Regulation of cancer stem cells by CXCL1, a chemokine whose secretion is controlled by MCM2.

BACKGROUND: A high expression pattern of minichromosome maintenance 2 (MCM2) has been observed in various cancers. MCM2 is a protein involved in the cell cycle and plays a role in cancer growth and differentiation by binding to six members of the MCM subfamily. The MCM protein family includes MCM2 through MCM7. METHODS: MCM2 has shown high expression in both lung cancer stem cells (LCSCs) and glioma stem cells (GSCs). We investigated the characteristics of CSCs and the regulation of the epithelial-to-mesenchymal transition (EMT) phenomenon in LCSCs and GSCs by MCM2. Additionally, we explored secreted factors regulated by MCM2. RESULTS: There was a significant difference in survival rates between lung cancer patients and brain cancer patients based on MCM2 expression. MCM2 was found to regulate both markers and regulatory proteins in LCSCs. Moreover, MCM2 is thought to be involved in cancer metastasis by regulating cell migration and invasion, not limited to lung cancer but also identified in glioma. Among chemokines, chemokine (C-X-C motif) ligand 1 (CXCL1) was found to be regulated by MCM2. CONCLUSIONS: MCM2 not only participates in the cell cycle but also affects cancer cell growth by regulating the external microenvironment to create a favorable environment for cells. MCM2 is highly expressed in malignant carcinomas, including CSCs, and contributes to the malignancy of various cancers. Therefore, MCM2 may represent a crucial target for cancer therapeutics.

MCM2 is involved in subtyping and tamoxifen resistance of ERα-positive breast cancer by acting as the downstream factor of ERα.

Tamoxifen (TAM) resistance is finally developed in over 40% of patients with estrogen receptor α-positive breast cancer (ERα+ -BC), documenting that discovering new molecular subtype is needed to confer perception to the heterogeneity of ERα+ -BC. We obtained representative gene sets subtyping ERα+ -BC using gene set variation analysis (GSVA), non-negative matrix factorization (NMF), and COX regression methods on the basis of METABRIC, TCGA, and GEO databases. Furthermore, the risk score of ERα+ -BC subtyping was established using least absolute shrinkage and selection operator (LASSO) regression on the basis of genes in the representative gene sets, thereby generating the two subtypes of ERα+ -BC. We further found that minichromosome maintenance complex component 2 (MCM2) functioned as the hub gene subtyping ERα+ -BC using GO, KEGG, and MCODE. MCM2 expression was capable for specifically predicting 1-year overall survival (OS) of ERα+ -BC and correlated with T stage, AJCC stage, and tamoxifen (TAM) sensitivity of ERα+ -BC. The downregulation of MCM2 expression inhibited proliferation, migration, and invasion of TAM-resistant cells and promoted G0/G1 arrest. Altogether, tamoxifen resistance entails that MCM2 is a hub gene subtyping ERα+ -BC, providing a novel dimension for discovering a potential target of TAM-resistant BC.

Coiled-coil domain-containing 154 promotes colorectal cancer proliferation and metastasis via interacting with minichromosome maintenance complex component 2.

Coiled-Coil Domain-Containing (CCDC) is a large class of structural proteins containing left-handed supercoiled structure. The clinical value and the functional implication of CCDC in colorectal cancer (CRC) remain unknown. Based on the genetic, transcriptional, and clinical data from The Cancer Genome Atlas, five of thirty-six CCDC proteins were differentially expressed in the CRC and associated with the survival of patients with CRC. A CCDC-score model was established to evaluate the prognosis of patients. The potential function of Coiled-Coil Domain-Containing 154 (CCDC154) was investigated using bioinformatical methods, which unveiled that high expression of CCDC154 indicates poor survival for patients with CRC and correlates with low infiltration of CD8+ T cells and high infiltration of neutrophils, indicating that CCDC154 enhances tumor growth and metastasis. CCDC154 interacts with Minichromosome Maintenance Complex Component 2 (MCM2) protein and promotes malignant phenotype via MCM2. We validated the expression level and survival prediction value of CCDC154 in clinical samples, and analyzed its co-expression of MCM2, Ki-67 and p53. This work discloses the role of CCDC in clinical setting and CCDC154 functions in CRC.

