The Plk1 Inhibitor BI 2536 Temporarily Arrests Primary Cardiac Fibroblasts in Mitosis and Generates Aneuploidy In Vitro

Abstract

BI 2536 is a new anti-mitotic drug that targets polo-like kinase 1 (Plk1) and is currently under clinical development for cancer therapy. The effect of this drug on cancer cells has been extensively investigated, but information about the effects on primary dividing cells and differentiated non-dividing cells is scarce. We have investigated the effects of this drug on primary neonatal rat cardiac fibroblasts and on differentiated cardiomyocytes and explored the possibility to use this drug to enrich differentiated cell populations in vitro. BI 2536 had a profound effect on cardiac fibroblast proliferation in vitro and arrested these cells in mitosis with an IC50 of about 43 nM. Similar results were observed with primary human cells (HUVEC, IC50 = 30 nM), whereas the cancer cell line HeLa was more sensitive (IC50 of 9 nM). Further analysis revealed that prolonged mitotic arrest resulted in cell death for about 40% of cardiac fibroblasts. The remaining cells showed an interphase morphology with mostly multi- and micro-nucleated nuclei. This indicates that a significant number of primary fibroblasts are able to escape BI 2536 induced mitotic arrest and apparently become aneuploid. No effects were observed on cardiomyocytes and hypertrophic response (growth) upon endothelin-1 and phenylephrine stimulation was normal in the presence of BI 2536. This indicates that BI 2536 has no adverse effects on terminally differentiated cells and still allows proliferation independent growth induction in these cells. In conclusion, cardiomyocytes could be enriched using BI 2536, but the formation of aneuploidy in proliferating cells most likely limits this in vitro application and does not allow its use in putative cell based therapies.

Polo-like kinase 1 inhibitors, mitotic stress and the tumor suppressor p53

“…Taken together, based on the data from our studies and others, 10,15,16,19,38 we suggest that loss of functional p53 does not directly facilitates the cytotoxicity of using a FACSCalibur (BD Biosciences, Heidelberg), as described. 57 The staining of phospho-histone H3 and active caspase-3 was also evaluated by FACS.…”

“…29,30 However, we and others observed that Plk1 inhibitors affect both tumor cells with functional or deficient p53 as well as primary/normal non-transformed proliferating cells. 16,19,32,38 In the previous work, based on different cancer cell lines, we have demonstrated that cancer cells with wild type p53 actually responded more sensitively to Plk1 inhibitors, as evidenced by a high degree of apoptosis induction. 15 In the present work, we have focused on the question whether mitotic stress influences the effect of Plk1 inhibitors in cancer cells in context of the p53 status.…”

“…Rather, by activating cell death signaling pathways, functional p53 strengths the efficacy of Plk1 inhibitors and prevents possibly genome instability caused by Plk1 inhibitors. 38 Further studies are required to investigate whether the long-term outcomes of losing p53, such as low differential grade of tumor cells, defective DNA damage checkpoint or unbalanced metabolism in tumor cells, which makes possible the survival of cancer cells sensitize Plk1 inhibitor GSK461364. 56 BI 2536 in combination with pemetrexed shows an encouraging antitumor activity.…”

Inhibition of Polo-Like Kinase 1 Induces Cell Cycle Arrest and Sensitizes Glioblastoma Cells to Ionizing Radiation

“…G2/ M arrest after PLK1 inhibition has already been shown in different cell types, including fibroblastic, primary, and cancer cells. [23][24][25] It is possible that after repeated efforts to divide, cells proceed toward death through apoptosis, which is the hallmark of PLK1 inhibition. Consistent with our findings, increased cell death rates have also been reported after PLK1 inhibition in pancreatic cancer cells, 26 hepatocellular carcinomas cells, 27 and squamous cell carcinoma cells, 28 among others.…”

Cooperative Phosphorylation of FADD by Aur-A and Plk1 in Response to Taxol Triggers Both Apoptotic and Necrotic Cell Death

“…Moreover, enhanced kinase activity associated with Plk1 and Aur-A overexpression confers a proliferative advantage in some tumors (11). Therefore, inhibitors of theses kinases are currently being evaluated for therapeutic use (11)(12)(13)(14)(15)(16). However, the feasibility of using these kinases as anticancer drug targets is a matter of controversy.…”