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BACKGROUND AND PURPOSE: Prostate cancer remains a major public health burden worldwide. Polo like kinase 4 (PLK4) has emerged as a promising therapeutic target in prostate cancer due to its key roles in cell cycle regulation and tumour progression. This study aims to develop and characterize the novel curcumin analogue NL13 as a potential therapeutic agent and PLK4 inhibitor against prostate cancer. EXPERIMENTAL APPROACH: NL13 was synthesized and its effects were evaluated in prostate cancer cells and mouse xenograft models. Kinome screening and molecular modelling identified PLK4 as the primary target. Antiproliferative and proapoptotic mechanisms were explored via cell cycle, apoptosis, gene and protein analyses. KEY RESULTS: Compared with curcumin, NL13 exhibited much greater potency in inhibiting PC3 (IC50, 3.51 µM vs. 35.45 µM) and DU145 (IC50, 2.53 µM vs. 29.35 µM) prostate cancer cells viability and PLK4 kinase activity (2.32 µM vs. 246.88 µM). NL13 induced G2/M cell cycle arrest through CCNB1/CDK1 down-regulation and triggered apoptosis via caspase-9/caspase-3 cleavage. These effects were mediated by PLK4 inhibition, which led to the inactivation of the AKT signalling pathway. In mice, NL13 significantly inhibited tumour growth and modulated molecular markers consistent with in vitro findings, including decreased p-AKT and increased cleaved caspase-9/3. CONCLUSION AND IMPLICATIONS: NL13, a novel PLK4-targeted curcumin analogue, exerts promising anticancer properties against prostate cancer by disrupting the PLK4-AKT-CCNB1/CDK1 and apoptosis pathways. NL13 represents a promising new agent for prostate cancer therapy.

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CRISPR/Cas9 genome editing is a pervasive research tool due to its relative ease of use. However, some systems are not amenable to generating edited clones due to genomic complexity and/or difficulty in establishing clonal lines. For example, Drosophila Schneider 2 (S2) cells possess a segmental aneuploid genome and are challenging to single-cell select. Here, we describe a streamlined CRISPR/Cas9 methodology for knock-in and knock-out experiments in S2 cells, whereby an antibiotic resistance gene is inserted in-frame with the coding region of a gene-of-interest. By using selectable markers, we have improved the ease and efficiency for the positive selection of null cells using antibiotic selection in feeder layers followed by cell expansion to generate clonal lines. Using this method, we generated the first acentrosomal S2 cell lines by knocking-out centriole genes Polo-like Kinase 4/Plk4 or Ana2 as proof of concept. These strategies for generating gene-edited clonal lines will add to the collection of CRISPR tools available for cultured Drosophila cells by making CRISPR more practical and therefore improving gene function studies.

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Objective: This study aimed to assess the value of PLK4 as a biomarker in papillary thyroid carcinoma (PTC). Methods: This study reviewed 230 PTC patients receiving surgical resections. PLK4 was detected in tumor tissues and samples of normal thyroid gland tissues by immunohistochemistry. Results: PLK4 was elevated in tumor tissues versus normal thyroid gland tissues (p < 0.001). Tumor PLK4 was linked with extrathyroidal invasion (p = 0.036), higher pathological tumor stage (p = 0.030), node stage (p = 0.045) and tumor/node/metastasis stage (p = 0.022) in PTC patients. Tumor PLK4 immunohistochemistry score >3 was linked with shortened disease-free survival (p = 0.026) and overall survival (p = 0.028) and independently predicted poorer disease-free survival (hazard ratio: 2.797; p = 0.040). Conclusion: Tumor PLK4 reflects extrathyroidal invasion, higher tumor stage and shortened survival in PTC.