Characterization of histone chaperone MCM2 as a key regulator in arsenic-induced depletion of H3.3 at genomic loci.

Arsenic exposure is associated with an increased risk of many cancers, and epigenetic mechanisms play a crucial role in arsenic-mediated carcinogenesis. Our previous studies have shown that arsenic exposure induces polyadenylation of H3.1 mRNA and inhibits the deposition of H3.3 at critical gene regulatory elements. However, the precise underling mechanisms are not yet understood. To characterize the factors governing arsenic-induced inhibition of H3.3 assembly through H3.1 mRNA polyadenylation, we utilized mass spectrometry to identify the proteins, especially histone chaperones, with reduced binding affinity to H3.3 under conditions of arsenic exposure and polyadenylated H3.1 mRNA overexpression. Our findings reveal that the interaction between H3.3 and the histone chaperon protein MCM2 is diminished by both polyadenylated H3.1 mRNA overexpression and arsenic treatment in human lung epithelial BEAS-2B cells. The increased binding of MCM2 to H3.1, resulting from elevated H3.1 protein levels, appears to contribute to the reduced availability of MCM2 for H3.3. To further investigate the role of MCM2 in H3.3 deposition during arsenic exposure and H3.1 mRNA polyadenylation, we overexpressed MCM2 in BEAS-2B cells overexpressing polyadenylated H3.1 or exposed to arsenic. Our results demonstrate that MCM2 overexpression attenuates H3.3 depletion at several genomic loci, suggesting its involvement in the arsenic-induced displacement of H3.3 mediated by H3.1 mRNA polyadenylation. These findings suggest that changes in the association between histone chaperone MCM2 and H3.3 due to polyadenylation of H3.1 mRNA may play a pivotal role in arsenic-induced carcinogenesis.

KNTC1 and MCM2 are the molecular targets of gallbladder cancer.

BACKGROUND: Gallbladder carcinoma is a malignant epithelial tumor of gallbladder with a high degree of malignancy. However, relationship between KNTC1 and MCM2 and gallbladder cancer is unclear. METHODS: GSE139682 and GSE202479 were downloaded from gene expression omnibus (GEO). Differentially expressed genes (DEGs) were screened. Functional enrichment analysis and gene set enrichment analysis (GSEA) were performed. Protein-protein interaction (PPI) Network was constructed and analyzed. Gene expression heat map was drawn. Comparative toxicogenomics database (CTD) analysis was performed to find diseases most related to core genes. TargetScan was performed for screening miRNAs that regulated central DEGs. RESULTS: 230 DEGs were identified. According to GObp analysis, they were mainly concentrated in regulation of ossification, regulation of spindle microtubule and centromere attachment, cytoskeleton tissue of cortical actin. According to GOcc analysis, they are mainly concentrated in plasma membrane part, cell junction, plasma membrane region and anterior membrane. According to GOmf analysis, they are mainly enriched in protein homodimerization activity, proximal promoter sequence-specific DNA binding and sulfur compound binding. KEGG showed that target genes were mainly enriched in Hippo signal pathway, p53 signal pathway and cancer pathway. KIFC2, TUBG1, RACGAP1, CHMP4C, SFN and MYH11 were identified as core genes. Gene expression heat map showed that KNTC1, MCM2, CKAP2, RACGAP1, CCNB1 were highly expressed in gallbladder carcinoma samples. CTD analysis showed that KNTC1, MCM2, CKAP2, RACGAP1, CCNB1 were associated with head and neck squamous cell carcinoma, necrosis, inflammation and hepatomegaly. CONCLUSIONS: KNTC1 and MCM2 are highly expressed in gallbladder cancer. Higher expression level correlates with worse prognosis.

Circ_0104700 contributes to acute myeloid leukemia progression by enhancing MCM2 expression through targeting miR-665.

