Targeting Of PLK1 Research Articles

Therapeutic targeting of PLK1 in TERT promoter-mutant hepatocellular carcinoma.

Hotspot mutations in the promoter of telomerase reverse transcriptase (TERT) gene are the most common genetic variants in hepatocellular carcinoma (HCC) and associated with poor prognosis of the disease. However, no drug was currently approved for treating TERT promoter mutation positive HCC patients. Here, we aim to explore the potential therapeutic strategy for targeting TERT promoter mutation in HCC. The Liver Cancer Model Repository database was used for screening potential drugs to selectively suppress the growth of TERT promoter mutant HCC cells. RNA-seq, CRISPR-Cas9 technology and siRNA transfection were performed for mechanistic studies. Cell counting kit-8 (CCK8) assay and the xenograft tumour models were used for cell growth detection in vitro and in vivo, respectively. Cell apoptosis and cell cycle arrest were analysed by Annexin V-FITC staining and/or propidium iodide staining. PLK1 inhibitors were remarkably more sensitive to HCC cells harbouring TERT promoter mutation than wild-type cells in vitro and in vivo, which were diminished after TERT promoter mutation was edited to the wild-type nucleotide. Comparing the HCC cells with wild-type promoter of TERT, PLK1 inhibitors specifically downregulated Smad3 to regulate TERT for inducing apoptosis and G2/M arrest in TERT mutant HCC cells. Moreover, knockout of Smad3 counteracted the effects of PLK1 inhibitors in TERT mutant HCC cells. Finally, a cooperative effect of PLK1 and Smad3 inhibition was observed in TERT mutant cells. PLK1 inhibition selectively suppressed the growth of TERT mutant HCC cells through Smad3, thus contributed to discover a novel therapeutic strategy to treat HCC patients harbouring TERT promoter mutations. TERT promoter mutation confers sensitivity to PLK1 inhibitors in HCC. The selective growth inhibition of TERT mutant HCC cells induced by PLK1 inhibitor was mediated by Smad3. Combined inhibition of PLK1 and Smad3 showed a cooperative anti-tumor effect in TERT mutant HCC cells.

Green Stereoselective Synthesis and In Silico Anticancer Evaluation of Tetrahydro-β-carboline-Derived Spiro Heterocycles

AbstractA green stereoselective synthesis of spiroheterocycles incorporating a spirooxindole and a 1,2,3,4-tetrahydro-β-carboline (THβC) are demonstrated here by the one-pot, three-component reaction of THβC, isatins, and chalcones. Operational simplicity and chromatography-free isolation are the highlights of the reaction which resulted in densely substituted spiroheterocycles with four-contiguous stereocenters in excellent yields. The activity of the compounds as anticancer agents was studied in silico against MDM2 and PLK1 target proteins and they show excellent binding interactions compared to reference drugs.

Development of Cell Permeable NanoBRET Probes for the Measurement of PLK1 Target Engagement in Live Cells.

PLK1 is a protein kinase that regulates mitosis and is both an important oncology drug target and a potential antitarget of drugs for the DNA damage response pathway or anti-infective host kinases. To expand the range of live cell NanoBRET target engagement assays to include PLK1, we developed an energy transfer probe based on the anilino-tetrahydropteridine chemotype found in several selective PLK inhibitors. Probe 11 was used to configure NanoBRET target engagement assays for PLK1, PLK2, and PLK3 and measure the potency of several known PLK inhibitors. In-cell target engagement for PLK1 was in good agreement with the reported cellular potency for the inhibition of cell proliferation. Probe 11 enabled the investigation of the promiscuity of adavosertib, which had been described as a dual PLK1/WEE1 inhibitor in biochemical assays. Live cell target engagement analysis of adavosertib via NanoBRET demonstrated PLK activity at micromolar concentrations but only selective engagement of WEE1 at clinically relevant doses.

PLK1 regulates CtIP and DNA2 interplay in long-range DNA end resection.

