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Polo-like kinase 1 (PLK1) is a serine/threonine–protein kinase essential for regulating multiple stages of cell cycle progression in mammals. Aberrant regulation of PLK1 has been observed in numerous human cancers and is linked to poor prognoses. However, its role in the pathogenesis of colorectal cancer (CRC) in the Middle East remains unexplored. PLK1 overexpression was noted in 60.3% (693/1149) of CRC cases and was significantly associated with aggressive clinico-pathological parameters and p-ERK1/2 overexpression. Intriguingly, multivariate logistic regression analysis identified PLK1 as an independent predictor of lymph node metastasis. Our in vitro experiments demonstrated that CRC cells with high PLK1 levels were resistant to 5-Fu treatment, while those with low PLK1 expression were sensitive. To investigate PLK1′s role in chemoresistance, we used the specific inhibitor volasertib, which effectively reversed 5-Fu resistance. Interestingly, forced PLK1 expression activated the CRAF-MEK-ERK signaling cascade, while its inhibition suppressed this cascade. PLK1 knockdown reduced epithelial-to-mesenchymal transition (EMT) progression and stem cell-like traits in 5-Fu-resistant cells, implicating PLK1 in EMT induction and stemness in CRC. Moreover, silencing ERK1/2 significantly mitigated chemoresistance, EMT, and stemness properties in CRC cell lines that express PLK1. Furthermore, the knockdown of Zeb1 attenuated EMT and stemness, suggesting a possible link between EMT activation and the maintenance of stemness in CRC. Our findings underscore the pivotal role of PLK1 in mediating chemoresistance and suggest that PLK1 inhibition may represent a potential therapeutic strategy for the management of aggressive colorectal cancer subtypes. [ABSTRACT FROM AUTHOR]

Simple Summary: Patients with hormone receptor-positive (HR+)/HER2-negative metastatic breast cancer who have PIK3CA mutations and have progressed on first-line therapies are considered for treatment with alpelisib alongside endocrine therapy (ET). Combination therapeutic strategies may extend the clinical benefit of alpelisib. This study investigated the potential of onvansertib, a highly specific inhibitor of polo-like kinase 1 (PLK1), to enhance the efficacy of alpelisib in preclinical models of HR+ breast cancer. Our findings demonstrated that onvansertib synergizes with alpelisib in PIK3CA-mutant HR+ breast cancer cell lines and patient-derived xenografts that are resistant to ET and cyclin-dependent kinase 4/6 inhibitors. The combination inhibited PLK1 and PI3K signaling, induced cell cycle arrest, and triggered apoptotic death in both cell lines and xenograft tumors. Overall, our preclinical data encourage the clinical exploration of this combination for PIK3CA-mutant HR+ metastatic breast cancer patients progressing on standard-of-care therapies. Background: Endocrine therapy (ET) combined with cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) is the preferred first-line treatment for hormone receptor-positive (HR+)/HER2- metastatic breast cancer. Although this is beneficial, acquired resistance leads to disease progression, and patients harboring PIK3CA mutations are treated with targeted therapies such as the PI3Kα inhibitor, alpelisib, alongside ET. Drug-associated resistance mechanisms limit the efficacy of alpelisib, highlighting the need for better combination therapies. This study aimed to evaluate the efficacy of combining alpelisib with a highly specific PLK1 inhibitor, onvansertib, in PIK3CA-mutant HR+ breast cancer preclinical models. Methods: We assessed the effect of the alpelisib and onvansertib combination on cell viability, PI3K signaling pathway, cell cycle phase distribution and apoptosis in PI3K-activated HR+ breast cancer cell lines. The antitumor activity of the combination was evaluated in three PIK3CA-mutant HR+ breast cancer patient-derived xenograft (PDX) models, resistant to ET and CDK4/6 inhibitor palbociclib. Pharmacodynamics studies were performed using immunohistochemistry and Simple Western analyses in tumor tissues. Results: The combination synergistically inhibited cell viability, suppressed PI3K signaling, induced G2/M arrest and apoptosis in PI3K-activated cell lines. In the three PDX models, the combination demonstrated superior anti-tumor activity compared to the single agents. Pharmacodynamic studies confirmed the inhibition of both PLK1 and PI3K activity and pronounced apoptosis in the combination-treated tumors. Conclusions: Our findings support that targeting PLK1 and PI3Kα with onvansertib and alpelisib, respectively, may be a promising strategy for patients with PIK3CA-mutant HR+ breast cancer failing ET + CDK4/6i therapies and warrant clinical evaluation. [ABSTRACT FROM AUTHOR]

The accuracy of cell division requires precise regulation of the cellular machinery governing DNA/genome duplication, ensuring its equal distribution among the daughter cells. The control of the centrosome cycle is crucial for the formation of a bipolar spindle, ensuring error‐free segregation of the genome. The cell and centrosome cycles operate in close synchrony along similar principles. Both require a single duplication round in every cell cycle, and both are controlled by the activity of key protein kinases. Nevertheless, our comprehension of the precise cellular mechanisms and critical regulators synchronizing these two cycles remains poorly defined. Here, we present our hypothesis that the spatiotemporal regulation of a dynamic equilibrium of mitotic kinases activities forms a molecular clock that governs the synchronous progression of both the cell and the centrosome cycles. [ABSTRACT FROM AUTHOR]

Polo-like kinase 1 (Plk1), an evolutionarily conserved serine/threonine protein kinase, is a key regulator involved in the mitotic process of the cell cycle. Mounting evidence suggests that Plk1 is also involved in a variety of nonmitotic events, including the DNA damage response, DNA replication, cytokinesis, embryonic development, apoptosis, and immune regulation. The DNA damage response (DDR) includes activation of the DNA checkpoint, DNA damage recovery, DNA repair, and apoptosis. Plk1 is not only an important target of the G2/M DNA damage checkpoint but also negatively regulates the G2/M checkpoint commander Ataxia telangiectasia-mutated (ATM), promotes G2/M phase checkpoint recovery, and regulates homologous recombination repair by interacting with Rad51 and BRCA1, the key factors of homologous recombination repair. This article briefly reviews the function of Plk1 in response to DNA damage. [ABSTRACT FROM AUTHOR]

Oocyte meiotic maturation failure and chromosome abnormality is one of the main causes of infertility, abortion, and diseases. The mono-orientation of sister chromatids during the first meiosis is important for ensuring accurate chromosome segregation in oocytes. MEIKIN is a germ cell-specific protein that can regulate the mono-orientation of sister chromatids and the protection of the centromeric cohesin complex during meiosis I. Here we found that MEIKIN is a maternal protein that was highly expressed in mouse oocytes before the metaphase I (MI) stage, but became degraded by the MII stage and dramatically reduced after fertilization. Strikingly, MEIKIN underwent phosphorylation modification after germinal vesicle breakdown (GVBD), indicating its possible function in subsequent cellular event regulation. We further showed that MEIKIN phosphorylation was mediated by PLK1 at its carboxyl terminal region and its C-terminus was its key functional domain. To clarify the biological significance of meikin degradation during later stages of oocyte maturation, exogenous expression of MEIKIN was employed, which showed that suppression of MEIKIN degradation resulted in chromosome misalignment, cyclin B1 and Securin degradation failure, and MI arrest through a spindle assembly checkpoint (SAC)-independent mechanism. Exogenous expression of MEIKIN also inhibited metaphase II (MII) exit and early embryo development. These results indicate that proper MEIKIN expression level and its C-terminal phosphorylation by PLK1 are critical for regulating the metaphase–anaphase transition in meiotic oocyte. The findings of this study are important for understanding the regulation of chromosome segregation and the prevention meiotic abnormality. [ABSTRACT FROM AUTHOR]

Abstract Echovirus 30 (E30), a member of the species B Enterovirus family, is a primary pathogen responsible for aseptic meningitis and encephalitis. E30 is associated with severe nervous system diseases and is a primary cause of child illness, disability, and even mortality. However, the mechanisms underlying E30-induced brain injury remain poorly understood. In this study, we used a neonatal mouse model of E30 to investigate the possible mechanisms of brain injury. E30 infection triggered the activation of microglia in the mouse brain and efficiently replicated within HMC3 cells. Subsequent transcriptomic analysis revealed inflammatory activation of microglia in response to E30 infection. We also detected a significant upregulation of polo-like kinase 1 (PLK1) and found that its inhibition could limit E30 infection in a sucking mouse model. Collectively, E30 infection led to brain injury in a neonatal mouse model, which may be related to excessive inflammatory responses. Our findings highlight the intricate interplay between E30 infection and neurological damage, providing crucial insights that could guide the development of interventions and strategies to address the severe clinical manifestations associated with this pathogen.

Background: Polo-like kinase 1 (PLK1) is a critical therapeutic target in the treatment of head and neck squamous cell carcinoma (HNSCC). The objective of this study was to investigate the therapeutic effect of the combination of BI 2536, a PLK1 inhibitor, and erastin, a ferroptosis inducer, in HNSCC. Methods: The proliferation, invasion, and migration abilities of Tu177 and FaDu cells upon exposure to BI 2536 and erastin, used in combination or alone, were tested. Fe2+, glutathione (GSH), and malondialdehyde (MDA) detection kits were used to determine whether the addition of BI 2536 enhanced the accumulation of Fe2+ and MDA, along with the depletion of GSH. Quantitative real-time PCR, western blot analyses were performed to investigate whether BI 2536 further altered the mRNA and expression level of ferroptosis genes. Furthermore, si PLK1 was used to investigate whether targeting PLK1 gene promoted erastin-induced ferroptosis. Results: The combination of BI 2536 and erastin exerted a stronger cytotoxicity than treatment with a single agent. Compared with erastin treatment alone, the combination of BI 2536 and erastin lowered the ability of tumor cells to self-clone, invade, and migrate. BI 2536 enhanced the accumulation of Fe2+ and MDA, and the depletion of GSH. BI 2536 increased erastin-induced changes in ferroptosis-related gene mRNA and expression. Importantly, targeting PKL1 enhanced the anti-cancer effect of erastin. Conclusion: BI 2536 enhanced the sensitivity of HNSCC cells to erastin, which provides a new perspective for cancer treatment. [ABSTRACT FROM AUTHOR]