OBJECTIVE: Acute myeloid leukemia (AML) is a common blood cancer associated with poor prognosis and high mortality. In this study, we investigated the role and underlying mechanism of action of circ_0104700 in the pathogenesis of AML. METHODS: Circ_0104700 was screened from the GEO database and detected in AML samples and cell lines. The effect of circ_0104700 on AML was analyzed using a methylcellulose colony assay, CCK-8 assay, and cell cycle and apoptosis analyses. The mechanism was explored using bioinformatic analysis, quantitative reverse transcription-PCR, dual-luciferase reporter assays, northern blotting and western blot analysis in AML cells. RESULTS: Circ_0104700 expression was higher in AML patients and AML cell lines. Functionally, circ_0104700 depletion attenuated cell viability and induced apoptosis in MV-4-11 and Kasumi-1 cells. Circ_0104700 depletion enhanced the G0/G1-phase proportion but reduced the proportion of S-phase cells in MV-4-11 and Kasumi-1 cells. circ_0104700 served as a competing endogenous RNA of miR-665 and enhanced MCM2 expression by sponging miR-665 in MV-4-11 and Kasumi-1 cells. Silencing circ_0104700 repressed the proliferation and cell cycle and induced apoptosis of MV-4-11 and Kasumi-1 cells by inhibiting miR-665. MCM2 depletion alleviated the proliferation and cell cycle and enhanced the apoptosis of MV-4-11 and Kasumi-1 cells by inactivating JAK/STAT signaling. JAK/STAT signaling was involved in circ_0104700-mediated malignant phenotypes of MV-4-11 and Kasumi-1 cells. CONCLUSION: circ_0104700 contributed to AML progression by enhancing MCM2 expression by targeting miR-665. Our findings provide novel potential therapeutic targets for AML, including circ_0104700, miR-665, and MCM2.

RAD51 bypasses the CMG helicase to promote replication fork reversal.

Replication fork reversal safeguards genome integrity as a replication stress response. DNA translocases and the RAD51 recombinase catalyze reversal. However, it remains unknown why RAD51 is required and what happens to the replication machinery during reversal. We find that RAD51 uses its strand exchange activity to circumvent the replicative helicase, which remains bound to the stalled fork. RAD51 is not required for fork reversal if the helicase is unloaded. Thus, we propose that RAD51 creates a parental DNA duplex behind the helicase that is used as a substrate by the DNA translocases for branch migration to create a reversed fork structure. Our data explain how fork reversal happens while maintaining the helicase in a position poised to restart DNA synthesis and complete genome duplication.

The importance of nuclear RAGE-Mcm2 axis in diabetes or cancer-associated replication stress.

An elevated frequency of DNA replication defects is associated with diabetes and cancer. However, data linking these nuclear perturbations to the onset or progression of organ complications remained unexplored. Here, we report that RAGE (Receptor for Advanced Glycated Endproducts), previously believed to be an extracellular receptor, upon metabolic stress localizes to the damaged forks. There it interacts and stabilizes the minichromosome-maintenance (Mcm2-7) complex. Accordingly, RAGE deficiency leads to slowed fork progression, premature fork collapse, hypersensitivity to replication stress agents and reduction of viability, which was reversed by the reconstitution of RAGE. This was marked by the 53BP1/OPT-domain expression and the presence of micronuclei, premature loss-of-ciliated zones, increased incidences of tubular-karyomegaly, and finally, interstitial fibrosis. More importantly, the RAGE-Mcm2 axis was selectively compromised in cells expressing micronuclei in human biopsies and mouse models of diabetic nephropathy and cancer. Thus, the functional RAGE-Mcm2/7 axis is critical in handling replication stress in vitro and human disease.

ESRG is critical to maintain the cell survival and self-renewal/pluripotency of hPSCs by collaborating with MCM2 to suppress p53 pathway.