DNA double-strand break (DSB) repair is initiated by DNA end resection. CtIP acts in short-range resection to stimulate MRE11-RAD50-NBS1 (MRN) to endonucleolytically cleave 5'-terminated DNA to bypass protein blocks. CtIP also promotes the DNA2 helicase-nuclease to accelerate long-range resection downstream from MRN. Here, using AlphaFold2, we identified CtIP-F728E-Y736E as a separation-of-function mutant that is still proficient in conjunction with MRN but is not able to stimulate ssDNA degradation by DNA2. Accordingly, CtIP-F728E-Y736E impairs physical interaction with DNA2. Cellular assays revealed that CtIP-F728E-Y736E cells exhibit reduced DSB-dependent chromatin-bound RPA, impaired long-range resection, and increased sensitivity to DSB-inducing drugs. Previously, CtIP was shown to be targeted by PLK1 to inhibit long-range resection, yet the underlying mechanism was unclear. We show that the DNA2-interacting region in CtIP includes the PLK1 target site at S723. The integrity of S723 in CtIP is necessary for the stimulation of DNA2, and phosphorylation of CtIP by PLK1 in vitro is consequently inhibitory, explaining why PLK1 restricts long-range resection. Our data support a model in which CDK-dependent phosphorylation of CtIP activates resection by MRN in S phase, and PLK1-mediated phosphorylation of CtIP disrupts CtIP stimulation of DNA2 to attenuate long-range resection later at G2/M.

Development of Cell Permeable NanoBRET Probes for the Measurement of PLK1 Target Engagement in Live Cells

Development of Cell Permeable NanoBRET Probes for the Measurement of PLK1 Target Engagement in Live Cells

Sequential Targeting of PLK1 and PARP1 Reverses the Resistance to PARP Inhibitors and Enhances Platin-Based Chemotherapy in BRCA-Deficient High-Grade Serous Ovarian Cancer with KRAS Amplification.

Ovarian cancer (OC) accounts for approximately 4% of cancer deaths in women worldwide and is the deadliest gynecologic malignancy. High-grade serous ovarian cancer (HGSOC) is the most predominant ovarian cancer, in which BRCA1/2 gene mutation ranges from 3 to 27%. PARP inhibitors (PARPi) have shown promising results as a synthetically lethal therapeutic approach for BRCA mutant and recurrent OC in clinical use. However, emerging data indicate that BRCA-deficient cancers may be resistant to PARPi, and the mechanisms of this resistance remain elusive. We found that amplification of KRAS likely underlies PARPi resistance in BRCA2-deficient HGSOC. Our data suggest that PLK1 inhibition restores sensitivity to PARPi in HGSOC with KRAS amplification. The sequential combination of PLK1 inhibitor (PLK1i) and PARPi drastically reduces HGSOC cell survival and increases apoptosis. Furthermore, we were able to show that a sequential combination of PLK1i and PARPi enhanced the cellular apoptotic response to carboplatin-based chemotherapy in KRAS-amplified resistant HGSOC cells and 3D spheroids derived from recurrent ovarian cancer patients. Our results shed new light on the critical role of PLK1 in reversing PARPi resistance in KRAS-amplified HGSOC, and offer a new therapeutic strategy for this class of ovarian cancer patients where only limited options currently exist.

Identification of polo-like kinase 1 as a therapeutic target in murine lupus.

IntroductionThe signalling cascades that contribute to lupus pathogenesis are incompletely understood. We address this by using an unbiased activity‐based kinome screen of murine lupus.MethodsAn unbiased activity‐based kinome screen (ABKS) of 196 kinases was applied to two genetically different murine lupus strains. Systemic and renal lupus were evaluated following in vivo PLK1blockade. The upstream regulators and downstream targets of PLK1 were also interrogated.ResultsMultiple signalling cascades were noted to be more active in murine lupus spleens, including PLK1. In vivo administration of a PLK1‐specific inhibitor ameliorated splenomegaly, anti‐dsDNA antibody production, proteinuria, BUN and renal pathology in MRL.lpr mice (P < 0.05). Serum IL‐6, IL‐17 and kidney injury molecule 1 (KIM‐1) were significantly decreased after PLK1 inhibition. PLK1 inhibition reduced germinal centre and marginal zone B cells in the spleen, but changes in T cells were not significant. In vitro, splenocytes were treated with anti‐mouse CD40 Ab or F(ab’)2 fragment anti‐mouse IgM. After 24‐h stimulation, IL‐6 secretion was significantly reduced upon PLK1 blockade, whereas IL‐10 production was significantly increased. The phosphorylation of mTOR was assessed in splenocyte subsets, which revealed a significant change in myeloid cells. PLK1 blockade reduced phosphorylation associated with mTOR signalling, while Aurora‐A emerged as a potential upstream regulator of PLK1.ConclusionThe Aurora‐A → PLK1 → mTOR signalling axis may be central in lupus pathogenesis, and emerges as a potential therapeutic target.