Inflammatory signals from immunological cells may cause damage to intestinal epithelial cells (IECs), resulting in intestinal inflammation and tissue impairment. Interferon‐γ‐inducible protein 16 (IFI16) was reported to be involved in the pathogenesis of Behçet's syndrome (BS). This study aimed to investigate how inflammatory cytokines released by immunological cells and IFI16 participate in the pathogenesis of intestinal BS. RNA sequencing and real‐time quantitative PCR (qPCR) showed that the positive regulation of tumor necrosis factor‐α (TNF‐α) production in peripheral blood mononuclear cells (PBMCs) of intestinal BS patients may be related to the upregulation of polo like kinase 1 (PLK1) in PBMCs (P = 0.012). The plasma TNF‐α protein level in intestinal BS was significantly higher than in healthy controls (HCs; P = 0.009). PBMCs of intestinal BS patients and HCs were co‐cultured with human normal IECs (NCM460) to explore the interaction between immunological cells and IECs. Using IFI16 knockdown, PBMC‐NCM460 co‐culture, TNF‐α neutralizing monoclonal antibody (mAb), stimulator of interferon genes (STING) agonist 2′3′‐cGAMP, and the PLK1 inhibitor SBE 13 HCL, we found that PLK1 promotes the secretion of TNF‐α from PBMCs of intestinal BS patients, which causes overexpression of IFI16 and induces apoptosis of IECs via the STING–TBK1 pathway. The expressions of IFI16, TNF‐α, cleaved caspase 3, phosphorylated STING (pSTING) and phosphorylated tank binding kinase 1 (pTBK1) in the intestinal ulcer tissue of BS patients were significantly higher than that of HCs (all P < 0.05). PLK1 in PBMCs of intestinal BS patients increased TNF‐α secretion, inducing IEC apoptosis via activation of the IFI16–STING–TBK1 pathway. PLK1 and the IFI16–STING–TBK1 pathway may be new therapeutic targets for intestinal BS. [ABSTRACT FROM AUTHOR]

Echovirus 30 (E30), a member of the species B Enterovirus family, is a primary pathogen responsible for aseptic meningitis and encephalitis. E30 is associated with severe nervous system diseases and is a primary cause of child illness, disability, and even mortality. However, the mechanisms underlying E30-induced brain injury remain poorly understood. In this study, we used a neonatal mouse model of E30 to investigate the possible mechanisms of brain injury. E30 infection triggered the activation of microglia in the mouse brain and efficiently replicated within HMC3 cells. Subsequent transcriptomic analysis revealed inflammatory activation of microglia in response to E30 infection. We also detected a significant upregulation of polo-like kinase 1 (PLK1) and found that its inhibition could limit E30 infection in a sucking mouse model. Collectively, E30 infection led to brain injury in a neonatal mouse model, which may be related to excessive inflammatory responses. Our findings highlight the intricate interplay between E30 infection and neurological damage, providing crucial insights that could guide the development of interventions and strategies to address the severe clinical manifestations associated with this pathogen. [ABSTRACT FROM AUTHOR]

Novel therapeutic approaches are needed for the treatment of Ewing sarcoma tumors. We previously identified that Ewing sarcoma cell lines are sensitive to drugs that inhibit protein translation. However, translational and therapeutic approaches to inhibit protein synthesis in tumors are limited. In this work, we identified that reactive oxygen species, which are generated by a wide range of chemotherapy and other drugs, inhibit protein synthesis and reduce the level of critical proteins that support tumorigenesis in Ewing sarcoma cells. In particular, we identified that both hydrogen peroxide and auranofin, an inhibitor of thioredoxin reductase and regulator of oxidative stress and reactive oxygen species, activate the repressor of protein translation 4E-BP1 and reduce the levels of the oncogenic proteins RRM2 and PLK1 in Ewing and other sarcoma cell lines. These results provide novel insight into the mechanism of how ROSinducing drugs target cancer cells via inhibition of protein translation and identify a mechanistic link between ROS and the DNA replication (RRM2) and cell cycle regulatory (PLK1) pathways. [ABSTRACT FROM AUTHOR]

This review aims to outline mitotic kinase inhibitors' roles as potential therapeutic targets and assess their suitability as a stand-alone clinical therapy or in combination with standard treatments for advanced-stage solid tumors, including triple-negative breast cancer (TNBC). Breast cancer poses a significant global health risk, with TNBC standing out as the most aggressive subtype. Comprehending the role of mitosis is crucial for understanding how TNBC advances from a solid tumor to metastasis. Chemotherapy is the primary treatment used to treat TNBC. Some types of chemotherapeutic agents target cells in mitosis, thus highlighting the need to comprehend the molecular mechanisms governing mitosis in cancer. This understanding is essential for devising targeted therapies to disrupt these mitotic processes, prevent or treat metastasis, and improve patient outcomes. Mitotic kinases like Aurora kinase A, Aurora Kinase B, never in mitosis gene A-related kinase 2, Threonine-Tyrosine kinase, and Polo-kinase 1 significantly impact cell cycle progression by contributing to chromosome separation and centrosome homeostasis. When these kinases go awry, they can trigger chromosome instability, increase cell proliferation, and activate different molecular pathways that culminate in a transition from epithelial to mesenchymal cells. Ongoing clinical trials investigate various mitotic kinase inhibitors as potential biological treatments against advanced solid tumors. While clinical trials against mitotic kinases have shown some promise in the clinic, more investigation is necessary, since they induce severe adverse effects, particularly affecting the hematopoietic system. [ABSTRACT FROM AUTHOR]

This article explores the role of Polo-like kinase 1 (PLK1) in tumorigenesis, specifically in colorectal cancer (CRC) and lung cancer. The study analyzes the mutational landscape of PLK1 and concomitant mutations in these cancers. The findings suggest that PLK1 may have tumor-suppressor features and caution should be exercised when using PLK1 inhibitors. The study also identifies different mutation subtypes of PLK1 and their associations with other genetic abnormalities in CRC. However, the study has limitations, such as missing information on cancer stage and treatment data, and further research is needed to validate the findings. [Extracted from the article]

During mitosis, chromosomes assemble kinetochores to dynamically couple with spindle microtubules.1,2 Kinetochores also function as signaling hubs directing mitotic progression by recruiting and controlling the fate of the anaphase promoting complex/cyclosome (APC/C) activator CDC-20.3,4,5 Kinetochores either incorporate CDC-20 into checkpoint complexes that inhibit the APC/C or dephosphorylate CDC-20, which allows it to interact with and activate the APC/C.4,6 The importance of these two CDC-20 fates likely depends on the biological context. In human somatic cells, the major mechanism controlling mitotic progression is the spindle checkpoint. By contrast, progression through mitosis during the cell cycles of early embryos is largely checkpoint independent.7,8,9,10 Here, we first show that CDC-20 phosphoregulation controls mitotic duration in the C. elegans embryo and defines a checkpoint-independent temporal mitotic optimum for robust embryogenesis. CDC-20 phosphoregulation occurs at kinetochores and in the cytosol. At kinetochores, the flux of CDC-20 for local dephosphorylation requires an ABBA motif on BUB-1 that directly interfaces with the structured WD40 domain of CDC-20.6,11,12,13 We next show that a conserved STP motif in BUB-1 that docks the mitotic kinase PLK-114 is necessary for CDC-20 kinetochore recruitment and timely mitotic progression. The kinase activity of PLK-1 is required for CDC-20 to localize to kinetochores and phosphorylates the CDC-20-binding ABBA motif of BUB-1 to promote BUB-1-CDC-20 interaction and mitotic progression. Thus, the BUB-1-bound pool of PLK-1 ensures timely mitosis during embryonic cell cycles by promoting CDC-20 recruitment to the vicinity of kinetochore-localized phosphatase activity.

This research aims to investigate the role and underlying biological mechanism of FBXO45 in regulating ferroptosis of renal fibrocytes in a diabetic nephropathy (DN) model.

Polo‐like kinase 1 (PLK1; also known as serine/threonine‐protein kinase PLK1) serves as a central player in cell proliferation, exerting critical regulatory roles in mitotic processes and cell survival. We conducted an analysis of PLK1 protein expression in a large cohort of samples from papillary thyroid carcinoma (PTC) patients and examined its functional significance in PTC cell lines, both in vitro and in vivo. PLK1 overexpression was noted in 54.2% of all PTC and was significantly associated with aggressive clinicopathological parameters; it was also found to be an independent prognostic marker for shorter recurrence‐free survival. Given the significant association between PLK1 and forkhead box protein M1 (FoxM1), and their concomitant overexpression in a large proportion of PTC samples, we explored their correlation and their combined inhibitions in PTC in vitro and in vivo. Inhibition of PLK1 expression indeed suppressed cell proliferation, leading to cell cycle arrest and apoptosis in PTC cell lines. Significantly, the downregulation of PLK1 reduced the self‐renewal capability of spheroids formed from PTC cells. Immunoprecipitation analysis shows that PLK1 binds to FoxM1 and vice versa in vitro. Mechanistically, PLK1 knockdown suppresses FoxM1 expression, whereas inhibition of FoxM1 does not affect PLK1 expression, which suggests that PLK1 acts through the FoxM1 pathway. The combined treatment of a PLK1 inhibitor (volasertib) and a FoxM1 inhibitor (thiostrepton) demonstrated a synergistic effect in reducing PTC cell growth in vitro and delaying tumor growth in vivo. This study highlights the important role of PLK1 in PTC tumorigenesis and prognosis. It also highlights the synergistic therapeutic potential of dual‐targeting PLK1 and FoxM1 in PTC, unveiling a potential innovative therapeutic strategy for managing aggressive forms of PTC.