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BACKGROUND: Polo-like kinase 4 (PLK4) serves as a marker for tumor features and poor outcomes in cancers. This study aimed to explore the associations of tumor PLK4 protein expression with tumor characteristics and survival in endometrial cancer (EC) patients who underwent surgical resection. METHODS: This study included 142 EC patients who underwent surgical resection. Tumor tissue samples were obtained for tumor PLK4 protein expression detection via immunohistochemistry (IHC). RESULTS: Among EC patients, 26.1% had a PLK4 IHC score of 0, 24.6% had a score of 1-3, 27.5% had a score of 4-6, and 21.8% had a score of 7-12. Tumor PLK4 protein expression positively associated with lymphovascular invasion (P = 0.008) and Federation of Gynecology and Obstetrics (FIGO) stage (P = 0.005). Disease-free survival (DFS) was not different between patients with tumor PLK4 IHC scores > 0 and ≤ 0 (P = 0.154) but was reduced in patients with scores > 3 vs. ≤ 3 (P = 0.009) and > 6 vs. ≤ 6 (P < 0.001). Similarly, overall survival (OS) was not different between patients with scores > 0 and ≤ 0 (P = 0.322) but was shorter in patients with scores > 3 vs. ≤ 3 (P = 0.011) and > 6 vs. ≤ 6 (P = 0.006). After adjustment, a tumor PLK4 IHC score > 6 (vs. ≤ 6) (hazard ratio (HR): 3.156, P = 0.008) or > 3 (vs. ≤ 3) (HR: 3.918, P = 0.026) was independently associated with shortened DFS and OS. CONCLUSION: A tumor PLK4 IHC score > 6 or > 3 associates with shortened DFS and OS in EC patients who undergo surgical resection.

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Polo-like kinase 4 (PLK4), a highly conserved serine/threonine kinase, masterfully regulates centriole duplication in a spatiotemporal manner to ensure the fidelity of centrosome duplication and proper mitosis. Abnormal expression of PLK4 contributes to genomic instability and associates with a poor prognosis in cancer. Inhibition of PLK4 is demonstrated to exhibit significant efficacy against various types of human cancers, further highlighting its potential as a promising therapeutic target for cancer treatment. As such, numerous small-molecule inhibitors with distinct chemical scaffolds targeting PLK4 have been extensively investigated for the treatment of different human cancers, with several undergoing clinical evaluation (e.g., CFI-400945). Here, we review the structure, distribution, and biological functions of PLK4, encapsulate its intricate regulatory mechanisms of expression, and highlighting its multifaceted roles in cancer development and metastasis. Moreover, the recent advancements of PLK4 inhibitors in patent or literature are summarized, and their therapeutic potential as monotherapies or combination therapies with other anticancer agents are also discussed.

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The anti-tumor effect of polo-like kinase 4 (PLK4) inhibitor has been explored in several solid carcinomas, while its application in anaplastic thyroid cancer (ATC) remains scarce. Hence, the current study aimed to investigate the effect of PLK4 inhibitor on the malignant behaviors of ATC cell lines and its synergistic antitumor effect with sorafenib. C643 and 8305c cells were cultured in various concentrations of centrinone (PLK4 inhibitor) with or without sorafenib. Meanwhile, the cell viability, cell apoptosis, cell cycle and expressions of glycogen synthetase kinase beta (GSK3ß), p-GSK3ß, ß-catenin were determined. PLK4 mRNA and protein expressions were higher in most ATC cell lines than the normal thyroid epithelial cell line (all P < .05). Centrinone decreased cell viability, induced cell apoptosis, arrested cell cycle at G2/M phase and inactivated Wnt/ß-catenin signaling with dose-dependent manners in C643 and 8305c cells (all P < .05). Interestingly, centrinone plus sorafenib further improved antitumor effect (P < .05 at most concentrations), with the highest combination index at 5 nM centrinone plus 4 µM sorafenib in C643 cells, then 4 nM centrinone plus 4 µM sorafenib in C643 cells. Subsequently, centrinone plus sorafenib reduced cell viability, promoted cell apoptosis, facilitated cell cycle at G2/M phase and repressed Wnt/ß-catenin signaling more effectively compared with centrinone or sorafenib monotherapy in C643 and 8305c cells (all P < .05). PLK4 inhibitor exhibits antitumor effect and synergizes sorafenib via arresting cell cycle and inactivating Wnt/ß-catenin pathway in ATC.