PLK1 and NOTCH Positively Correlate in Melanoma and Their Combined Inhibition Results in Synergistic Modulations of Key Melanoma Pathways.

Melanoma is one of the most serious forms of skin cancer, and its increasing incidence coupled with nonlasting therapeutic options for metastatic disease highlights the need for additional novel approaches for its management. In this study, we determined the potential interactions between polo-like kinase 1 (PLK1, a serine/threonine kinase involved in mitotic regulation) and NOTCH1 (a type I transmembrane protein deciding cell fate during development) in melanoma. Employing an in-house human melanoma tissue microarray (TMA) containing multiple cases of melanomas and benign nevi, coupled with high-throughput, multispectral quantitative fluorescence imaging analysis, we found a positive correlation between PLK1 and NOTCH1 in melanoma. Furthermore, The Cancer Genome Atlas database analysis of patients with melanoma showed an association of higher mRNA levels of PLK1 and NOTCH1 with poor overall, as well as disease-free, survival. Next, utilizing small-molecule inhibitors of PLK1 and NOTCH (BI 6727 and MK-0752, respectively), we found a synergistic antiproliferative response of combined treatment in multiple human melanoma cells. To determine the molecular targets of the overall and synergistic responses of combined PLK1 and NOTCH inhibition, we conducted RNA-sequencing analysis employing a unique regression model with interaction terms. We identified the modulations of several key genes relevant to melanoma progression/metastasis, including MAPK, PI3K, and RAS, as well as some new genes such as Apobec3G, BTK, and FCER1G, which have not been well studied in melanoma. In conclusion, our study demonstrated a synergistic antiproliferative response of concomitant targeting of PLK1 and NOTCH in melanoma, unraveling a potential novel therapeutic approach for detailed preclinical/clinical evaluation.

PLK-1 promotes the merger of the parental genome into a single nucleus by triggering lamina disassembly.

Life of sexually reproducing organisms starts with the fusion of the haploid egg and sperm gametes to form the genome of a new diploid organism. Using the newly fertilized Caenorhabditis elegans zygote, we show that the mitotic Polo-like kinase PLK-1 phosphorylates the lamin LMN-1 to promote timely lamina disassembly and subsequent merging of the parental genomes into a single nucleus after mitosis. Expression of non-phosphorylatable versions of LMN-1, which affect lamina depolymerization during mitosis, is sufficient to prevent the mixing of the parental chromosomes into a single nucleus in daughter cells. Finally, we recapitulate lamina depolymerization by PLK-1 in vitro demonstrating that LMN-1 is a direct PLK-1 target. Our findings indicate that the timely removal of lamin is essential for the merging of parental chromosomes at the beginning of life in C. elegans and possibly also in humans, where a defect in this process might be fatal for embryo development.

Peptidomimetic Polo-Box-Targeted Inhibitors that Engage PLK1 in Tumor Cells and Are Selective against the PLK3 Tumor Suppressor.

The polo-box domain (PBD) of PLK1 determines mitotic substrate recognition and subcellular localization. Compounds that target PLK1 selectively are required due to the tumor-suppressor roles of PLK3. A structure-activity analysis of the PBD phosphopeptide binding motif has identified potent peptides that delineate the determinants required for mimicry by nonpeptidic inhibitors and provide insights into the structural basis for the selectivity of inhibitors for the PLK1 PBD. Fragment-ligated inhibitory peptides (FLIPs) obtained through REPLACE have been optimized to enhance in vitro binding and a systematic analysis of selectivity for PLK1 vs PLK3 has been carried out for peptides and peptidomimetics. Furthermore, these more drug-like non-ATP-competitive inhibitors had on-target engagement in a cellular context, as evidenced by stabilization of PLK1 in a thermal-shift assay and by inhibition of the phosphorylation of TCTP, a target of PLK1. Investigation in cells expressing a mutant PLK1 showed that these cells are sensitive to PBD inhibitors but dramatically resistant to clinically investigated ATP-competitive compounds. These results further validate targeting the PBD binding site in the move towards PLK1 inhibitors that are active against tumors resistant to ATP inhibitors.