Osteosarcoma is the most common primary bone malignancy in children and adolescents. Overexpression of polo‐like kinase 1 (PLK1) is frequent in osteosarcoma and drives disease progression and metastasis, making it a promising therapeutic target. In this study, we explored PLK1 knockdown in osteosarcoma cells using RNA interference mediated by high‐fidelity Cas13d (hfCas13d). PLK1 was found to be significantly upregulated in osteosarcoma tumour tissues compared to normal bone. sgRNA‐mediated PLK1 suppression via hfCas13d transfection inhibited osteosarcoma cell proliferation, induced G2/M cell cycle arrest, promoted apoptosis, reduced cell invasion and increased expression of the epithelial marker E‐cadherin. Proximity labelling by TurboID coupled with co‐immunoprecipitation identified novel PLK1 interactions with Smad3, a key intracellular transducer of TGF‐β signalling. PLK1 knockdown impaired Smad2/3 phosphorylation and modulated TGF‐β/Smad3 pathway inactivation. Finally, in vivo delivery of hfCas13d vectors targeting PLK1 substantially attenuated osteosarcoma xenograft growth in nude mice. Taken together, this study highlights PLK1 as a potential therapeutic target and driver of disease progression in osteosarcoma. It also demonstrates the utility of hfCas13d‐mediated gene knockdown as a strategy for targeted therapy. Further optimization of PLK1 suppression approaches may ultimately improve clinical outcomes for osteosarcoma patients. [ABSTRACT FROM AUTHOR]

Background: 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. Methods: 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. Results: 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. Conclusions: 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. Key points: 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. [ABSTRACT FROM AUTHOR]

The host cytoskeleton plays crucial roles in various stages of virus infection, including viral entry, transport, replication, and release. However, the specific mechanisms by which intermediate filaments are involved in orthoflavivirus infection have not been well understood. In this study, we demonstrate that the Japanese encephalitis virus (JEV) remodels the vimentin network, resulting in the formation of cage-like structures that support viral replication. Mechanistically, JEV NS1 and NS1' proteins induce the translocation of CDK1 from the nucleus to the cytoplasm and interact with it, leading to the phosphorylation of vimentin at Ser56. This phosphorylation event recruits PLK1, which further phosphorylates vimentin at Ser83. Consequently, these phosphorylation modifications convert the typically filamentous vimentin into non-filamentous "particles" or "squiggles." These vimentin "particles" or "squiggles" are then transported retrogradely along microtubules to the endoplasmic reticulum, where they form cage-like structures. Notably, NS1' is more effective than NS1 in triggering the CDK1-PLK1 cascade response. Overall, our study provides new insights into how JEV NS1 and NS1' proteins manipulate the vimentin network to facilitate efficient viral replication. [ABSTRACT FROM AUTHOR]

Simple Summary: The kinesin superfamily is an important member of motor proteins, playing important roles in cell growth and development. Studies on KIF2A of the kinesin-13 family have mainly focused on cell division, while the relationship between KIF2A and signaling pathways remains unclear. The purpose of the present study was to determine whether KIF2A has a regulatory effect on the PI3K/AKT pathway, thus affecting spermatogenic cell proliferation and apoptosis during spermatogenesis, and whether its interacting protein PLK1 is involved in this process. RNAi experiments were performed in our study. Our results showed that KIF2A upregulates the PI3K/AKT signaling pathway through its interacting protein PLK1 during the spermatogenesis of Eriocheir sinensis. E. sinensis is an animal model for studying the reproduction and development of crustaceans. In this study, we knocked down the Es-Kif2a gene by injecting dsRNA into E. sinensis and inhibited Es-Plk1 gene expression by injecting PLK1 inhibitor BI6727 into E. sinensis. Then, the cell proliferation level, apoptosis level, and PI3K/AKT signaling expression level were detected. Our results showed that the proliferation level of spermatogenic cells decreased, while the apoptosis level increased after Es-Kif2a knockdown or Es-Plk1 inhibition. In order to verify whether these changes are caused by regulating the PI3K/AKT pathway, we detected the expression of PI3K and AKT proteins after Es-Kif2a knockdown or Es-Plk1 inhibition. Western Blot showed that in both the Es-Kif2a knockdown group and the Es-Plk1 inhibition group, the expression of PI3K and AKT proteins decreased. In addition, immunofluorescence showed that Es-KIF2A and Es-PLK1 proteins were co-localized during E. sinensis spermatogenesis. To further explore the upstream and downstream relationship between Es-KIF2A and Es-PLK1, we detected the expression level of Es-PLK1 after Es-Kif2a knockdown as well as the expression level of Es-KIF2A after Es-Plk1 inhibition. Western Blot showed that the expression of Es-PLK1 decreased after Es-Kif2a knockdown, while there was no significant change of Es-KIF2A after Es-Plk1 inhibition, indicating that Es-PLK1 may be a downstream factor of Es-KIF2A. Taken together, these results suggest that Es-KIF2A upregulates the PI3K/AKT signaling pathway through Es-PLK1 during the spermatogenesis of E. sinensis, thereby affecting the proliferation and apoptosis levels of spermatogenic cells. [ABSTRACT FROM AUTHOR]

Elimination of virally infected or tumoral cells is mediated by cytotoxic T cells (CTL). Upon antigen recognition, CTLs assemble a specialized signaling and secretory domain at the interface with their target, the immune synapse (IS). During IS formation, CTLs acquire a transient polarity, marked by re-orientation of the centrosome and microtubule cytoskeleton toward the IS, thus directing the transport and delivery of the lytic granules to the target cell. Based on the implication that the kinase Aurora A has a role in CTL function, we hypothesized that its substrate, the mitotic regulator Polo-like kinase 1 (PLK1), might participate in CTL IS assembly. We demonstrate that PLK1 is phosphorylated upon TCR triggering and polarizes to the IS. PLK1 silencing or inhibition results in impaired IS assembly and function, as witnessed by defective synaptic accumulation of T cell receptors (TCRs), as well as compromised centrosome and lytic granule polarization to the IS, resulting in impaired target cell killing. This function is achieved by coupling early signaling to microtubule dynamics, a function pivotal for CTL-mediated cytotoxicity. These results identify PLK1 as a new player in CTL IS assembly and function. [ABSTRACT FROM AUTHOR]

Polo‐like kinase 1 (PLK1; also known as serine/threonine‐protein kinase PLK1) serves as a central player in cell proliferation, exerting critical regulatory roles in mitotic processes and cell survival. We conducted an analysis of PLK1 protein expression in a large cohort of samples from papillary thyroid carcinoma (PTC) patients and examined its functional significance in PTC cell lines, both in vitro and in vivo. PLK1 overexpression was noted in 54.2% of all PTC and was significantly associated with aggressive clinicopathological parameters; it was also found to be an independent prognostic marker for shorter recurrence‐free survival. Given the significant association between PLK1 and forkhead box protein M1 (FoxM1), and their concomitant overexpression in a large proportion of PTC samples, we explored their correlation and their combined inhibitions in PTC in vitro and in vivo. Inhibition of PLK1 expression indeed suppressed cell proliferation, leading to cell cycle arrest and apoptosis in PTC cell lines. Significantly, the downregulation of PLK1 reduced the self‐renewal capability of spheroids formed from PTC cells. Immunoprecipitation analysis shows that PLK1 binds to FoxM1 and vice versa in vitro. Mechanistically, PLK1 knockdown suppresses FoxM1 expression, whereas inhibition of FoxM1 does not affect PLK1 expression, which suggests that PLK1 acts through the FoxM1 pathway. The combined treatment of a PLK1 inhibitor (volasertib) and a FoxM1 inhibitor (thiostrepton) demonstrated a synergistic effect in reducing PTC cell growth in vitro and delaying tumor growth in vivo. This study highlights the important role of PLK1 in PTC tumorigenesis and prognosis. It also highlights the synergistic therapeutic potential of dual‐targeting PLK1 and FoxM1 in PTC, unveiling a potential innovative therapeutic strategy for managing aggressive forms of PTC. [ABSTRACT FROM AUTHOR]

BackgroundThreonine tyrosine kinase (TTK) and polo-like kinase 1 (PLK1) are common essential kinases that collaborate in activating the spindle assembly checkpoint (SAC) at the kinetochore, ensuring appropriate chromosome alignment and segregation prior to mitotic exit. Targeting of either TTK or PLK1 has been clinically evaluated in cancer patients; however, dual inhibitors have not yet been pursued. Here we present the in vitro and in vivo characterization of a first in class, dual TTK/PLK1 inhibitor (BAL0891).MethodsMechanism of action studies utilized biochemical kinase and proteomics-based target-engagement assays. Cellular end-point assays included immunoblot- and flow cytometry-based cell cycle analyses and SAC integrity evaluation using immunoprecipitation and immunofluorescence approaches. Anticancer activity was assessed in vitro using cell growth assays and efficacy was evaluated, alone and in combination with paclitaxel and carboplatin, using mouse models of triple negative breast cancer (TNBC).ResultsBAL0891 elicits a prolonged effect on TTK, with a transient activity on PLK1. This unique profile potentiates SAC disruption, forcing tumor cells to aberrantly exit mitosis with faster kinetics than observed with a TTK-specific inhibitor. Broad anti-proliferative activity was demonstrated across solid tumor cell lines in vitro. Moreover, intermittent intravenous single-agent BAL0891 treatment of the MDA-MB-231 mouse model of TNBC induced profound tumor regressions associated with prolonged TTK and transient PLK1 in-tumor target occupancy. Furthermore, differential tumor responses across a panel of thirteen TNBC patient-derived xenograft models indicated profound anticancer activity in a subset (~40%). Using a flexible dosing approach, pathologically confirmed cures were observed in combination with paclitaxel, whereas synergy with carboplatin was schedule dependent.ConclusionsDual TTK/PLK1 inhibition represents a novel approach for the treatment of human cancer, including TNBC patients, with a potential for potent anticancer activity and a favorable therapeutic index. Moreover, combination approaches may provide an avenue to expand responsive patient populations.

Novel therapeutic approaches are needed for the treatment of Ewing sarcoma tumors. We previously identified that Ewing sarcoma cell lines are sensitive to drugs that inhibit protein translation. However, translational and therapeutic approaches to inhibit protein synthesis in tumors are limited. In this work, we identified that reactive oxygen species, which are generated by a wide range of chemotherapy and other drugs, inhibit protein synthesis and reduce the level of critical proteins that support tumorigenesis in Ewing sarcoma cells. In particular, we identified that both hydrogen peroxide and auranofin, an inhibitor of thioredoxin reductase and regulator of oxidative stress and reactive oxygen species, activate the repressor of protein translation 4E-BP1 and reduce the levels of the oncogenic proteins RRM2 and PLK1 in Ewing and other sarcoma cell lines. These results provide novel insight into the mechanism of how ROS-inducing drugs target cancer cells via inhibition of protein translation and identify a mechanistic link between ROS and the DNA replication (RRM2) and cell cycle regulatory (PLK1) pathways.