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BACKGROUND: Polo-like kinase 4 (PLK4) is a crucial regulator for centriole replication and is reported to be aberrantly expressed in various cancers, where it participates to tumorigenesis. However, PLK4 effect in acute myeloid leukemia (AML), is still uncertain. This study investigates the function of PLK4 in AML. METHODS: Quantitative real-time PCR was used to measure the level of PLK4. Centrinone, a selective PLK4 small molecule inhibitor, was used for PLK4 inhibition and explore its effect in AML cells. The cell growth was detected by the CCK8, while the cell cycle and apoptosis were assessed by flow cytometry. The level of proteins associated with apoptosis, cell cycle and endoplasmic reticulum (ER) stress were analyzed by western blotting. RESULTS: PLK4 was overexpressed in AML cells. PLK4 knockdown or its specific inhibition by centrinone induced G2/M phase arrest via suppressing the expression of cyclin B1 and Cdc2 and promoting the level of proapoptotic proteins. Moreover, PLK4 targeting enhanced the level of proteins related to ER stress, such as GRP78, ATF4, ATF6, and CHOP. CONCLUSION: These findings demonstrated that targeting PLK4 can induce apoptosis, G2/M and ER stress in AML cells.

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The conserved Ser/Thr protein phosphatase 5 (PP5) is involved in the regulation of key cellular processes, including DNA damage repair and cell division in eukaryotes. As a co-chaperone of Hsp90, PP5 has been shown to modulate the maturation and activity of numerous oncogenic kinases. Here, we identify a novel substrate of PP5, the Polo-like kinase 4 (Plk4), which is the master regulator of centriole duplication in animal cells. We show that PP5 specifically interacts with Plk4, and is able to dephosphorylate the kinase in vitro and in vivo, which affects the interaction of Plk4 with its partner proteins. In addition, we provide evidence that PP5 and Plk4 co-localize to the centrosomes in Drosophila embryos and cultured cells. We demonstrate that PP5 is not essential; the null mutant flies are viable without a severe mitotic phenotype; however, its loss significantly reduces the fertility of the animals. Our results suggest that PP5 is a novel regulator of the Plk4 kinase in Drosophila.

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Polo-like kinase 4 (PLK4) promotes tumorigenesis and is associated with the prognosis of several solid tumors, while its clinical role in patients with renal cell carcinoma (RCC) remains unidentified. The present study aimed to analyze the association of PLK4 with clinicopathological characteristics and long-term prognosis in patients with RCC. The present study detected PLK4 protein and mRNA expression using immunohistochemical and reverse transcription-quantitative PCR assays in 120 patients with RCC. Disease-free survival (DFS) and overall survival (OS) time were calculated based on a median follow-up duration of 6.9 years (range, 1.2-9.9 years). PLK4 protein expression was elevated in tumor tissues compared with adjacent tissues (P<0.001). Upregulation of PLK4 protein was associated with increased T stage (P=0.023), N stage (P=0.014) and TNM stage (P=0.007). Additionally, elevated tumor PLK4 protein expression exhibited an associating trend (without statistical significance) with reduced DFS rate (P=0.066) and was associated with decreased OS rate (P=0.036). However, univariate Cox's regression analysis indicated that high PLK4 protein expression (compared with low PLK4 protein expression) was associated with reduced OS rate (P=0.040) but not with PFS rate (P=0.070). Following adjustment by multivariate Cox's regression analysis, PLK4 protein expression was associated with neither DFS nor OS rate (both P>0.050). Additionally, PLK4 mRNA expression was further detected in some patients (for which fresh specimens frozen in liquid nitrogen were available) to validate the aforementioned observations, and the expression was elevated in tumor tissues compared with adjacent tissues. Furthermore, increased PLK4 mRNA expression was associated with tumor size ≥7 cm, high TNM stage and reduced DFS rate (all P<0.050). PLK4 possesses a certain clinical utility in monitoring the clinical stage of patients with RCC, while its prognostic value requires further validation.