Subcellular Drug Targeting Illuminates Local Action of Polo‐like kinase 1 and Aurora A During Mitosis

Fundamental cellular processes such as cell division, migration, metabolism, and growth rely on the precise and timely regulation of signaling proteins. Yet how individual signal transduction events at specific microenvironments drive complex cellular processes remains elusive. During mitosis, two protein kinases, polo‐like kinase 1 (Plk1) and Aurora A (AurA) share several overlapping functions. Current approaches in which small‐molecule kinase inhibitor drugs are used to ascribe function to Plk1 and AurA provide important insight into the global roles of these proteins inside the cell. Unfortunately, the indiscriminate distribution of such traditional inhibitor drugs limits our ability to resolve how these mitotic enzymes function at distinct organelles. Here we exploit the SNAP‐tag system to create genetically encoded platforms that restrain kinase inhibitor drugs at defined subcellular locations. Local Kinase Inhibition (LoKI) allows us to ascribe organelle‐specific functions to broad‐specificity kinases. Using a combination of biochemical approaches, quantitative imaging, and live‐cell microscopy we uncover that local targeting of Plk1 and AurA kinase inhibitors to centrosomes disrupts substrate phosphorylation, spindle formation, and mitotic duration more profoundly than global drug distribution in human cells. Furthermore, we show that zebrafish embryos treated with centrosome‐targeted Plk1 inhibitors have more microtubule abnormalities than those treated with a non‐localized inhibitor. Finally, we expand our approach by targeting drugs to the kinetochore and find that inhibition of kinetochore‐associated pools of AurA blocks phosphorylation of microtubule‐kinetochore components. Future work will employ LoKI to investigate how downstream substrates of Plk1 and AurA function at centrosomes and kinetochore to coordinate mitotic signaling events.

Targeting Plk1 with siRNNs in primary cells from pediatric B-cell acute lymphoblastic leukemia patients

B-cell acute lymphoblastic leukemia (B-ALL) accounts for nearly one fifth of all childhood cancers and current challenges in B-ALL treatment include resistance, relapse and late-onset side effects due to the chemotherapy. To overcome these hurdles, novel therapies need to be investigated. One promising target is Polo-like kinase 1 (Plk1), a key regulator of the cell cycle. In this study, the Plk family expression is investigated in primary peripheral blood and bone marrow mononuclear cells from ten pediatric B-ALL patients. For the first time, short interfering RiboNucleic Neutrals (siRNNs) that enter cells without a transfection reagent are used to target Plk1 mRNA in primary cells from pediatric B-ALL patients. Our results show that the expression of Plk1 and Plk4 is significantly higher in pediatric B-ALL patients compared to healthy donors. Moreover, treatment of primary peripheral blood and bone marrow mononuclear cells from pediatric B-ALL patients, cultured ex vivo, with Plk1-targeting siRNNs results in cleavage of Plk1 mRNA. Importantly, the Plk1 knockdown is specific and does not affect other Plk members in contrast to many small molecule Plk1 inhibitors. Thus, Plk1 is a potential therapeutic target in pediatric B-ALL and selective targeting of Plk1 can be achieved by the use of siRNNs.