The mechanisms of self-renewal and pluripotency maintenance of human pluripotent stem cells (hPSCs) have not been fully elucidated, especially for the role of those poorly characterized long noncoding RNAs (lncRNAs). ESRG is a lncRNA highly expressed in hPSCs, and its functional roles are being extensively explored in the field. Here, we identified that the transcription of ESRG can be directly regulated by OCT4, a key self-renewal factor in hPSCs. Knockdown of ESRG induces hPSC differentiation, cell cycle arrest, and apoptosis. ESRG binds to MCM2, a replication-licensing factor, to sustain its steady-state level and nuclear location, safeguarding error-free DNA replication. Further study showed that ESRG knockdown leads to MCM2 abnormalities, resulting in DNA damage and activation of the p53 pathway, ultimately impairs hPSC self-renewal and pluripotency, and induces cell apoptosis. In summary, our study suggests that ESRG, as a novel target of OCT4, plays an essential role in maintaining the cell survival and self-renewal/pluripotency of hPSCs in collaboration with MCM2 to suppress p53 signaling. These findings provide critical insights into the mechanisms underlying the maintenance of self-renewal and pluripotency in hPSCs by lncRNAs.

Hypoxia Inhibits Cell Cycle Progression and Cell Proliferation in Brain Microvascular Endothelial Cells via the miR-212-3p/MCM2 Axis.

Hypoxia impairs blood-brain barrier (BBB) structure and function, causing pathophysiological changes in the context of stroke and high-altitude brain edema. Brain microvascular endothelial cells (BMECs) are major structural and functional elements of the BBB, and their exact role in hypoxia remains unknown. Here, we first deciphered the molecular events that occur in BMECs under 24 h hypoxia by whole-transcriptome sequencing assay. We found that hypoxia inhibited BMEC cell cycle progression and proliferation and downregulated minichromosome maintenance complex component 2 (Mcm2) expression. Mcm2 overexpression attenuated the inhibition of cell cycle progression and proliferation caused by hypoxia. Then, we predicted the upstream miRNAs of MCM2 through TargetScan and miRanDa and selected miR-212-3p, whose expression was significantly increased under hypoxia. Moreover, the miR-212-3p inhibitor attenuated the inhibition of cell cycle progression and cell proliferation caused by hypoxia by regulating MCM2. Taken together, these results suggest that the miR-212-3p/MCM2 axis plays an important role in BMECs under hypoxia and provide a potential target for the treatment of BBB disorder-related cerebrovascular disease.

MCM2 in human cancer: functions, mechanisms, and clinical significance.

BACKGROUND: Aberrant DNA replication is the main source of genomic instability that leads to tumorigenesis and progression. MCM2, a core subunit of eukaryotic helicase, plays a vital role in DNA replication. The dysfunction of MCM2 results in the occurrence and progression of multiple cancers through impairing DNA replication and cell proliferation. CONCLUSIONS: MCM2 is a vital regulator in DNA replication. The overexpression of MCM2 was detected in multiple types of cancers, and the dysfunction of MCM2 was correlated with the progression and poor prognoses of malignant tumors. According to the altered expression of MCM2 and its correlation with clinicopathological features of cancer patients, MCM2 was thought to be a sensitive biomarker for cancer diagnosis, prognosis, and chemotherapy response. The anti-tumor effect induced by MCM2 inhibition implies the potential of MCM2 to be a novel therapeutic target for cancer treatment. Since DNA replication stress, which may stimulate anti-tumor immunity, frequently occurs in MCM2 deficient cells, it also proposes the possibility that MCM2 targeting improves the effect of tumor immunotherapy.

ITRAQ-based quantitative proteomic analysis reveals that VPS35 promotes the expression of MCM2-7 genes in HeLa cells.

Vacuolar protein sorting 35 (VPS35) is a major component of the retromer complex that regulates endosomal trafficking in eukaryotic cells. Recent studies have shown that VPS35 promotes tumor cell proliferation and affects the nuclear accumulation of its interacting partner. In this study, isobaric tags for relative and absolute quantitation (iTRAQ)-based mass spectrometry were used to measure the changes in nuclear protein abundance in VPS35-depleted HeLa cells. A total of 47 differentially expressed proteins were identified, including 27 downregulated and 20 upregulated proteins. Gene ontology (GO) analysis showed that the downregulated proteins included several minichromosome maintenance (MCM) proteins described as cell proliferation markers, and these proteins were present in the MCM2-7 complex, which is essential for DNA replication. Moreover, we validated that loss of VPS35 reduced the mRNA and protein expression of MCM2-7 genes. Notably, re-expression of VPS35 in VPS35 knockout HeLa cells rescued the expression of these genes. Functionally, we showed that VPS35 contributes to cell proliferation and maintenance of genomic stability of HeLa cells. Therefore, these findings reveal that VPS35 is involved in the regulation of MCM2-7 gene expression and establish a link between VPS35 and cell proliferation.