Abstract Background Human discs large-associated protein 5 (DLGAP5) is reported to play a pivotal role in regulating the cell cycle and implicate in tumorigenesis and progression of various cancers. Our current research endeavored to explore the prognostic value, immune implication, biological function and targeting strategy of DLGAP5 in LUAD through approaches including bioinformatics, network pharmacology analysis and experimental study. Methods Multiple databases, including TCGA, GEO, CPTAC and Human Protein Atlas, were utilized to explore the expression and clinical significance of DLGAP5 in LUAD. The genetic alterations of DLGAP5 were assessed through cBioPortal and COSMIC databases. The relationship between DLGAP5 expression and genetic abnormalities of driver genes in LUAD was analyzed through TIMER2.0 database. CancerSEA database was utilized to explore the function of DLGAP5 in 14 different states in LUAD at single-cell resolution. GDSC database was utilized to analyze the impact of DLGAP5 on IC50 of frequently-used anti-LUAD drugs. CIBERSORT method and TIMER2.0 database was utilized to explore the relationship between DLGAP5 and tumor immune infiltration. Network pharmacology was applied to screen potential DLGAP5 inhibitor. In vitro and in vivo experiments were utilized to evaluate biological function and downstream targets of DLGAP5, and the effect of screened DLGAP5 inhibitor on LUAD growth. Results High DLGAP5 expression was commonly observed in LUAD and associated with mutation of major driver genes, poor prognosis, high IC50 values of frequently-used anti-LUAD drugs, increasing immune infiltration and elevated immune checkpoint blockade-related genes in LUAD. PLK1 was revealed as a potential DLGAP5 downstream target in LUAD. DLGAP5 overexpression or knockdown significantly promoted or inhibited LUAD cell proliferation and PLK1 expression. PLK1 overexpression well rescued DLGAP5 knockdown-induced cell proliferation inhibition, or vice versa. Furthermore, by virtual screening of an investigational drug library from the DrugBank database, AT9283 was screened and identified as a novel DLGAP5 inhibitor. AT9283 effectively suppressed growth of LUAD cells both in vitro and in vivo. DLGAP5 overexpression significantly reversed AT9283-induced proliferation inhibition. Moreover, AT9283 significantly suppressed DLGAP5 and PLK1 expression, while DLGAP5 overexpression significantly reversed AT9283-induced PLK1 suppression. Conclusion Our research has demonstrated that DLGAP5 is upregulated in LUAD and exhibits a strong correlation with unfavorable prognosis. Furthermore, DLGAP5 assumes a significant function in the regulation of tumor immunity and treatment outcome of immune checkpoint inhibitors. Of note, we found that DLGAP5 promotes cell proliferation of LUAD via upregulating PLK1. Targeting DLGAP5 by AT9283, our newly identified DLGAP5 inhibitor, suppresses LUAD growth. DLGAP5 may become a promising prognostic biomarker and therapeutic target for patients with LUAD.

Abstract Background Rho guanine nucleotide dissociation inhibitor 1 (RhoGDI1) plays an important role in diverse cellular processes by regulating Rho guanosine triphosphate (GTP)ases activity. RhoGDI1 phosphorylation regulates the spatiotemporal activation of Rho GTPases during cell migration. In this study, we identified polo-like kinase 1 (PLK1) as a novel kinase of RhoGDI1 and investigated the molecular mechanism by which the interaction between RhoGDI1 and PLK1 regulates cancer cell migration. Methods Immunoprecipitation, GST pull-down assay, and proximity ligation assay (PLA) were performed to analyze the interaction between RhoGDI1 and PLK1. In vitro kinase assay and immunoprecipitation were performed with Phospho-(Ser/Thr) antibody. We evaluated RhoA activation using RhoGTPases activity assay. Cell migration and invasion were analyzed by transwell assays. Results GST pull-down assays and PLA showed that PLK1 directly interacted with RhoGDI1 in vitro and in vivo. Truncation mutagenesis revealed that aa 90-111 of RhoGDI1 are critical for interacting with PLK1. We also showed that PLK1 phosphorylated RhoGDI1 at Thr7 and Thr91, which induces cell motility. Overexpression of the GFP-tagged RhoGDI1 truncated mutant (aa 90-111) inhibited the interaction of PLK1 with RhoGDI1 and attenuated RhoA activation by PLK1. Furthermore, the overexpression of the RhoGDI1 truncated mutant reduced cancer cell migration and invasion in vitro and suppressed lung metastasis in vivo. Conclusions Collectively, we demonstrate that the phosphorylation of RhoGDI1 by PLK1 promotes cancer cell migration and invasion through RhoA activation. This study connects the interaction between PLK1 and RhoGDI1 to the promotion of cancer cell behavior associated with malignant progression, thereby providing opportunities for cancer therapeutic interventions.

Abstract Background KRASG12C inhibitors (KRASG12Ci) AMG510 and MRTX849 have shown promising efficacy in clinical trials and been approved for the treatment of KRASG12C-mutant cancers. However, the emergence of therapy-related drug resistance limits their long-term potential. This study aimed to identify the critical mediators and develop overcoming strategies. Methods By using RNA sequencing, RT-qPCR and immunoblotting, we identified and validated the upregulation of c-Myc activity and the amplification of the long noncoding RNA ST8SIA6-AS1 in KRASG12Ci-resistant cells. The regulatory axis ST8SIA6-AS1/Polo-like kinase 1 (PLK1)/c-Myc was investigated by bioinformatics, RNA fluorescence in situ hybridization, RNA immunoprecipitation, RNA pull-down and chromatin immunoprecipitation. Gain/loss-of-function assays, cell viability assay, xenograft models, and IHC staining were conducted to evaluate the anti-cancer effects of co-inhibition of ST8SIA6-AS1/PLK1 pathway and KRAS both in vitro and in vivo. Results KRASG12Ci sustainably decreased c-Myc levels in responsive cell lines but not in cell lines with intrinsic or acquired resistance to KRASG12Ci. PLK1 activation contributed to this ERK-independent c-Myc stability, which in turn directly induced PLK1 transcription, forming a positive feedback loop and conferring resistance to KRASG12Ci. ST8SIA6-AS1 was found significantly upregulated in resistant cells and facilitated the proliferation of KRASG12C-mutant cancers. ST8SIA6-AS1 bound to Aurora kinase A (Aurora A)/PLK1 and promoted Aurora A-mediated PLK1 phosphorylation. Concurrent targeting of KRAS and ST8SIA6-AS1/PLK1 signaling suppressed both ERK-dependent and -independent c-Myc expression, synergistically led to cell death and tumor regression and overcame KRASG12Ci resistance. Conclusions Our study deciphers that the axis of ST8SIA6-AS1/PLK1/c-Myc confers both intrinsic and acquired resistance to KRASG12Ci and represents a promising therapeutic target for combination strategies with KRASG12Ci in the treatment of KRASG12C-mutant cancers.

Abstract Background More than 90% of the mortality of triple-negative breast cancer (TNBC) patients is attributed to cancer metastasis with organotropism. The lung is a frequent site of TNBC metastasis. However, the precise molecular mechanism for lung-specific metastasis of TNBC is not well understood. Methods RNA sequencing was performed to identify patterns of gene expression associated with lung metastatic behavior using 4T1-LM3, MBA-MB-231-LM3, and their parental cells (4T1-P, MBA-MB-231-P). Expressions of RGCC, called regulator of cell cycle or response gene to complement 32 protein, were detected in TNBC cells and tissues by qRT-PCR, western blotting, and immunohistochemistry. Kinase activity assay was performed to evaluate PLK1 kinase activity. The amount of phosphorylated AMP-activated protein kinase α2 (AMPKα2) was detected by immunoblotting. RGCC-mediated metabolism was determined by UHPLC system. Oxidative phosphorylation was evaluated by JC-1 staining and oxygen consumption rate (OCR) assay. Fatty acid oxidation assay was conducted to measure the status of RGCC-mediated fatty acid oxidation. NADPH and ROS levels were detected by well-established assays. The chemical sensitivity of cells was evaluated by CCK8 assay. Results RGCC is aberrantly upregulated in pulmonary metastatic cells. High level of RGCC is significantly related with lung metastasis in comparison with other organ metastases. RGCC can effectively promote kinase activity of PLK1, and the activated PLK1 phosphorylates AMPKα2 to facilitate TNBC lung metastasis. Mechanistically, the RGCC/PLK1/AMPKα2 signal axis increases oxidative phosphorylation of mitochondria to generate more energy, and promotes fatty acid oxidation to produce abundant NADPH. These metabolic changes contribute to sustaining redox homeostasis and preventing excessive accumulation of potentially detrimental ROS in metastatic tumor cells, thereby supporting TNBC cell survival and colonization during metastases. Importantly, targeting RGCC in combination with paclitaxel/carboplatin effectively suppresses pulmonary TNBC lung metastasis in a mouse model. Conclusions RGCC overexpression is significantly associated with lung-specific metastasis of TNBC. RGCC activates AMPKα2 and downstream signaling through RGCC-driven PLK1 activity to facilitate TNBC lung metastasis. The study provides implications for RGCC-driven OXPHOS and fatty acid oxidation as important therapeutic targets for TNBC treatment.