Abstract 302: The combined inhibition of Plk1 and Notch1 results in a synergistic anti-proliferative response in human melanoma cells

Abstract Melanoma is the most aggressive form of skin cancer, and its incidence and mortality have been increasing over the past thirty years. Therefore, additional therapeutic strategies are needed for an effective management of this neoplasm. We previously demonstrated that polo-like kinase 1 (Plk1, a serine/threonine kinase involved in mitotic regulation) and Notch1 (a type I transmembrane protein deciding cell fate during development) are strongly correlated in melanoma (Cancer Res 2018; 78 (13 Suppl): Abstract nr 2530). In addition, a number of studies from our lab and elsewhere have suggested the important functions of Plk1- as well as Notch- signaling in melanoma progression. Based on these observations, in this study, we determined the potential efficacy of a combined inhibition of Plk1 and Notch1 against melanoma cells. Employing Plk1 inhibitor volasertib (BI6727) and Notch1 inhibitor MK-0752, we determined the effects of concomitant targeting of these pathways in A375 (wild type TP53) and SK-MEL-2 (mutant TP53) human melanoma cell lines. Employing RealTime-Glo and trypan blue exclusion assays, we found that volasertib (10 and 20 nM) and MK-0752 (50 and 100 μM) resulted in a significant decrease (60-80%) in viability and growth of human melanoma cells A375 and SK-MEL-2. The Combination Index (CI), as calculated using the Chou-Talalay theorem, was less than 1 when volasertib at 10 nM was given with MK-0752 at 50 and 100 μM, indicating a synergism between these two drugs. As a result of the synergism, the melanoma cells treated with combined drugs showed decreased colony formation ability compared to individual drugs. In addition, the combined treatment also resulted in enhanced apoptosis of melanoma cells. Interestingly, cell cycle analysis by flow cytometry showed that combined inhibition of Plk1 and Notch1 caused a G2-M phase arrest of melanoma cells. Furthermore, our data demonstrated that volasertib and MK-0752 combination caused a marked increase of cleaved PARP (poly ADP ribose polymerase) and cleaved caspase-3 in both melanoma cell lines, as well as increased expression of tumor suppressor protein TP53 and its target p21 (CDKN1A) in A375 melanoma cells. Taken together, our data demonstrated that a combined targeting of Plk1- and Notch1- signaling pathways imparts a synergistic anti-proliferative response against melanoma. However, additional experiments are needed i) to validate our in vitro data to in vivo situations in human relevant melanoma models, and ii) to define the interactive mechanism(s) of the observed synergistic response. Citation Format: Shengqin Su, Gagan Chhabra, Chandra K. Singh, Mary A. Ndiaye, Nihal Ahmad. The combined inhibition of Plk1 and Notch1 results in a synergistic anti-proliferative response in human melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 302.

Synergistic activity of BET inhibitor MK-8628 and PLK inhibitor Volasertib in preclinical models of medulloblastoma

Medulloblastoma is the most prevalent central nervous system tumor in children. Targeted treatment approaches for patients with high-risk medulloblastoma are needed as current treatment regimens are not curative in many cases and cause significant therapy-related morbidity. Medulloblastoma harboring MYC amplification have the most aggressive clinical course and worst outcome. Targeting the BET protein BRD4 has significant anti-tumor effects in preclinical models of MYC-amplified medulloblastoma, however, in most cases these are not curative. We here assessed the therapeutic efficacy of the orally bioavailable BRD4 inhibitor, MK-8628, in preclinical models of medulloblastoma. MK-8628 showed therapeutic efficacy against in vitro and in vivo models of MYC-amplified medulloblastoma by inducing apoptotic cell death and cell cycle arrest. Gene expression analysis of cells treated with MK-8628 showed that anti-tumor effects were accompanied by significant repression of MYC transcription as well as disruption of MYC-regulated transcriptional programs. Additionally, we found that targeting of MYC protein stability through pharmacological PLK1 inhibition showed synergistic anti-medulloblastoma effects when combined with MK-8628 treatment. Thus, MK-8628 is effective against preclinical high-risk medulloblastoma models and its effects can be enhanced through simultaneous targeting of PLK1.

Co-targeting PLK1 and mTOR induces synergistic inhibitory effects against esophageal squamous cell carcinoma.

PLK1 potentiates both mTORC1 and mTORC2 activities in ESCC cells. PLK1 expression positively correlated with mTOR activity in a subset of ESCC. Co-targeting of PLK1 and mTOR produced stronger antitumor effects partially due to synergistic inhibition of AKT, 4E-BP1 and S6 through cooperatively blocking mTORC2 and mTORC1 cascades. Combination targeting of PLK1 and mTOR may be a novel and promising therapeutic strategy for ESCC treatment.