WASHC1 interacts with MCM2-7 complex to promote cell survival under replication stress.

BACKGROUND: WASHC1 is a member of the Wiskott-Aldrich syndrome protein (WASP) family and is involved in endosomal protein sorting and trafficking through the generation of filamentous actin (F-actin) via activation of the Arp2/3 complex. There is increasing evidence that WASHC1 is present in the nucleus and nuclear WASHC1 plays important roles in regulating gene transcription, DNA repair as well as maintaining nuclear organization. However, the multi-faceted functions of nuclear WASHC1 still need to be clarified. METHODS AND RESULTS: We show here that WASHC1 interacts with several components of the minichromosome maintenance (MCM) 2-7 complex by using co-immunoprecipitation and in situ proximity ligation assay. WASHC1-depleted cells display normal DNA replication and S-phase progression. However, loss of WASHC1 sensitizes HeLa cells to DNA replication inhibitor hydroxyurea (HU) and increases chromosome instability of HeLa and 3T3 cells under condition of HU-induced replication stress. Re-expression of nuclear WASHC1 in WASHC1KO 3T3 cells rescues the deficiency of WASHC1KO cells in the chromosomal stability after HU treatment. Moreover, chromatin immunoprecipitation assay indicates that WASHC1 associates with DNA replication origins, and knockdown of WASHC1 inhibits MCM protein loading at origins. CONCLUSIONS: Since efficient loading of excess MCM2-7 complexes is required for cells to survive replicative stress, these results demonstrate that WASHC1 promotes cell survival and maintain chromosomal stability under replication stress through recruitment of excess MCM complex to origins.

MCMBP promotes the assembly of the MCM2-7 hetero-hexamer to ensure robust DNA replication in human cells.

The MCM2-7 hetero-hexamer is the replicative DNA helicase that plays a central role in eukaryotic DNA replication. In proliferating cells, the expression level of the MCM2-7 hexamer is kept high, which safeguards the integrity of the genome. However, how the MCM2-7 hexamer is assembled in living cells remains unknown. Here, we revealed that the MCM-binding protein (MCMBP) plays a critical role in the assembly of this hexamer in human cells. MCMBP associates with MCM3 which is essential for maintaining the level of the MCM2-7 hexamer. Acute depletion of MCMBP demonstrated that it contributes to MCM2-7 assembly using nascent MCM3. Cells depleted of MCMBP gradually ceased to proliferate because of reduced replication licensing. Under this condition, p53-positive cells exhibited arrest in the G1 phase, whereas p53-null cells entered the S phase and lost their viability because of the accumulation of DNA damage, suggesting that MCMBP is a potential target for killing p53-deficient cancers.

Expression of MCM2 as a Proliferative Marker in Actinic Keratosis and Cutaneous Squamous Cell Carcinoma.

BACKGROUND/AIM: Minichromosome maintenance protein 2 (MCM2) can be considered an indicator of cancer clinical outcome. In this study, we tried to estimate the usefulness of assessing MCM2 protein expression in actinic keratosis (AK) and cutaneous squamous cell carcinoma (cSCC). MATERIALS AND METHODS: The study included 22 lesions of AK, 57 of cSCC and 17 tissue samples of the healthy skin. RESULTS: Higher average expression of MCM2 protein in cSCC and AK was demonstrated in comparison to healthy skin (p=0.01). Likewise, the level of MCM2 expression differed statistically significantly (p=0.02) between SCC, AK, and healthy skin. Significant correlations between MCM2 expression and Ki-67 and p53 antigen were found (r=0.51, p=0.01; r=0.45, p=0.04 respectively) in AK lesions, however these relationships were not noted in cSCC. CONCLUSION: MCM2 is overexpressed in both AK and cSCC lesions, however this protein cannot be considered an important indicator of proliferation in cSCC.

MCM2 promotes the proliferation, migration and invasion of cholangiocarcinoma cells by reducing the p53 signaling pathway.