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High expression of polo-like kinase 4 (PLK4) promotes tumorigenesis and is correlated with poor prognosis in several kinds of cancer. However, the prognostic value of PLK4 in colorectal cancer (CRC) has not been elucidated. The aim of the present study was to investigate the association between PLK4 and the prognosis and effect of PLK4 inhibition on chemosensitivity in CRC. A total of 142 patients with CRC were enrolled, and 142 pairs of CRC and para-carcinoma tissues were used to measure PLK4 protein expression using immunohistochemistry (IHC). Among them, 69 pairs were used to detect PLK4 mRNA expression using reverse transcription-quantitative PCR. In addition, PLK4-small interfering RNA (siRNA) was transfected into CRC cells, followed by 5-fluorouracil (5-FU) treatment for it was a fundamental chemotherapy for CRC. In addition, western blotting was used to detect PLK4 protein expression among human colonic epithelial cell and human CRC cell lines, including HCT-116, LoVo, SW480 and HT-29, as well as nuclear translocation of ß-catenin. The IHC score and mRNA expression of PLK4 were higher in CRC tissues compared with para-carcinoma tissues (both P<0.001). Furthermore, the IHC score of tumor PLK4 was not correlated with pathological grade (P=0.585), T stage (P=0.357), N stage (P=0.107225) or tumor-node-metastasis (TNM) stage (P=0.093). The mRNA expression of tumor PLK4 was positively correlated with N stage (P=0.019) and TNM stage (P=0.004), but not with pathological grade (P=0.498) or T stage (P=0.112). Of note, the high protein expression of tumor PLK4 was an independent factor for poor overall survival (OS; P=0.048). In addition, PLK4 was elevated in CRC cell lines; PLK4-siRNA reduced the 50% inhibitory concentration value of 5-FU in HCT-116 (4.4±0.1 µM vs. 7.6±1.4 µM) and LoVo cells (5.5±0.6 µM vs. 9.9±1.8 µM) (both P<0.05). Besides, PLK4-siRNA decreased nuclear translocation of ß-catenin. In conclusion, the high expression of tumor PLK4 was associated with advanced TNM stage and shorter OS in patients with CRC. In addition, targeting PLK4 improved chemosensitivity in CRC cells.

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Polo-like kinase 4 (Plk4) is the master regulator of centriole assembly. Several evolutionarily conserved mechanisms strictly regulate Plk4 abundance and activity to ensure cells maintain a proper number of centrioles. In this issue of Genes & Development, Phan et al. (pp. 718-736) add to this growing list by describing a new mechanism of control that restricts Plk4 translation through competitive ribosome binding at upstream open reading frames (uORFs) in the mature Plk4 mRNA. Fascinatingly, this mechanism is especially critical in the development of primordial germ cells in mice that are transcriptionally hyperactive and thus exquisitely sensitive to Plk4 mRNA regulation.

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Centrioles are composed of a central cartwheel tethered to nine-fold symmetric microtubule (MT) blades. The centriole cartwheel and MTs are thought to grow from opposite ends of these organelles, so it is unclear how they coordinate their assembly. We previously showed that in Drosophila embryos an oscillation of Polo-like kinase 4 (Plk4) helps to initiate and time the growth of the cartwheel at the proximal end. Here, in the same model, we show that CP110 and Cep97 form a complex close to the distal-end of the centriole MTs whose levels rise and fall as the new centriole MTs grow, in a manner that appears to be entrained by the core cyclin-dependent kinase (Cdk)-Cyclin oscillator that drives the nuclear divisions in these embryos. These CP110 and Cep97 dynamics, however, do not appear to time the period of centriole MT growth directly. Instead, we find that changing the levels of CP110 and Cep97 appears to alter the Plk4 oscillation and the growth of the cartwheel at the proximal end. These findings reveal an unexpected potential crosstalk between factors normally concentrated at opposite ends of the growing centrioles, which might help to coordinate centriole growth. This article has an associated First Person interview with the first authors of the paper.