Amphiphilic nanocarrier-induced modulation of PLK1 and miR-34a leads to improved therapeutic response in pancreatic cancer

The heterogeneity of pancreatic ductal adenocarcinoma (PDAC) suggests that successful treatment might rely on simultaneous targeting of multiple genes, which can be achieved by RNA interference-based therapeutic strategies. Here we show a potent combination of microRNA and siRNA delivered by an efficient nanocarrier to PDAC tumors. Using proteomic-microRNA profiles and survival data of PDAC patients from TCGA, we found a novel signature for prolonged survival. Accordingly, we used a microRNA-mimic to increase miR-34a together with siRNA to silence PLK1 oncogene. For in vivo dual-targeting of this combination, we developed a biodegradable amphiphilic polyglutamate amine polymeric nanocarrier (APA). APA-miRNA–siRNA polyplexes systemically administered to orthotopically inoculated PDAC-bearing mice showed no toxicity and accumulated at the tumor, resulting in an enhanced antitumor effect due to inhibition of MYC oncogene, a common target of both miR-34a and PLK1. Taken together, our findings warrant this unique combined polyplex’s potential as a novel nanotherapeutic for PDAC.

In silico identification of putative bifunctional Plk1 inhibitors by integrative virtual screening and structural dynamics approach

Polo like kinase (Plk1) is a master regulator of cell cycle and considered as next generation antimitotic target in human. As Plk1 predominantly expresses in the dividing cells with a much higher expression in cancerous cells, it serves as a discriminative target for cancer therapeutics. Here we implied a novel and promising integrative strategy to identify "bifunctional" Plk1 inhibitors that compete simultaneously with ATP and substrate for their binding sites. We integrated structure-based virtual screening (SBVS) and molecular dynamics simulations with emphasis on unique structural properties of Plk1. Through screening of 20,000 compounds, nearly ~2000 hits were enriched and subjected to SBVS against ATP and substrate binding sites of Plk1. Subsequently, on the basis of their binding abilities to Plk1 kinase and polo box domains, filtration of candidate hits resulted in the isolation of 26 compounds. By exclusion of close analogs or isomers, 10 unique compounds were selected for detailed study. A representative compound was subjected to molecular dynamics simulation assay to have deep structural insights and to gauge critical structural crunch for inhibitor binding against kinase and polo box domains. Our integrative approach may complement high-throughput screening and identify bifunctional Plk1 inhibitors that may contribute in selective targeting of Plk1 to elicit desired biological process.

Abstract 1991: FOXM1 expression in epithelial ovarian cancer coincides with gene amplification and functional loss of p53 and Rb and drives G2-M progression and target gene expression