Polo-like kinase 1 (PLK1) is a serine/threonine–protein kinase essential for regulating multiple stages of cell cycle progression in mammals. Aberrant regulation of PLK1 has been observed in numerous human cancers and is linked to poor prognoses. However, its role in the pathogenesis of colorectal cancer (CRC) in the Middle East remains unexplored. PLK1 overexpression was noted in 60.3% (693/1149) of CRC cases and was significantly associated with aggressive clinico-pathological parameters and p-ERK1/2 overexpression. Intriguingly, multivariate logistic regression analysis identified PLK1 as an independent predictor of lymph node metastasis. Our in vitro experiments demonstrated that CRC cells with high PLK1 levels were resistant to 5-Fu treatment, while those with low PLK1 expression were sensitive. To investigate PLK1′s role in chemoresistance, we used the specific inhibitor volasertib, which effectively reversed 5-Fu resistance. Interestingly, forced PLK1 expression activated the CRAF-MEK-ERK signaling cascade, while its inhibition suppressed this cascade. PLK1 knockdown reduced epithelial-to-mesenchymal transition (EMT) progression and stem cell-like traits in 5-Fu-resistant cells, implicating PLK1 in EMT induction and stemness in CRC. Moreover, silencing ERK1/2 significantly mitigated chemoresistance, EMT, and stemness properties in CRC cell lines that express PLK1. Furthermore, the knockdown of Zeb1 attenuated EMT and stemness, suggesting a possible link between EMT activation and the maintenance of stemness in CRC. Our findings underscore the pivotal role of PLK1 in mediating chemoresistance and suggest that PLK1 inhibition may represent a potential therapeutic strategy for the management of aggressive colorectal cancer subtypes.

Background: Rho guanine nucleotide dissociation inhibitor 1 (RhoGDI1) plays an important role in diverse cellular processes by regulating Rho guanosine triphosphate (GTP)ases activity. RhoGDI1 phosphorylation regulates the spatiotemporal activation of Rho GTPases during cell migration. In this study, we identified polo-like kinase 1 (PLK1) as a novel kinase of RhoGDI1 and investigated the molecular mechanism by which the interaction between RhoGDI1 and PLK1 regulates cancer cell migration. Methods: Immunoprecipitation, GST pull-down assay, and proximity ligation assay (PLA) were performed to analyze the interaction between RhoGDI1 and PLK1. In vitro kinase assay and immunoprecipitation were performed with Phospho-(Ser/Thr) antibody. We evaluated RhoA activation using RhoGTPases activity assay. Cell migration and invasion were analyzed by transwell assays. Results: GST pull-down assays and PLA showed that PLK1 directly interacted with RhoGDI1 in vitro and in vivo. Truncation mutagenesis revealed that aa 90-111 of RhoGDI1 are critical for interacting with PLK1. We also showed that PLK1 phosphorylated RhoGDI1 at Thr7 and Thr91, which induces cell motility. Overexpression of the GFP-tagged RhoGDI1 truncated mutant (aa 90-111) inhibited the interaction of PLK1 with RhoGDI1 and attenuated RhoA activation by PLK1. Furthermore, the overexpression of the RhoGDI1 truncated mutant reduced cancer cell migration and invasion in vitro and suppressed lung metastasis in vivo. Conclusions: Collectively, we demonstrate that the phosphorylation of RhoGDI1 by PLK1 promotes cancer cell migration and invasion through RhoA activation. This study connects the interaction between PLK1 and RhoGDI1 to the promotion of cancer cell behavior associated with malignant progression, thereby providing opportunities for cancer therapeutic interventions. [ABSTRACT FROM AUTHOR]

Stable transmission of genetic information during cell division requires faithful chromosome segregation. Mounting evidence has demonstrated that polo-like kinase 1 (PLK1) dynamics at kinetochores control correct kinetochore–microtubule attachments and subsequent silencing of the spindle assembly checkpoint. However, the mechanisms underlying PLK1-mediated silencing of the spindle checkpoint remain elusive. Here, we identified a regulatory mechanism by which PLK1-elicited zeste white 10 (ZW10) phosphorylation regulates spindle checkpoint silencing in mitosis. ZW10 is a cognate substrate of PLK1, and the phosphorylation of ZW10 at Ser12 enables dynamic ZW10–Zwint1 interactions. Inhibition of ZW10 phosphorylation resulted in misaligned chromosomes, while persistent expression of phospho-mimicking ZW10 mutant caused premature anaphase, in which sister chromatids entangled as cells entered anaphase. These findings reveal the previously uncharacterized PLK1–ZW10 interaction through which dynamic phosphorylation of ZW10 fine-tunes accurate chromosome segregation in mitosis. [ABSTRACT FROM AUTHOR]

Background: Despite advancements in treatment approaches, patients diagnosed with aggressive breast cancer (BC) subtypes typically face an unfavorable prognosis. Globally, these cancers continue to pose a significant threat to women's health, leading to substantial morbidity and mortality. Consequently, there has been a significant struggle to identify viable molecular targets for therapeutic intervention in these patients. Pololike Kinase-1 (PLK1) represents one of these molecular targets currently undergoing rigorous scrutiny for the treatment of such tumors. Yet, its role in the pathogenesis of BC in Middle Eastern ethnicity remains unexplored. Methods: We investigated the expression of PLK1 protein in a cohort of more than 1500 Middle Eastern ethnicity BC cases by immunohistochemistry. Association with clinicopathological parameters and prognosis were performed. In vitro studies were conducted using the PLK1 inhibitor volasertib and the PARP inhibitor olaparib, either alone or in combination, in PTC cell lines. Results: Overexpression of PLK1 was detected in 27.4% of all BC cases, and this was notably correlated with aggressive clinicopathological markers. PLK1 was enriched in the triple-negative breast cancer (TNBC) subtype and exhibited poor overall survival (p = 0.0347). Notably, there was a positive correlation between PLK1 and PARP overexpression, with co-expression of PLK1 and PARP observed in 15.7% of cases and was associated with significantly poorer overall survival (OS) compared to the overexpression of either protein alone (p = 0.0050). In vitro, we studied the effect of PLK1 and PARP inhibitors either single or combined treatments in two BRCA mutated, and one BRCA proficient TNBC cell lines. We showed that combined inhibition significantly reduced cell survival and persuaded apoptosis in TNBC cell lines. Moreover, our findings indicate that inhibition of PLK1 can reinstate sensitivity in PARP inhibitor (PARPi) resistant TNBC cell lines. Conclusion: Our results shed light on the role of PLK1 in the pathogenesis and prognosis of Middle Eastern BC and support the potential clinical development of combined inhibition of PLK1 and PARP, a strategy that could potentially broaden the use of PLK1 and PARP inhibitors beyond BC cases lacking BRCA. [ABSTRACT FROM AUTHOR]

Simple Summary: Hepatocellular carcinoma (HCC) has the highest incidence and mortality rate in the Asian population. However, it is unclear how ethnic genetic variability affects this. Our research shows that hsa-miR-100-5p, a miRNA associated with HCC prognosis, improves prognosis in Asians. In contrast, Polo-like kinase 1 (PLK1), inhibited by hsa-miR-100-5p, worsens HCC prognosis in Asians. A single-nucleotide polymorphism (SNP) named rs27770, located in the 3′UTR of PLK1, has a significantly higher frequency of the G allele in the East Asian population. The A>G change in rs27770 affects the PLK1 mRNA secondary structure and alters the hsa-miR-100-5p/PLK1 interaction by creating an additional seedless binding site. This could make hsa-miR-100-5p more favorable for HCC prognosis in the Asian population. Our findings could help explain why the Asian population is more susceptible to HCC. Hepatocellular carcinoma (HCC) has the highest incidence and mortality in the Asian population, and race is an independent risk factor affecting survival time in liver cancer. Micro RNAs (miRNAs) are remarkably dysregulated in HCC and closely associated with HCC prognosis. Recent studies show that genetic variability between ethnic groups may result in differences in the specificity of HCC miRNA biomarkers. Here, we reveal a high expression level of hsa-miR-100-5p, an HCC prognosis-related miRNA, which improves HCC prognosis in the Asian Population with Polo-like kinase 1 (PLK1) variant rs27770A>G. In this study, we discovered that hsa-miR-100-5p was downregulated in various HCC cell lines. While mimics transient transfection and mouse liver cancer model confirmed the interaction between hsa-miR-100-5p and PLK1, a stratified analysis based on the Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA-LIHC) data suggest both low hsa-miR-100-5p expression level and high PLK1 expression level associated with poor HCC prognosis, especially in the Asian population. According to the 1000 Genomes Project database, the SNP rs27770 located in 3′UTR of PLK1 had a significantly higher G allele frequency in the East Asian population. Bioinformatics analysis suggested that rs27770 A>G affects PLK1 mRNA secondary structure and alters the hsa-miR-100-5p/PLK1 interaction by forming an additional seedless binding site. This racial variation caused PLK1 to be more vulnerable to hsa-miR-100-5p inhibition, resulting in hsa-miR-100-5p being more favorable for HCC prognosis in the Asian population. [ABSTRACT FROM AUTHOR]

The development of castration‐resistant prostate cancer (CRPC) is a significant factor that reduces life expectancy among patients with prostate cancer. Previously, it is reported that CDK4/6 inhibitors can overcome the resistance of CRPC to BET inhibitors by destabilizing BRD4, suggesting that the combination of CDK4/6 inhibitors and BET inhibitors is a promising approach for treating CRPC. In this study, candidates that affect the combined antitumor effect of CDK4/6 inhibitors and BET inhibitors on CRPC is aimed to examine. The data demonstrates that CBX3 is abnormally upregulated in CDK4/6 inhibitors‐resistant cells. CBX3 is almost positively correlated with the cell cycle in multiple malignancies and is downregulated by BET inhibitors. Mechanistically, it is showed that CBX3 is transcriptionally upregulated by BRD4 in CRPC cells. Moreover, it is demonstrated that CBX3 modulated the sensitivity of CRPC to CDK4/6 inhibitors by binding with RB1 to release E2F1. Furthermore, it is revealed that PLK1 phosphorylated CBX3 to enhance the interaction between RB1 and CBX3, and desensitize CRPC cells to CDK4/6 inhibitors. Given that BRD4 regulates CBX3 expression and PLK1 affects the binding between RB1 and CBX3, it is proposed that a dual BRD4/PLK1 inhibitor can increase the sensitivity of CRPC cells to CDK4/6 inhibitors partially through CBX3. [ABSTRACT FROM AUTHOR]

