Differential Cellular Effects of Plk1 Inhibitors Targeting the ATP-binding Domain or Polo-box Domain.

The expression of polo-like kinase 1 (Plk1) correlates with malignancy and is thus recognized as a target for cancer therapy. In addition to the development of ATP-competitive Plk1 inhibitors, the polo-box domain (PBD), a unique functional domain of PLKs, is being targeted to develop Plk1-specific inhibitors. However, the action mechanisms of these two classes of Plk1 inhibitors have not been thoroughly evaluated. Here, we evaluate the differences in cellular effects of ATP-binding domain inhibitors (BI 2536, GSK 461364) and PBD inhibitors (poloxin, thymoquinone) to determine their mechanisms of Plk1 inhibition. Our data show that BI 2536 and GSK461364 increased the population of cells in the G2/M phase compared with controls, while treatment with poloxin and thymoquinone increased cell population in the S phase as well as in G2/M, in a p53-independent manner. The population of cells staining positively for p-Histone H3 and MPM2, mitotic index, was increased by treatment with BI 2536 or GSK461364, but not by treatment with poloxin or thymoquinone. Furthermore, treatment with BI 2536 or GSK461364 resulted in activation of the BubR1 spindle checkpoint kinase, suggesting that treatment with ATP-binding domain inhibitors induces metaphase arrest. However, the administration of poloxin and thymoquinone resulted in an increase in p21(WAF1) and S arrest, indicating that PBD inhibitors also affected interphase before mitotic entry. Taken together, these data suggest that the PDB of Plk1 plays a role in S phase progression through interaction with other proteins, while its ATP-binding domain is important for regulating mitotic progression mediated by its catalytic activity involving consumption of ATP.

Shin SB ，Woo SU ，Yim H 《-》

Small molecule inhibition of polo-like kinase 1 by volasertib (BI 6727) causes significant melanoma growth delay and regression in vivo.

The objective of this study was to determine the therapeutic potential of polo-like kinase 1 (Plk1) inhibition in melanoma, in vivo. Employing Vectra technology, we assessed the Plk1 expression profile in benign nevi, malignant (stages I-IV) and metastatic melanomas. We found a significant elevation of Plk1 immunostaining in melanoma tissues. Further, a second generation small molecule Plk1 inhibitor, BI 6727, resulted in reductions in growth, viability and clonogenic survival, as well as an increase in apoptosis of A375 and Hs 294T melanoma cells. BI 6727 treatment also resulted in a G2/M-as well as S-phase cell cycle arrest in melanoma cells. Importantly, BI 6727 (intravenous injection; 10 and 25 mg/kg body weight) treatment resulted in significant tumor growth delay and regression in vivo in A375-and Hs 294T-implanted xenografts in athymic nude mice. These anti-melanoma effects were accompanied with a decreased cellular proliferation (Ki-67 staining) and induction of apoptosis (caspase 3 activation). In addition, BI 6727 treatment caused a marked induction of p53 and p21 in vitro as well as in vivo. Overall, we suggest that Plk1 inhibition may be a useful approach as a monotherapy as well as in combination with other existing therapeutics, for melanoma management.

Cholewa BD ，Ndiaye MA ，Huang W ，Liu X ，Ahmad N ... -  《-》

Loss of p21Cip1/CDKN1A renders cancer cells susceptible to Polo-like kinase 1 inhibition.

Kreis NN ，Louwen F ，Zimmer B ，Yuan J ... -  《Oncotarget》

Polo-box domain inhibitor poloxin activates the spindle assembly checkpoint and inhibits tumor growth in vivo.

Polo-like kinase 1 (Plk1) is widely established as one of the most promising targets in oncology. Although the protein kinase domain of Plk1 is highly conserved, the polo-box domain (PBD) of Plk1 provides a much more compelling site to specifically inhibit the localization and target binding of Plk1. We recently identified, via fluorescence polarization assay, the natural product derivative, Poloxin, as the first small-molecule inhibitor specifically targeting the function of the Plk1 PBD. In this study, we characterized its mitotic phenotype and its function in vitro and in vivo. Poloxin induces centrosome fragmentation and abnormal spindle and chromosome misalignment, which activate the spindle assembly checkpoint and prolong mitosis. Notably, centrosomal fragmentation induced by Poloxin is partially attributable to dysfunctional Kizuna, a key substrate of Plk1 at centrosomes. Moreover, Poloxin strongly inhibits proliferation of a panel of cancer cells by inducing mitotic arrest, followed by a surge of apoptosis. More important, we report, for the first time to our knowledge, that the PBD inhibitor, Poloxin, significantly suppresses tumor growth of cancer cell lines in xenograft mouse models by lowering the proliferation rate and triggering apoptosis in treated tumor tissues. The data highlight that targeting the PBD by Poloxin is a powerful approach for selectively inhibiting Plk1 function in vitro and in vivo.

Yuan J ，Sanhaji M ，Krämer A ，Reindl W ，Hofmann M ，Kreis NN ，Zimmer B ，Berg T ，Strebhardt K ... -  《-》

Optimized Plk1 PBD Inhibitors Based on Poloxin Induce Mitotic Arrest and Apoptosis in Tumor Cells.

Scharow A ，Raab M ，Saxena K ，Sreeramulu S ，Kudlinzki D ，Gande S ，Dötsch C ，Kurunci-Csacsko E ，Klaeger S ，Kuster B ，Schwalbe H ，Strebhardt K ，Berg T ... -  《-》