Abstract Introduction: According to The Cancer Genome Atlas (TCGA), the FOXM1 transcription factor network is activated in 84% of high-grade serous ovarian cancers (HGS-OC), a common and lethal subtype of epithelial ovarian cancer (EOC). FOXM1 plays a key role in cell cycle progression and genomic stability, and is overexpressed in various malignancies. The goal of this study was to understand the genetic determinants of FOXM1 expression in EOC, and to define its functional role in relevant cell models. Methods: We analyzed FOXM1 gene amplification data in HGS-OC and other cancers using cBioPortal. We determined FOXM1 expression and its relationship to clinico-pathology in a large set of primary EOC tissues. We used primary and immortalized murine (MOSE) and human ovarian surface epithelial (hOSE) cells to determine the contribution of p53, Rb, and E2F1 to FOXM1 expression. We used FOXM1 knockdown and overexpression approaches to dissect its functional contribution to cell cycle progression and target gene activation in OSE and EOC cells. We used human EOC cell lines to assess genetic mechanisms driving the expression of the three known FOXM1 splice variants. Results: The FOXM1 locus was amplified in ∼12% of HGS-OC, greater than any other tumor type examined and suggestive of selective pressure in HGS-OC. FOXM1 mRNA was frequently upregulated in EOC, in association with advanced stage and grade. In MOSE cells, combined knockout of Rb1 and Trp53 synergistically induced Foxm1. In agreement, hOSE cells immortalized with SV40 T antigen (IOSE-SV) showed dramatically higher FOXM1 expression as compared to hOSE immortalized with hTERT. Knockdown experiments revealed that FOXM1 induction in IOSE-SV is partially dependent on E2F1. FOXM1 knockdown in IOSE-SV blocked G2-M transition and downregulated known targets of FOXM1, including CCNB1 and PLK1. Conversely, FOXM1 overexpression in primary hOSE upregulated FOXM1 target genes. FOXM1 expression in EOC cell lines was heterogeneous and the highest expression was observed in cell lines with FOXM1 gene amplification and TP53 and RB1 mutations. The level of expression of FOXM1 splice variants in EOC cell lines was consistently FOXM1c &amp;gt; FOXM1b &amp;gt; FOXM1a, indicating that transcriptionally active isoforms show higher expression. Conclusions: Our data reveal that genetic alterations in TP53 and RB1, along with FOXM1 gene amplification, are critical determinants of FOXM1 expression in EOC. FOXM1 functionally contributes to cell cycle progression in EOC models, likely via the transactivation of gene targets driving G2-M transition. Current studies are dissecting the role of post-translational modifications in FOXM1 activation in EOC, and extending work into relevant human fallopian tube secretory epithelial cell systems. Finally, our findings support the development of FOXM1-targeted therapies for EOC. Citation Format: Carter J. Barger, Wa Zhang, Kunle Odunsi, Adam R. Karpf. FOXM1 expression in epithelial ovarian cancer coincides with gene amplification and functional loss of p53 and Rb and drives G2-M progression and target gene expression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1991. doi:10.1158/1538-7445.AM2015-1991

Plk1 phosphorylation of PTEN causes a tumor-promoting metabolic state.

One outcome of activation of the phosphatidylinositol 3-kinase (PI3K) pathway is increased aerobic glycolysis, but the upstream signaling events that regulate the PI3K pathway, and thus the Warburg effect, are elusive. Increasing evidence suggests that Plk1, a cell cycle regulator, is also involved in cellular events in addition to mitosis. To test whether Plk1 contributes to activation of the PI3K pathway, and thus aerobic glycolysis, we examined potential targets of Plk1 and identified PTEN as a Plk1 substrate. We hypothesize that Plk1 phosphorylation of PTEN leads to its inactivation, activation of the PI3K pathway, and the Warburg effect. Our data show that overexpression of Plk1 leads to activation of the PI3K pathway and enhanced aerobic glycolysis. In contrast, inhibition of Plk1 causes markedly reduced glucose metabolism in mice. Mechanistically, we show that Plk1 phosphorylation of PTEN and Nedd4-1, an E3 ubiquitin ligase of PTEN, results in PTEN inactivation. Finally, we show that Plk1 phosphorylation of PTEN promotes tumorigenesis in both its phosphatase-dependent and -independent pathways, revealing potentially new drug targets to arrest tumor cell growth.

The Role of Polo-like Kinase 1 in Carcinogenesis: Cause or Consequence?

Polo-like kinase 1 (Plk1) is a well-established mitotic regulator with a diverse range of biologic functions continually being identified throughout the cell cycle. Preclinical evidence suggests that the molecular targeting of Plk1 could be an effective therapeutic strategy in a wide range of cancers; however, that success has yet to be translated to the clinical level. The lack of clinical success has raised the question of whether there is a true oncogenic addiction to Plk1 or if its overexpression in tumors is solely an artifact of increased cellular proliferation. In this review, we address the role of Plk1 in carcinogenesis by discussing the cell cycle and DNA damage response with respect to their associations with classic oncogenic and tumor suppressor pathways that contribute to the transcriptional regulation of Plk1. A thorough examination of the available literature suggests that Plk1 activity can be dysregulated through key transformative pathways, including both p53 and pRb. On the basis of the available literature, it may be somewhat premature to draw a definitive conclusion on the role of Plk1 in carcinogenesis. However, evidence supports the notion that oncogene dependence on Plk1 is not a late occurrence in carcinogenesis and it is likely that Plk1 plays an active role in carcinogenic transformation.