Background: KRASG12C inhibitors (KRASG12Ci) AMG510 and MRTX849 have shown promising efficacy in clinical trials and been approved for the treatment of KRASG12C-mutant cancers. However, the emergence of therapy-related drug resistance limits their long-term potential. This study aimed to identify the critical mediators and develop overcoming strategies. Methods: By using RNA sequencing, RT-qPCR and immunoblotting, we identified and validated the upregulation of c-Myc activity and the amplification of the long noncoding RNA ST8SIA6-AS1 in KRASG12Ci-resistant cells. The regulatory axis ST8SIA6-AS1/Polo-like kinase 1 (PLK1)/c-Myc was investigated by bioinformatics, RNA fluorescence in situ hybridization, RNA immunoprecipitation, RNA pull-down and chromatin immunoprecipitation. Gain/loss-of-function assays, cell viability assay, xenograft models, and IHC staining were conducted to evaluate the anti-cancer effects of co-inhibition of ST8SIA6-AS1/PLK1 pathway and KRAS both in vitro and in vivo. Results: KRASG12Ci sustainably decreased c-Myc levels in responsive cell lines but not in cell lines with intrinsic or acquired resistance to KRASG12Ci. PLK1 activation contributed to this ERK-independent c-Myc stability, which in turn directly induced PLK1 transcription, forming a positive feedback loop and conferring resistance to KRASG12Ci. ST8SIA6-AS1 was found significantly upregulated in resistant cells and facilitated the proliferation of KRASG12C-mutant cancers. ST8SIA6-AS1 bound to Aurora kinase A (Aurora A)/PLK1 and promoted Aurora A-mediated PLK1 phosphorylation. Concurrent targeting of KRAS and ST8SIA6-AS1/PLK1 signaling suppressed both ERK-dependent and -independent c-Myc expression, synergistically led to cell death and tumor regression and overcame KRASG12Ci resistance. Conclusions: Our study deciphers that the axis of ST8SIA6-AS1/PLK1/c-Myc confers both intrinsic and acquired resistance to KRASG12Ci and represents a promising therapeutic target for combination strategies with KRASG12Ci in the treatment of KRASG12C-mutant cancers. [ABSTRACT FROM AUTHOR]

Background: More than 90% of the mortality of triple-negative breast cancer (TNBC) patients is attributed to cancer metastasis with organotropism. The lung is a frequent site of TNBC metastasis. However, the precise molecular mechanism for lung-specific metastasis of TNBC is not well understood. Methods: RNA sequencing was performed to identify patterns of gene expression associated with lung metastatic behavior using 4T1-LM3, MBA-MB-231-LM3, and their parental cells (4T1-P, MBA-MB-231-P). Expressions of RGCC, called regulator of cell cycle or response gene to complement 32 protein, were detected in TNBC cells and tissues by qRT-PCR, western blotting, and immunohistochemistry. Kinase activity assay was performed to evaluate PLK1 kinase activity. The amount of phosphorylated AMP-activated protein kinase α2 (AMPKα2) was detected by immunoblotting. RGCC-mediated metabolism was determined by UHPLC system. Oxidative phosphorylation was evaluated by JC-1 staining and oxygen consumption rate (OCR) assay. Fatty acid oxidation assay was conducted to measure the status of RGCC-mediated fatty acid oxidation. NADPH and ROS levels were detected by well-established assays. The chemical sensitivity of cells was evaluated by CCK8 assay. Results: RGCC is aberrantly upregulated in pulmonary metastatic cells. High level of RGCC is significantly related with lung metastasis in comparison with other organ metastases. RGCC can effectively promote kinase activity of PLK1, and the activated PLK1 phosphorylates AMPKα2 to facilitate TNBC lung metastasis. Mechanistically, the RGCC/PLK1/AMPKα2 signal axis increases oxidative phosphorylation of mitochondria to generate more energy, and promotes fatty acid oxidation to produce abundant NADPH. These metabolic changes contribute to sustaining redox homeostasis and preventing excessive accumulation of potentially detrimental ROS in metastatic tumor cells, thereby supporting TNBC cell survival and colonization during metastases. Importantly, targeting RGCC in combination with paclitaxel/carboplatin effectively suppresses pulmonary TNBC lung metastasis in a mouse model. Conclusions: RGCC overexpression is significantly associated with lung-specific metastasis of TNBC. RGCC activates AMPKα2 and downstream signaling through RGCC-driven PLK1 activity to facilitate TNBC lung metastasis. The study provides implications for RGCC-driven OXPHOS and fatty acid oxidation as important therapeutic targets for TNBC treatment. [ABSTRACT FROM AUTHOR]

53BP1 acts at the crossroads between DNA repair and p53‐mediated stress response. With its interactors p53 and USP28, it is part of the mitotic surveillance (or mitotic stopwatch) pathway (MSP), a sensor that monitors the duration of cell division, promoting p53‐dependent cell cycle arrest when a critical time threshold is surpassed. Here, we show that Polo‐like kinase 1 (PLK1) activity is essential for the time‐dependent release of 53BP1 from kinetochores. PLK1 inhibition, which leads to 53BP1 persistence at kinetochores, prevents cytosolic 53BP1 association with p53 and results in a blunted MSP. Strikingly, the identification of CENP‐F as the kinetochore docking partner of 53BP1 enabled us to show that measurement of mitotic timing by the MSP does not take place at kinetochores, as perturbing CENP‐F‐53BP1 binding had no measurable impact on the MSP. Taken together, we propose that PLK1 supports the MSP by generating a cytosolic pool of 53BP1 and that an unknown cytosolic mechanism enables the measurement of mitotic duration. Synopsis: Following an abnormally extended mitosis, cells activate p53 via 53BP1. Surprisingly, 53BP1 kinetochore localization is dispensable for this process, whereas PLK1 activity, a negative regulator of 53BP1 localization, appears to be essential.53BP1 kinetochore localization is dispensable for the mitotic surveillance (or mitotic stopwatch) pathway (MSP).PLK1 is required for the MSP.PLK1 activity promotes 53BP1 kinetochore dynamics and 53BP1‐p53 complex formation. [ABSTRACT FROM AUTHOR]

Abstract Background 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. Methods 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. Results 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. Conclusions 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. Key points 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.

Abstract Background Understanding the mechanism behind immune cell plasticity in cancer metastasis is crucial for identifying key regulators. Previously we found that mitotic factors regulate epithelial-mesenchymal transition, but how these factors convert to metastatic players in the tumor microenvironment (TME) is not fully understood. Methods The clinical importance of mitotic factors was analyzed by heatmap analysis, a KM plot, and immunohistochemistry in lung adenocarcinoma (LUAD) patients. Immunoprecipitation, LC–MS/MS, kinase assay, and site-directed mutagenesis were performed for the interaction and phosphorylation. A tail-vein injection mouse model, Transwell-based 3D culture, microarray analysis, coculture with monocytes, and chromatin immunoprecipitation assays were used to elucidate the function of phosphorylated FoxM1 in metastasis of TME. Results The phosphorylated FoxM1 at Ser25 by PLK1 acquires the reprogramming ability to stimulate the invasive traits in cancer and influence immune cell plasticity. This invasive form of p-FoxM1 upregulates the expression of IL1A/1B, VEGFA, and IL6 by direct activation, recruiting monocytes and promoting the polarization of M2d-like tumor-associated macrophages (TAMs). Upregulation of PD-L1 in LUAD having phosphomimetic FoxM1 facilitates immune evasion. In invasive LUAD with phosphomimetic FoxM1, IFITM1 is the most highly expressed through the activation of the STING-TBK1-IRF3 signaling, which enhances FoxM1-mediated signaling. Clinically, higher expression of FOXM1, PLK1, and IFITM1 is inversely correlated with the survival rate of advanced LUAD patients, providing a promising therapeutic strategy for the treatment of LUAD. Conclusion FoxM1-based therapy would be a potential therapeutic strategy for LUAD to reduce TAM polarization, immune escape, and metastasis, since FoxM1 functions as a genetic reprogramming factor reinforcing LUAD malignancy in the TME.

HuaChanSu is active water extracts from the skin of Bufo bufo gargarizans Cantor. It has been already used to treat clinical cancers including HCC (Hepatocellular carcinoma, HCC), however, the molecular mechanisms under HuaChanSu's anti-cancer effects remain unclear. PPP (Pentose phosphate pathway, PPP), the major source of ribose and NADPH (Nicotinamide adenine dinucleotide phosphate, NADPH), is always over-activated and particularly critical for tumor cells growth. In this study, firstly, we illustrate that HuaChanSu restrains the growth of human hepatoma cells. More importantly, we demonstrate that the expression of G6PD (Glucose-6-phosphate dehydrogenase, G6PD), the first rate-limiting enzyme of the PPP, is restrained in human hepatoma cells after treatment with HuaChanSu. Additionally, our results show that G6PD enzyme activity and dimer formation are inhibited by HuaChanSu. Furthermore, we find that HuaChanSu could inhibit NADPH production and nucleotide level. In addition, we identify that expression of PLK1 (Polo-like kinase 1, PLK1) is also reduced in response to HuaChanSu, and knockdown of PLK1 restrains enzyme activity and dimer formation of G6PD, but has no effect on G6PD protein level. Subsequently, we demonstrate that inhibition of G6PD could restrain the proliferation of tumor cells and enhance the inhibitory effect of HuaChanSu on cell proliferation of human hepatoma cells. In conclusion, for the first time, our study reveals that HuaChanSu interferes with PPP via suppression of G6PD expression and enzyme activity to restrain growth of tumor cells, and these results provide a novel insight for the anti-hepatoma mechanisms of HuaChanSu and promote the innovation of the research model of TCM. Moreover, the development of drugs targeting abnormal tumor metabolism is currently a hot topic, our works provide theoretical support for further drug development from HuaChanSu, meanwhile, the revelation of the new molecular mechanism also provides a new perspective for the study of the pathogenesis of liver cancer. Short abstract: HuaChanSu suppresses expression of G6PD, the first rate-limiting enzyme of the PPP, restrains G6PD enzyme activity and dimer formation via inhibition of PLK1, knockdown of G6PD could impair the growth of human hepatoma cells and increase the blocking effect of HuaChanSu on cell proliferation of cancer cells. In addition, HuaChanSu restrains NADPH production and nucleotide level, implying the suppression of PPP flux. Our study suggests that HuaChanSu interferes with PPP via G6PD inhibition to exert anti-hepatoma effects.

