USA.
J Biol Chem. 2010 Jul 23;285(30):23105-14. doi: 10.1074/jbc.M110.124990. Epub 2010 May 20.
Nutlin-3a is a preclinical drug that stabilizes p53 by blocking the interaction between p53 and MDM2. In our previous study, Nutlin-3a promoted a tetraploid G(1) arrest in two p53 wild-type cell lines (HCT116 and U2OS), and both cell lines underwent endoreduplication after Nutlin-3a removal. Endoreduplication gave rise to stable tetraploid clones resistant to therapy-induced apoptosis. Prior knowledge of whether cells are susceptible to Nutlin-induced endoreduplication and therapy resistance could help direct Nutlin-3a-based therapies. In the present study, Nutlin-3a promoted a tetraploid G(1) arrest in multiple p53 wild-type cell lines. However, some cell lines underwent endoreduplication to relatively high extents after Nutlin-3a removal whereas other cell lines did not. The resistance to endoreduplication observed in some cell lines was associated with a prolonged 4N arrest after Nutlin-3a removal. Knockdown of either p53 or p21 immediately after Nutlin-3a removal could drive endoreduplication in otherwise resistant 4N cells. Finally, 4N-arrested cells retained persistent p21 expression; expressed senescence-associated beta-galactosidase; displayed an enlarged, flattened phenotype; and underwent a proliferation block that lasted at least 2 weeks after Nutlin-3a removal. These findings demonstrate that transient Nutlin-3a treatment can promote an apparently permanent proliferative block in 4N cells of certain cell lines associated with persistent p21 expression and resistance to endoreduplication.
Nutlin-3a 是一种通过阻断 p53 与 MDM2 之间的相互作用来稳定 p53 的临床前药物。在我们之前的研究中，Nutlin-3a 使两个 p53 野生型细胞系（HCT116 和 U2OS）中的细胞发生四倍体 G1 期阻滞，并且在 Nutlin-3a 去除后，这两个细胞系都经历了内复制。内复制导致对治疗诱导的细胞凋亡具有抗性的稳定四倍体克隆。预先了解细胞是否容易受到 Nutlin 诱导的内复制和治疗耐药性的影响，可能有助于指导基于 Nutlin-3a 的治疗。在本研究中，Nutlin-3a 在多个 p53 野生型细胞系中促进了四倍体 G1 期阻滞。然而，在 Nutlin-3a 去除后，一些细胞系经历了相对较高程度的内复制，而其他细胞系则没有。在一些细胞系中观察到的对内复制的抗性与 Nutlin-3a 去除后 4N 阻滞的延长有关。Nutlin-3a 去除后立即敲低 p53 或 p21 可使原本耐受的 4N 细胞发生内复制。最后，4N 期阻滞的细胞保留持续的 p21 表达；表达衰老相关的β-半乳糖苷酶；表现出扩大的、扁平的表型；并且在 Nutlin-3a 去除后至少持续 2 周的增殖阻滞。这些发现表明，短暂的 Nutlin-3a 处理可以在某些细胞系的 4N 细胞中促进明显的永久性增殖阻滞，这与持续的 p21 表达和对内复制的抗性有关。
