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Tamoxifen resistance is a multifaceted phenomenon, characterized by the constitutive activation of multiple signaling cascades that provide an additional survival advantage to cells. Ground studies related to reverse the tamoxifen resistance by employing chemotherapeutic drugs that specifically inhibit proteins, those of aberrantly expressed, are required. Seminal studies showed that p38 signaling and VEGF play crucial role in acquiring resistance to tamoxifen. In this view, we had chosen paclitaxel, a mitotic inhibitor with anti-proliferative effects against a wide array of cancers in this study. Further to mitigate the undesirable complications of paclitaxel (PAC), we employed this drug in combination along with BI2536 (BI), a PLK inhibitor for this study to sensitize the tamoxifen resistant cells to apoptosis. MCF 7/TAM and T-47D/TAM cells were treated with PAC, BI and in combination (BI-PAC) evaluated for its anticancer activity through apoptotic and western blot analysis. Modulatory effects of BI-PAC on p38 inactivation were affirmed through immunofluorescence and drug potential studies. Results reveal that cells were subjected to apoptosis on drug(s) treatment which was confirmed through cytotoxicity, annexin studies. Further, the anti-proliferative effects of the drug(s) were affirmed through nuclear morphological and TUNEL assays. Immunoblot results revealed the upregulation of proapoptotic Bax, cleaved caspase 9 along with Bcl-2, MDM2, Cox-2, and P-Gly down regulation after 24h drug treatments. Moreover, phospho studies further construed the rationale behind the apoptosis and deduced the inactivation of p38 and NF-κB role in inducing apoptosis in drug treated cells. The efficacy of drug combinations in inactivating p38 was evaluated through drug potential studies. Further, BI-PAC treatments showed inhibition of p38 mediated senescence in tamoxifen resistant cells. Overall, our observations provide a new therapeutic combination that sensitizes tamoxifen resistant cells to apoptosis by specifically targeting p38 signaling and its downstream molecules and subsequently reduces extracellular VEGF levels., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Polo-like kinase 1 (PLK1) is an important regulator of the cell cycle and is overexpressed in various solid and hematological malignancies. Small molecule inhibitors targeting PLK1, such as BI2536 or BI6727 (Volasertib) are a promising therapeutic approach in such malignancies. Here, we show a loss of specifically localized PLK1 in AML blasts in vivo, accompanied by mitotic arrest with transition into apoptosis, in bone marrow biopsies of AML patients after treatment with BI2536. We verify these results in live cell imaging experiments with the AML cell line HL-60, and demonstrate that non-neoplastic, immortalized lymphoblastoid cells are also sensitive to PLK1 inhibition. It is demonstrated that normal granulopoietic precursors have similar PLK1 expression levels as leukemic blasts. These results are in line with the adverse effects of PLK1 inhibition and underline the great potential of PLK1 inhibitors in the treatment of AML., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Cholangiocarcinoma (CCA) is a lethal cancer originating from the epithelial cells within the bile duct and ranks as the second most prevalent form of liver cancer in Thailand. Polo-like kinase 1 (PLK1), a protein serine/threonine kinase, regulates a number of steps in cell mitosis and is upregulated in several types of cancer, including CCA. Our previous study identified PLK1 as a biomarker of the C1 subtype, correlating with poor prognosis in intrahepatic CCA. The present study aimed to examine the effect of PLK1 inhibition on CCA cells. Different CCA cell lines developed from Thai patients, HuCCA1, KKU055, KKU100 and KKU213A, were treated with two PLK1 inhibitors, BI2536 and BI6727, and were transfected with small interfering RNA, followed by analysis of cell proliferation, cell cycle distribution and cell apoptosis. It was discovered that BI2536 and BI6727 inhibited cell proliferation and caused G2/M-phase arrest in CCA cells. Furthermore, the number of total apoptotic cells was increased in PLK1 inhibitor-treated CCA cells. The expression levels of mitotic proteins, aurora kinase A, phosphorylated PLK1 (T210) and cyclin B1, were augmented in PLK1-inhibited CCA cells. Additionally, inhibition of PLK1 led to increased DNA damage, as determined by the upregulated levels of γH2AX and increased cleavage of poly (ADP-ribose) polymerase, an apoptotic marker. These results suggested that inhibiting PLK1 prolonged mitotic arrest and subsequently triggered cell apoptosis. Validation of the antiproliferative effects of PLK1 inhibition was accomplished through silencing of the PLK1 gene. In conclusion, targeting PLK1 provided promising results for further study as a potential candidate for targeted therapy in CCA. [ABSTRACT FROM AUTHOR]