The abnormal expressions of minichromosome maintenance protein 2 (MCM2) are closely related to the development of various kinds of cancers. We aimed to explore the functions and potential molecular mechanisms of MCM2 gene in cholangiocarcinoma (CCA) cell lines (Huh28 and RBE). First, the cell counting kit-8 (CCK-8), plate clone formation, transwell and invasion assays showed that MCM2 promotes the proliferation, migration and invasion of CCA cells. Flow cytometry assays showed that MCM2 significantly promotes the cell cycle, and inhibits the apoptosis of CCA cells. Further, by analyzing the RNA sequencing data of cholangiocarcinoma, we found that MCM2 gene is significantly negatively correlated with p53 signaling pathway. Quantitative real time polymerase chain reaction (qRT-PCR) and Western blotting (WB) assays confirmed that MCM2 in CCA cells significantly down-regulated the mRNA and protein expression levels of p53 and BAX, and up-regulated the mRNA and protein expression levels of BCL2 and CCND1. Flow cytometry, qRT-PCR and WB assays confirmed that MCM2 promotes CCA through p53 pathway. Finally, we found that MCM2 is up-regulated in CCA tissues compared to the matched non-tumor cholangiocarcinoma tissues, and the high expressions of MCM2 are significantly associated with the poor clinical outcomes of CCA patients. In conclusion, this study revealed that MCM2 promotes the development of CCA by reducing the p53 pathway, and its high expression levels predict poor prognosis in CCA patients. These results provide a theoretical basis for the development of new clinical diagnosis and treatment of cholangiocarcinoma in the future.

Clinicopathologic significance of DNA replication licensing factors in head and neck diffuse large B-cell lymphoma.

OBJECTIVE: Diffuse large B-cell lymphoma (DLBCL) harbors defects in the proliferation pathway. We performed multiparameter analysis of proteins expressed during different cell cycle phases and correlated them with clinical parameters of head and neck DLBCLs. STUDY DESIGN: Thirty-nine DLBCLs were staged and immunohistochemically stained with MCM2, Ki67, and geminin. The receiver operating characteristic curve and its area under the curve were calculated, and sensitivity vs specificity curve analysis was performed. RESULTS: The highest labeling index was in MCM2, followed by Ki67 and geminin (P < .001). All pairs showed significant differences (P < .001). The best cutoff points to differentiate limited from advanced disease were 68% and 45% for MCM2 and Ki67, respectively. There was no acceptable cutoff for geminin (area under the curve = 0.667, P = .134). MCM2/Ki67 (P = .293) and geminin/Ki67 (P = .233) ratios did not differ between the stages. The median (interquartile range) of the geminin/Ki67 ratio was 0.57 (0.68), translating to a reduced G1. CONCLUSIONS: We suggest a role for cell cycle-related proteins in the biology and behavior of DLBCLs. MCM2 and Ki67 cutoffs can be a potential option to differentiate limited from advanced disease, where imaging and laboratory techniques are unavailable. The G1 decrease and the significantly higher MCM2 expression compared to Ki67 indicate replication disturbances, making factors involved in the G1 phase targets for treatment.

Distinctive expression of DNA replication factors in squamous cell carcinomas of the lip, face and oral cavity.

OBJECTIVE: Uncontrolled proliferation and aberrations in cell-cycle progression are fundamental issues in cancer. In this study we aimed to determine and compare deoxyribonucleic acid (DNA) replication licensing factors at the mRNA and protein levels among squamous cell carcinomas (SCCs) of the lip, facial-skin and oral cavity. MATERIALS AND METHODS: A total of 103 lip, oral and face SCCs were immunohistochemically stained with MCM2 (mini-chromosome maintenance 2), geminin, and ki67, and their labeling-indices were calculated. Also, 57 SCCs from the same regions along with their adjacent normal tissues underwent quantitative reverse transcription-polymerase chain reaction analysis. RESULTS: All three proteins were overexpressed in the studied SCCs, but only geminin (P = 0.004) showed significant difference among the three regions, with higher levels in oral SCCs compared to lip (P = 0.005) and skin (P = 0.024) tumors. Geminin expression did not differ between skin- and lip-SCCs (P = 0.822). MCM2/ki67 ratio was higher in oral- compared to skin-neoplasms (P = 0.039), but no difference was found in geminin/ki67 among the SCC-subsites. There were significant differences in MCM2 and geminin mRNA between carcinomatous- and normal-tissues in all tumors, but not among the three locations. CONCLUSION: MCM2 and geminin are involved in the tumorigenesis of lip, face and oral SCC at both mRNA- and protein-levels. Geminin may have a role in the site-specific biologic behavior of SCC. Skin SCCs had the highest proportion of licensed non-proliferating cells, while actively proliferating cells were more prominent in oral tumors. Regarding DNA replication, lip SCCs seem to be closer to skin tumors compared to their oral counterparts.