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Centrosomes are microtubule-organizing centers comprised of a pair of centrioles and the surrounding pericentriolar material. Abnormalities in centriole number are associated with cell division errors and can contribute to diseases such as cancer. Centriole duplication is limited to once per cell cycle and is controlled by the dosage-sensitive Polo-like kinase 4 (PLK4). Here, we show that PLK4 abundance is translationally controlled through conserved upstream open reading frames (uORFs) in the 5' UTR of the mRNA. Plk4 uORFs suppress Plk4 translation and prevent excess protein synthesis. Mice with homozygous knockout of Plk4 uORFs (Plk4 Δu/Δu ) are viable but display dramatically reduced fertility because of a significant depletion of primordial germ cells (PGCs). The remaining PGCs in Plk4 Δu/Δu mice contain extra centrioles and display evidence of increased mitotic errors. PGCs undergo hypertranscription and have substantially more Plk4 mRNA than somatic cells. Reducing Plk4 mRNA levels in mice lacking Plk4 uORFs restored PGC numbers and fully rescued fertility. Together, our data uncover a specific requirement for uORF-dependent control of PLK4 translation in counterbalancing the increased Plk4 transcription in PGCs. Thus, uORF-mediated translational suppression of PLK4 has a critical role in preventing centriole amplification and preserving the genomic integrity of future gametes.

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Fraxetin, a natural product isolated from herb Cortex Fraxini, has been demonstrated to exhibit anti-cancer effects on various cancers. The aim of this work is to investigate the anti-tumor effect of Fraxetin in prostate cancer and the potential mechanisms. In this study, the prostatic epithelial cell RWPE-1 and prostate cancer cell DU145 were exposed to Fraxetin (10, 20, 40, and 80 µM) to detect the changes in cell viability using cell counting kit-8 (CCK-8) assay. Fraxetin (10, 20, and 40 µM) was utilized to treat DU145 cell, then the changes in cell proliferation, apoptosis, migration, and invasion were assessed. Western blot assay was employed to detect the expression of proteins that participate in the above cellular processes as well as Polo-like kinase 4 (PLK4), phosphatidylinositol 3-kinase (PI3K). In addition to 40 µM Fraxetin treatment, DU145 cells were overexpressed with PLK4, and then the above experiments were repeated. Results revealed that Fraxetin markedly decreased DU145 cell viability, but didn't affect the cell viability of RWPE-1. Fraxetin suppressed cell proliferation, migration, invasion, and induced apoptosis of DU145 cells in a concentration-dependent manner. Furthermore, the expression of PLK4 and phosphorylated PI3K and protein kinase B (Akt) were reduced upon Fraxetin treatment. Finally, PLK4 overexpression significantly reversed all the effects of Fraxetin on DU145 cells. Collectively, Fraxetin acted as a cancer suppressor in prostate cancer through inhibiting PLK4 expression thereby inactivating PI3K/Akt signaling.

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Aberrated PLK4 expression has been reported in different malignancies and causes centrosome amplification, aneuploidy, and genomic instability. However, the mechanism by which PLK4 is regulated in carcinogenesis remains not fully characterised. Here, we showed that PLK4 was overexpressed in human HCC and overexpression of PLK4 predicted poorer patient prognosis. Unexpectedly, we found that induced expression of PLK4 promotes, but knockdown of PLK4 inhibits, HCC cell migration and invasion. Mechanistically, we found that TEC tyrosine kinase, which also promotes HCC cell migration, stabilizes PLK4 by phosphorylation. TEC directly phosphorylates PLK4 at tyrosine 86 residue, which not only stabilizes the protein but also enhances PLK4-mediated HCC cell invasion. Further investigation by transcriptome sequencing indicated that PLK4 promotes the phosphorylation of focal adhesion kinase to regulate the focal adhesion pathway in HCC cell migration. Taken together, our results demonstrated that PLK4 plays an important role in HCC metastasis and revealed for the first time the mechanism by which PLK4 promotes HCC metastasis via TEC phosphorylation.

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Subsequently to the publication of the above article, and a Corrigendum that has already been published with the intention of showing the corrected version of Fig. 5 (DOI: 10.3892/ijo.2021.5213; published online on May 5, 2021), the authors have realized that only the data for Fig. 5C needed to be corrected with respect to the original figure published in the article, and there was no need to have presented replacement data for Fig. 5E in the previous Corrigendum. The further revised version of Fig. 5, featuring the corrected data for Fig. 5C, is shown on the next page. All the authors agree to this Corrigendum. Note that the revisions made to this figure do not adversely affect the results reported in the paper, or the conclusions stated therein. The authors regret that Fig. 5 was not presented in its finally revised form in the previous Corrigendum, and offer their apologies to the Editor and to the readers of the Journal. [the original article was published in International Journal of Oncology 54: 479­490, 2019; DOI: 10.3892/ijo.2018.4659].