BackgroundDespite advancements in treatment approaches, patients diagnosed with aggressive breast cancer (BC) subtypes typically face an unfavorable prognosis. Globally, these cancers continue to pose a significant threat to women's health, leading to substantial morbidity and mortality. Consequently, there has been a significant struggle to identify viable molecular targets for therapeutic intervention in these patients. Polo-like Kinase-1 (PLK1) represents one of these molecular targets currently undergoing rigorous scrutiny for the treatment of such tumors. Yet, its role in the pathogenesis of BC in Middle Eastern ethnicity remains unexplored.MethodsWe investigated the expression of PLK1 protein in a cohort of more than 1500 Middle Eastern ethnicity BC cases by immunohistochemistry. Association with clinicopathological parameters and prognosis were performed. In vitro studies were conducted using the PLK1 inhibitor volasertib and the PARP inhibitor olaparib, either alone or in combination, in PTC cell lines. ResultsOverexpression of PLK1 was detected in 27.4% of all BC cases, and this was notably correlated with aggressive clinicopathological markers. PLK1 was enriched in the triple-negative breast cancer (TNBC) subtype and exhibited poor overall survival (p = 0.0347). Notably, there was a positive correlation between PLK1 and PARP overexpression, with co-expression of PLK1 and PARP observed in 15.7% of cases and was associated with significantly poorer overall survival (OS) compared to the overexpression of either protein alone (p = 0.0050). In vitro, we studied the effect of PLK1 and PARP inhibitors either single or combined treatments in two BRCA mutated, and one BRCA proficient TNBC cell lines. We showed that combined inhibition significantly reduced cell survival and persuaded apoptosis in TNBC cell lines. Moreover, our findings indicate that inhibition of PLK1 can reinstate sensitivity in PARP inhibitor (PARPi) resistant TNBC cell lines. ConclusionOur results shed light on the role of PLK1 in the pathogenesis and prognosis of Middle Eastern BC and support the potential clinical development of combined inhibition of PLK1 and PARP, a strategy that could potentially broaden the use of PLK1 and PARP inhibitors beyond BC cases lacking BRCA.

Abstract The development of castration‐resistant prostate cancer (CRPC) is a significant factor that reduces life expectancy among patients with prostate cancer. Previously, it is reported that CDK4/6 inhibitors can overcome the resistance of CRPC to BET inhibitors by destabilizing BRD4, suggesting that the combination of CDK4/6 inhibitors and BET inhibitors is a promising approach for treating CRPC. In this study, candidates that affect the combined antitumor effect of CDK4/6 inhibitors and BET inhibitors on CRPC is aimed to examine. The data demonstrates that CBX3 is abnormally upregulated in CDK4/6 inhibitors‐resistant cells. CBX3 is almost positively correlated with the cell cycle in multiple malignancies and is downregulated by BET inhibitors. Mechanistically, it is showed that CBX3 is transcriptionally upregulated by BRD4 in CRPC cells. Moreover, it is demonstrated that CBX3 modulated the sensitivity of CRPC to CDK4/6 inhibitors by binding with RB1 to release E2F1. Furthermore, it is revealed that PLK1 phosphorylated CBX3 to enhance the interaction between RB1 and CBX3, and desensitize CRPC cells to CDK4/6 inhibitors. Given that BRD4 regulates CBX3 expression and PLK1 affects the binding between RB1 and CBX3, it is proposed that a dual BRD4/PLK1 inhibitor can increase the sensitivity of CRPC cells to CDK4/6 inhibitors partially through CBX3.

ABSTRACTTank-binding kinase 1 (TBK1) plays a pivotal role as a cellular factor in regulating the immune response against herpes simplex virus type 1 (HSV-1). TBK1’s role in defending against HSV-1 is attributed to its capacity to induce the type I interferon response. Recent discoveries emphasize TBK1’s involvement in the regulation of optineurin (OPTN), an autophagy adaptor protein that plays a direct role in enhancing the antiviral response against HSV-1 through selective autophagy. Intriguingly, OPTN can also reciprocally regulate TBK1, leading to a limited understanding of the precise role of the OPTN-TBK1 axis in HSV-1 infection. In our study, we shed light on the relationship between OPTN and TBK1 in influencing the outcome of HSV-1 infection. Surprisingly, when TBK1 or OPTN was absent in cells, the spread and infectivity of HSV-1 did not align as expected. Furthermore, it was unexpected to note that cells lacking OPTN showed higher interferon expression compared to normal cells, yet they exhibited enhanced viral growth. Our investigations revealed that OPTN can directly degrade HSV-1 through autophagy even in the absence of active TBK1. Interestingly, we identified that another protein, Polo-like kinase 1 (PLK1), can compensate for the absence of TBK1 and trigger autophagy to restrict HSV-1. In summary, our findings highlight that OPTN significantly impacts the outcome of HSV-1 infection, regardless of the conventional antiviral response mediated by TBK1. Additionally, by introducing a new role for PLK1, this research provides valuable insights into the intricate interplay among OPTN, TBK1, and autophagy in shaping the course of HSV-1 infection.IMPORTANCEHerpes simplex virus type 1 (HSV-1) is globally prevalent, with latent infections observed in up to 80% of the population. The virus is known for subverting host defense mechanisms and infiltrating the nervous system to establish latency in peripheral ganglia. Multiple stressors can reactivate the virus, and recurrent herpes has been linked to vision loss and neurodegeneration. Identifying critical host factors that limit the spread of HSV-1 and the subsequent establishment of latent infection holds the potential to drive new intervention strategies for eradicating the virus. Numerous pieces of evidence underscore the significance of Tank-binding kinase 1 (TBK1) in restricting HSV-1. Reports have also suggested that phosphorylation of optineurin (OPTN) by TBK1 is required for triggering OPTN-mediated autophagy for HSV degradation. This report adds new insights into the roles of OPTN and TBK1 in HSV-1 infection and provides proof of a TBK1-independent HSV-1 restriction through OPTN. It confirms that TBK1 activation can be substituted by PLK1 to provide protection against HSV-1. In contrast, the activation of OPTN is likely an indispensable host defense mechanism for optimal defense against HSV-1.

Background: Esophageal squamous cell carcinoma (ESCC) is the most common pathological type of esophageal cancer in China, accounting for more than 90 %. Most patients were diagnosed with advanced-stage ESCC, for whom new adjuvant therapy is recommended. Therefore, it is urgent to explore new therapeutic targets for ESCC. Ferroptosis, a newly discovered iron-dependent programmed cell death, has been shown to play an important role in carcinogenesis by many studies. This study explored the effect of Polo like kinase 1 (PLK1) on chemoradiotherapy sensitivity of ESCC through ferroptosis Methods: In this study, we knocked out the expression of PLK1 (PLK1-KO) in ESCC cell lines (KYSE150 and ECA109) with CRISPR/CAS9. The effects of PLK1-knock out on G6PD, the rate-limiting enzyme of pentose phosphate pathway (PPP), and downstream NADPH and GSH were explored. The lipid peroxidation was observed by flow cytometry, and the changes in mitochondria were observed by transmission electron microscopy. Next, through the CCK-8 assay and clone formation assay, the sensitivity to cobalt 60 rays, paclitaxel, and cisplatin were assessed after PLK1-knock out, and the nude mouse tumorigenesis experiment further verified it. The regulation of transcription factor YY1 on PLK1 was evaluated by dual luciferase reporter assay. The expression and correlation of PLK1 and YY1, and their impact on prognosis were analyzed in more than 300 ESCC cases from the GEO database and our center. Finally, the above results were further proved by single-cell sequencing. Results: After PLK1 knockout, the expression of G6PD dimer and the level of NADPH and GSH in KYSE150 and ECA109 cells significantly decreased. Accordingly, lipid peroxidation increased, mitochondria became smaller, membrane density increased, and ferroptosis was more likely to occur. However, with the stimulation of exogenous GSH (10 mM), there was no significant difference in lipid peroxidation and ferroptosis between the PLK1-KO group and the control group. After ionizing radiation, the PLK1-KO group had higher lipid peroxidation ratio, more cell death, and was more sensitive to radiation, while exogenous GSH (10 mM) could eliminate this difference. Similar results could also be observed when receiving paclitaxel combined with cisplatin and chemoradiotherapy. The expression of PLK1, G6PD dimer, and the level of NADPH and GSH in KYSE150, ECA109, and 293 T cells stably transfected with YY1-shRNAs significantly decreased, and the cells were more sensitive to radiotherapy and chemotherapy. ESCC patients from the GEO database and our center, YY1 and PLK1 expression were significantly positively-correlated, and the survival of patients with high expression of PLK1 was significantly shorter. Further analysis of single-cell sequencing specimens of ESCC in our center confirmed the above results. Conclusion: In ESCC, down-regulation of PLK1 can inhibit PPP, and reduce the level of NADPH and GSH, thereby promoting ferroptosis and improving their sensitivity to radiotherapy and chemotherapy. Transcription factor YY1 has a positive regulatory effect on PLK1, and their expressions were positively correlated. PLK1 may be a target for predicting and enhancing the chemoradiotherapy sensitivity of ESCC.