The efficacy of antimitotics is limited by slippage, whereby treated cells arrested in mitosis exit mitosis without cell division and, eventually, escape apoptosis, constituting a serious resistance mechanism to antimitotics. Strategies to overcome slippage should potentiate the cancer cell killing activity of these antimitotics. Such strategies should accelerate cell death in mitosis before slippage. Here, we undertook a mechanistic analysis to test whether the apoptosis activator Navitoclax potentiates apoptosis triggered by the antimitotic BI2536, a potent inhibitor of Polo-like kinase 1 (PLK1) with the goal of overcoming slippage. We found that cancer cells in 2D cultures treated with BI2536 alone accumulate in mitosis, but a significant fraction of arrested cells undergo slippage and survive. Remarkably, combining BI2536 with Navitoclax dramatically reduces slippage, shifting the cell fate to accelerated death in mitosis. The results are confirmed in 3D spheroids, a preclinical system that mimics in vivo tumor drug responses. Importantly, in 3D spheroids, the effect of the BI2536/Navitoclax combination requires a lower therapeutic dosage of each drug, underlying its potential to improve the therapeutic index. Our results highlight the relevance of apoptosis potentiators to circumvent slippage associated with antimitotics. The combination of BI2536 with Navitoclax shows in vitro synergy/additive effect, which warrants further clinical research. [ABSTRACT FROM AUTHOR]

Muscle development and regeneration require delicate cell cycle regulation of embryonic myoblasts and adult muscle satellite cells (MuSCs). Through analysis of the Polo-like kinase (Plk) family cell-cycle regulators in mice, we show that Plk1’s expression closely mirrors myoblast dynamics during embryonic and postnatal myogenesis. Cell-specific deletion of Plk1 in embryonic myoblasts leads to depletion of myoblasts, developmental failure and prenatal lethality. Postnatal deletion of Plk1 in MuSCs does not perturb their quiescence but depletes activated MuSCs as they enter the cell cycle, leading to regenerative failure. The Plk1-null MuSCs are arrested at the M-phase, accumulate DNA damage, and apoptose. Mechanistically, Plk1 deletion upregulates p53, and inhibition of p53 promotes survival of the Plk1-null myoblasts. Pharmacological inhibition of Plk1 similarly inhibits proliferation but promotes differentiation of myoblasts in vitro, and blocks muscle regeneration in vivo. These results reveal for the first time an indispensable role of Plk1 in developmental and regenerative myogenesis.

Background Merkel cell carcinoma (MCC) is a rare and aggressive malignancy of the skin. Treatment options for MCC include surgery, radiotherapy, and chemotherapy. The purpose of this study was to assess the expression of Polo-like kinase 1 (PLK1) in MCC and the role of the inhibitor, BI2536, as a potential therapeutic option in MCC. Methods PLK1 expression was assessed in tissue samples from 28 patients with MCC and 5 healthy skin samples via immunohistochemistry and furthermore in the 2 MCC cell lines, MCC13 and MCC26, via immunoblotting. The impact of increasing doses of BI2536 alone and in combination with cisplatin or irradiation on cell viability was measured using the CCK-8 assay. Colony forming assays were performed to evaluate long-term effects of combination treatments. Additionally, the induction of apoptotic cell death was measured via flow cytometry. Results PLK1 is moderately to strongly expressed in 75% of the patients with MCC. The PLK1 inhibitor, BI2536, demonstrated marked inhibition of cell proliferation with IC50 in the low nM range (from 10.07-12.39 nM). Furthermore, BI2536 induces apoptosis in MCC cell lines and acts synergistically in combination with irradiation and cisplatin. Conclusion Because of the marked upregulation of PLK1 in MCC tumor samples and potent inhibition of cell proliferation using a specific clinically available inhibitor, targeting of PLK1 qualifies as a potential novel therapeutic strategy in MCC. © 2015 Wiley Periodicals, Inc. Head Neck 38: E1918-E1925, 2016 [ABSTRACT FROM AUTHOR]