Demethylation at enhancer upregulates MCM2 and NUP37 expression predicting poor survival in hepatocellular carcinoma patients.

BACKGROUND: Identification of novel biomarker is important for development of molecular-targeted therapy agents for patients with hepatocellular carcinoma (HCC). This study aims to identify potential prognostic biomarkers and investigate epigenetic mechanism of HCC development. METHODS: Public bulk-RNA seq datasets and proteomic dataset were screened for identification of potential prognostic biomarkers for HCC patients. Public methylomic datasets were analyzed for deciphering the epigenetic mechanism regulating HCC-associated gene expression. Immunoblotting, immunohistochemistry, real-time PCR, and pyrosequencing were used to validate the findings from bioinformatic analyses. RESULTS: Minichromosome maintenance complex component 2 (MCM2) and nucleoporin 37 (NUP37) were overexpressed in human HCC tissues and hepatoma cell lines. MCM2 significantly positively correlated with NUP37 expression. Higher expression of MCM2 or NUP37 was significantly associated with advanced tumor stage and worse overall survival in 3 large independent HCC cohorts (n = 820). MCM2 and NUP37 overexpression are independent prognostic risk factors for HCC patients. Demethylation at an enhancer of MCM2 gene was a common event in patients with HCC, which significantly negatively correlated with MCM2 and NUP37 mRNA expression. CONCLUSIONS: Demethylation at enhancer regulates MCM2 and NUP37 expression in HCC. MCM2 and NUP37 are promising prognostic biomarkers and potential targets for epigenetic therapy in HCC patients.

Thiabendazole Inhibits Glioblastoma Cell Proliferation and Invasion Targeting Mini-chromosome Maintenance Protein 2.

Thiabendazole (TBZ), approved by the US Food and Drug Administration (FDA) for human oral use, elicits a potential anticancer activity on cancer cells in vitro and in animal models. Here, we evaluated the efficacy of TBZ in the treatment of human glioblastoma multiforme (GBM). TBZ reduced the viability of GBM cells (P3, U251, LN229, A172, and U118MG) relative to controls in a dose- and time-dependent manner. However, normal human astrocytes (NHA) exhibited a greater IC50 than tumor cell lines and were thus more resistant to its cytotoxic effects. 5-Ethynyl-2'-deoxyuridine (EdU)-positive cells and the number of colonies formed were decreased in TBZ-treated cells (at 150 µM, P < 0.05 and at 150 µM, P < 0.001, respectively). This decrease in proliferation was associated with a G2/M arrest as assessed with flow cytometry, and the downregulation of G2/M check point proteins. In addition, TBZ suppressed GBM cell invasion. Analysis of RNA sequencing data comparing TBZ-treated cells with controls yielded a group of differentially expressed genes, the functions of which were associated with the cell cycle and DNA replication. The most significantly downregulated gene in TBZ-treated cells was mini-chromosome maintenance protein 2 (MCM2). SiRNA knockdown of MCM2 inhibited proliferation, causing a G2/M arrest in GBM cell lines and suppressed invasion. Taken together, our results demonstrated that TBZ inhibited proliferation and invasion in GBM cells through targeting of MCM2. SIGNIFICANCE STATEMENT: TBZ inhibits the proliferation and invasion of glioblastoma cells by downregulating the expression of MCM2. These results support the repurposing of TBZ as a possible therapeutic drug in the treatment of GBM.