Background: Understanding the mechanism behind immune cell plasticity in cancer metastasis is crucial for identifying key regulators. Previously we found that mitotic factors regulate epithelial-mesenchymal transition, but how these factors convert to metastatic players in the tumor microenvironment (TME) is not fully understood. Methods: The clinical importance of mitotic factors was analyzed by heatmap analysis, a KM plot, and immunohistochemistry in lung adenocarcinoma (LUAD) patients. Immunoprecipitation, LC–MS/MS, kinase assay, and site-directed mutagenesis were performed for the interaction and phosphorylation. A tail-vein injection mouse model, Transwell-based 3D culture, microarray analysis, coculture with monocytes, and chromatin immunoprecipitation assays were used to elucidate the function of phosphorylated FoxM1 in metastasis of TME. Results: The phosphorylated FoxM1 at Ser25 by PLK1 acquires the reprogramming ability to stimulate the invasive traits in cancer and influence immune cell plasticity. This invasive form of p-FoxM1 upregulates the expression of IL1A/1B, VEGFA, and IL6 by direct activation, recruiting monocytes and promoting the polarization of M2d-like tumor-associated macrophages (TAMs). Upregulation of PD-L1 in LUAD having phosphomimetic FoxM1 facilitates immune evasion. In invasive LUAD with phosphomimetic FoxM1, IFITM1 is the most highly expressed through the activation of the STING-TBK1-IRF3 signaling, which enhances FoxM1-mediated signaling. Clinically, higher expression of FOXM1, PLK1, and IFITM1 is inversely correlated with the survival rate of advanced LUAD patients, providing a promising therapeutic strategy for the treatment of LUAD. Conclusion: FoxM1-based therapy would be a potential therapeutic strategy for LUAD to reduce TAM polarization, immune escape, and metastasis, since FoxM1 functions as a genetic reprogramming factor reinforcing LUAD malignancy in the TME. [ABSTRACT FROM AUTHOR]

PLK1 is overexpressed in acute myeloid leukemia (AML). A phase 1b trial of the PLK1 inhibitor onvansertib (ONV) combined with decitabine (DAC) demonstrated initial safety and efficacy in patients with relapsed/refractory (R/R) AML. The current study aimed to identify molecular predictors of response to ONV + DAC in R/R AML patients. A total of 44 R/R AML patients were treated with ONV + DAC and considered evaluable for efficacy. Bone marrow (BM) samples were collected at baseline for genomic and transcriptomic analysis (n = 32). A 10-gene expression signature, predictive of response to ONV + DAC, was derived from the leading-edge genes of gene set enrichment analyses (GSEA). The gene signature was evaluated in independent datasets and used to identify associated mutated genes. Twenty percent of the patients achieved complete remission, with or without hematologic count recovery (CR/CRi), and 32% exhibited a ≥50% reduction in bone marrow blasts. Patients who responded to treatment had elevated mitochondrial function and OXPHOS. The gene signature was not associated with response to DAC alone in an independent dataset. By applying the signature to the BeatAML cohort (n = 399), we identified a positive association between predicted ONV + DAC response and mutations in splicing factors (SF). In the phase 1b/2 trial, patients with SF mutations (SRSF2, SF3B1) had a higher CR/CRi rate (50%) compared to those without SF mutations (9%). PLK1 inhibition with ONV in combination with DAC could be a potential therapy in R/R AML patients, particularly those with high OXPHOS gene expression and SF mutations. [ABSTRACT FROM AUTHOR]

Two major mechanisms safeguard genome stability during mitosis: the mitotic checkpoint delays mitosis until all chromosomes have attached to microtubules, and the kinetochore–microtubule error‐correction pathway keeps this attachment process free from errors. We demonstrate here that the optimal strength and dynamics of these processes are set by a kinase–phosphatase pair (PLK1‐PP2A) that engage in negative feedback from adjacent phospho‐binding motifs on the BUB complex. Uncoupling this feedback to skew the balance towards PLK1 produces a strong checkpoint, hypostable microtubule attachments and mitotic delays. Conversely, skewing the balance towards PP2A causes a weak checkpoint, hyperstable microtubule attachments and chromosome segregation errors. These phenotypes are associated with altered BUB complex recruitment to KNL1‐MELT motifs, implicating PLK1‐PP2A in controlling auto‐amplification of MELT phosphorylation. In support, KNL1‐BUB disassembly becomes contingent on PLK1 inhibition when KNL1 is engineered to contain excess MELT motifs. This elevates BUB‐PLK1/PP2A complex levels on metaphase kinetochores, stabilises kinetochore–microtubule attachments, induces chromosome segregation defects and prevents KNL1‐BUB disassembly at anaphase. Together, these data demonstrate how a bifunctional PLK1/PP2A module has evolved together with the MELT motifs to optimise BUB complex dynamics and ensure accurate chromosome segregation. Synopsis: The mitotic checkpoint and kinetochore–microtubule error‐correction pathways safeguard chromosome segregation during mitosis. This work shows that a kinase–phosphatase recruitment module in BUBR1 integrates both processes by balancing PLK1 and PP2A levels at kinetochore scaffold KNL1. The kinase PLK1 and phosphatase PP2A‐B56 are engaged in an intramolecular negative feedback loop on BUBR1, which binds to kinetochores via phosphorylated MELT motifs in KNL1.This feedback balances PLK1 and PP2A levels at KNL1 to optimise strength and dynamics of the mitotic checkpoint (PLK1) and kinetochore–microtubule attachments (PP2A).The number of MELT motifs in KNL1 is also crucial for setting the right PLK1/PP2A levels because elevating MELT number causes defects that are associated with high PLK1/PP2A activity. [ABSTRACT FROM AUTHOR]

Lung cancer is still the most common cancer in the world, especially lung adenocarcinoma (LUAD). Despite years of effort, including the application of immunotherapy and targeted therapy, the survival rate of LUAD has not improved significantly. Exploring effective targets and combination drugs is crucial for the treatment of LUAD. We characterized differentially expressed genes between LUAD and normal lung tissue based on The Cancer Genome Atlas (TCGA) database and identified polo-like kinase 1 (PLK1) as the hub gene. Through an analysis using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), we obtained a combination of Chinese medicine with PLK1 inhibitor, whose biological function we confirmed by western blot and TdT-UTP nick-end labelling (TUNEL) assays. After combined analysis of protein expression with clinical characteristics, GNPNAT1, CCT6A, SMOX, UCK2, PLK1, HMMR and ANLN expression were significantly correlated with age, sex and stage. Among them, the survival rate was lower in patients with high PLK1 expression than in those with low PLK1 expression, making PLK1 a promising therapeutic target for LUAD. Stage and PLK1 expression could be used as independent prognostic factors for LUAD. By TCMSP analysis, tectoridin had the strongest correlation with PLK1. Tectoridin synergized with PLK1 inhibitor to suppress autophagy and ferroptosis but promoted caspase-3-mediated apoptosis in A549 cells. Our findings highlight a potential drug target and the combination therapy strategy of PLK1 inhibitor and tectoridin for LUAD patients. [ABSTRACT FROM AUTHOR]

Polo-Like Kinase 1 (PLK1), a key mediator of cell-cycle progression, is associated with poor prognosis and is a therapeutic target in a number of malignancies. Putative phosphorylation sites for PLK1 have been identified on Drosha, the main catalytic component of the microprocessor responsible for miR biogenesis. Several kinases, including GSK3β, p70 S6 kinase, ABL, PAK5, p38 MAPK, CSNK1A1 and ANKRD52-PPP6C, have been shown to phosphorylate components of the miR biogenesis machinery, altering their activity and/or localisation, and therefore the biogenesis of distinct miR subsets. We hypothesised that PLK1 regulates miR biogenesis through Drosha phosphorylation. In vitro kinase assays confirmed PLK1 phosphorylation of Drosha at S300 and/or S302. PLK1 inhibition reduced serine-phosphorylated levels of Drosha and its RNA-dependent association with DGCR8. In contrast, a "phospho-mimic" Drosha mutant showed increased association with DGCR8. PLK1 phosphorylation of Drosha alters Drosha Microprocessor complex subcellular localisation, since PLK1 inhibition increased cytosolic protein levels of both DGCR8 and Drosha, whilst nuclear levels were decreased. Importantly, the above effects are independent of PLK1's cell cycle-regulatory role, since altered Drosha:DGCR8 localisation upon PLK1 inhibition occurred prior to significant accumulation of cells in M-phase, and PLK1-regulated miRs were not increased in M-phase-arrested cells. Small RNA sequencing and qPCR validation were used to assess downstream consequences of PLK1 activity on miR biogenesis, identifying a set of ten miRs (miR-1248, miR-1306-5p, miR-2277-5p, miR-29c-5p, miR-93-3p, miR-152-3p, miR-509-3-5p, miR-511-5p, miR-891a-5p and miR-892a) whose expression levels were statistically significantly downregulated by two pharmacological PLK1 kinase domain inhibitors, RO-5203280 and GSK461364. Opposingly, increased levels of these miRs were observed upon transfection of wild-type or constitutively active PLK1. Importantly, pre-miR levels were reduced upon PLK1 inhibition, and pri-miR levels decreased upon PLK1 activation, and hence, PLK1 Drosha phosphorylation regulates MiR biogenesis at the level of pri-miR-to-pre-miR processing. In combination with prior studies, this work identifies Drosha S300 and S302 as major integration points for signalling by several kinases, whose relative activities will determine the relative biogenesis efficiency of different miR subsets. Identified kinase-regulated miRs have potential for use as kinase inhibitor response-predictive biomarkers, in cancer and other diseases. [ABSTRACT FROM AUTHOR]

Aim: Polo‐like kinase‐1 (PLK1) plays a crucial role in cell cycle progression, and it is considered a potential therapeutic target in many cancers. Although the role of PLK1 is well established in triple‐negative breast cancer (TNBC) as an oncogene, its role in luminal BC is still controversial. In this study, we aimed to evaluate the prognostic and predictive role of PLK1 in BC and its molecular subtypes. Methods: A large BC cohort (n = 1208) were immunohistochemically stained for PLK1. The association with clinicopathological, molecular subtypes, and survival data was analysed. PLK1 mRNA was evaluated in the publicly available datasets (n = 6774), including The Cancer Genome Atlas and the Kaplan–Meier Plotter tool. Results: 20% of the study cohort showed high cytoplasmic PLK1 expression. High PLK1 expression was significantly associated with a better outcome in the whole cohort, luminal BC. In contrast, high PLK1 expression was associated with a poor outcome in TNBC. Multivariate analyses indicated that high PLK1 expression is independently associated with longer survival in luminal BC, and in poorer prognosis in TNBC. At the mRNA levels, PLK1 expression was associated with short survival in TNBC consistent with the protein expression. However, in luminal BC, its prognostic value significantly varies between cohorts. Conclusion: The prognostic role of PLK1 in BC is molecular subtype‐dependent. As PLK1 inhibitors are introduced to clinical trials for several cancer types, our study supports evaluation of the pharmacological inhibition of PLK1 as an attractive therapeutic target in TNBC. However, in luminal BC, PLK1 prognostic role remains controversial. [ABSTRACT FROM AUTHOR]