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BACKGROUND: Polo-like kinase 4 (PLK4) is a serine/threonine protein kinase located in the centriole of the chromosome during the cell cycle. PLK4 overexpression has been described in a variety of many common human epithelial tumors. Conversely, PLK4 acts as a haploinsufficient tumor suppressor in some situations, highlighting the importance of strict regulation of PLK4 expression, activity, and function. Meanwhile, the importance of chemoradiation resistance in rectal cancer is being emphasized more than ever. We aimed to analyze PLK4 expression and the tumor regression grade (TRG) in patients with rectal cancer, treated with chemoradiotherapy (CRT). METHODS: A retrospective study was conducted on 102 patients with rectal cancer who received preoperative CRT. Immunohistochemistry for PLK4 in paraffin-embedded tissue was performed from the biopsy and surgical specimens. RESULTS: We found significant association between high expression of PLK4 and poor response to neoadjuvant CRT (according to both Mandard and The Korean Society of Pathologists TRG systems) in the pre-CRT specimens. Other clinicopathologic parameters did not reveal any correlation with PLK4 expression. CONCLUSIONS: This study revealed an association between high expression of PLK4 in the pre-CRT specimens and TRG. Our results indicated that PLK4 could potentially be a new predictor for CRT effect in patients with rectal cancer.

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Polo-like kinase 4 (PLK4) has been reported to contribute to tumor growth, invasion, and metastasis. However, the role of PLK4 in human bladder cancer (BC) remains unclear. Here, we demonstrate the regulatory function of PLK4 in human BC progression. PLK4 is overexpressed in BC cell lines and tissues, and its overexpression correlated with poor prognosis. Our transcriptome analysis combined with subsequent functional assays indicated that PLK4 inhibition can suppress BC cell growth and induce cell cycle arrest at G1 phase via activation of the p38/p53/p21 pathway in vitro and in vivo. Overall, our data suggest that PLK4 is a critical regulator of BC cell proliferation, and thus, it may have potential as a novel molecular target for BC treatment.

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PLK-4 kinase plays an essential role in the cell cycle from regulating centriole duplication till cytokinesis and is therefore an attractive drug target in cancers such as breast, lung, and central nervous system tumors. CFI-400945 is an efficient PLK-4 inhibitor and inhibits other non-PLK family proteins at nanomolar concentrations. We have compared PLK-4 with other kinases to understand its similarity based on multiple sequence alignments from protein sequences of primary structures, outer and buried residues, and compact active site conservation based on three-dimensional motifs. These in-depth studies provide information on known interface targets and design of more selective inhibitors to PLK-4. Further, pharmacophore features based on CFI-400945 bound to PLK-4 were used for searching library of compounds that were screened using deep learning methods to bind PLK-4. The shortlisted molecules were docked into PLK-4 active site and were validated using molecular docking and molecular dynamics simulations studies. MM-PBSA calculations revealed the stability of hit molecules and PLK-4 complexes in comparison with CFI-400945 and the contribution to binding from key active site residues.

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Subsequently to the publication of the above article, the author realized that Fig. 5 on p. 486 contained some errors on account of the figure having been compiled incorrectly; essentially, the published version of the figure contained incorrect images for the panels presented in Fig. 5C and E. The authors were able to re­examine their original data, and identify the data that was intended to have been shown for these figure parts. The corrected version of Fig. 5 is shown on the next page, featuring the correct data for Fig. 5C and E, including new bar charts showing the quantification of these data. The authors confirm that these data continue to support the main conclusions presented in their paper, and are grateful to the Editor of International Journal of Oncology for allowing them this opportunity to publish a Corrigendum. They also apologize to the readership for any inconvenience caused. [the original article was published in International Journal of Oncology 54: 479­490, 2019; DOI: 10.3892/ijo.2018.4659].