CircROR1 upregulates CCNE1 expression to promote melanoma invasion and metastasis by recruiting KAT2A.

Circular RNAs (circRNAs) play important roles in cancer occurrence and progression. To explore and elucidate the clinical significance of specific circular RNA in melanoma and its potential molecular mechanism. CircROR1 expression in melanoma cells and tissues was confirmed by qRT-PCR and ISH. qRT-PCR and Western blotting were performed to measure the levels of CCNE1, KAT2A, MMP9 and TIMP2. MTT, Transwell and wound healing assays were performed to evaluate cell proliferation, invasion and metastasis. A xenograft mouse model was established to further verify the CircROR1/CCNE1 axis in vivo. RNA pull-down and RIP assays were performed to detect the direct interaction KAT2A and CircROR1. A ChIP assay was used to investigate the enrichment of H3K9ac acetylation in the CCNE1 promoter. CircROR1 was significantly upregulated in metastatic melanoma cells and tissues, promoting proliferation, invasion and metastasis in vitro and tumour growth in vivo. CircROR1 overexpression increased CCNE1 and MMP9 protein expression and decreased TIMP2 protein expression. Functional rescue assays demonstrated that CircROR1 played a role in promoting malignant progression through CCNE1. CircROR1 specifically bound to the KAT2A protein without affecting its expression. CircROR1 overexpression increased the level of H3K9ac modification in the CCNE1 promoter region by recruiting KAT2A, thus upregulating CCNE1 expression. CircROR1 upregulates CCNE1 expression through KAT2A-mediated histone acetylation. Our research confirms the critical role of CircROR1 in melanoma invasion and metastasis, and CircROR1 could serve as a potential therapeutic target for melanoma treatment.

PPM1D activity promotes the replication stress caused by cyclin E1 overexpression.

Oncogene-induced replication stress has been recognized as a major cause of genome instability in cancer cells. Increased expression of cyclin E1 caused by amplification of the CCNE1 gene is a common cause of replication stress in various cancers. Protein phosphatase magnesium-dependent 1 delta (PPM1D) is a negative regulator of p53 and has been implicated in termination of the cell cycle checkpoint. Amplification of the PPM1D gene or frameshift mutations in its final exon promote tumorigenesis. Here, we show that PPM1D activity further increases the replication stress caused by overexpression of cyclin E1. In particular, we demonstrate that cells expressing a truncated mutant of PPM1D progress faster from G1 to S phase and fail to complete licensing of the replication origins. In addition, we show that transcription-replication collisions and replication fork slowing caused by CCNE1 overexpression are exaggerated in cells expressing the truncated PPM1D. Finally, replication speed and accumulation of focal DNA copy number alterations caused by induction of CCNE1 expression was rescued by pharmacological inhibition of PPM1D. We propose that increased activity of PPM1D suppresses the checkpoint function of p53 and thus promotes genome instability in cells expressing the CCNE1 oncogene.

Regulation of Interferon-ß-Modified Human Umbilical Cord Mesenchymal Stem Cell-Derived Exosomes in Proliferation and Apoptosis of Prostate Cancer Cells.

Prostate cancer (PCa) poses a serious burden to men. Interferon-ß (IFN-ß) is implicated in cancer cell growth. This study hence explored the regulation of IFN-ß-modified human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-Exos) in PCa cells. In vitro-cultured hUCMSCs were transfected with pcDNA3.1-IFN-ß plasmid or IFN-ß siRNA. hUCMSC-Exos were extracted by ultracentrifugation and identified. PCa cells (PC3 and LNCap) were treated with Exos. Cellular internalization of Exos by cells was detected by uptake assay. Cell proliferation, cycle, and apoptosis were evaluated by CCK-8, EdU staining, and flow cytometry. Levels of cell cycle-related proteins (cyclin D/cyclin E) were determined by Western blot. The effect of IFN-ß-modified hUCMSC-Exos in vivo was analyzed. IFN-ß-modified hUCMSC-Exos (Exooe-IFN-ß or Exosi-IFN-ß) were successfully isolated. IFN-ß was encapsulated in Exos, and PCa cells could uptake Exos. After treating with Exooe-IFN-ß, PCa cell proliferation was impeded, the percentage of cells in the G0/G1 phase, cyclin D/cyclin E levels, and cell apoptotic rate were elevated, while cells treated with Exooe-IFN-ß exhibited contrary trends. IFN-ß-modified hUCMSC-Exos reduced PCa tumor size and weight in vivo. Conjointly, IFN-ß-modified hUCMSC-Exos suppress PCa cell proliferation and facilitate apoptosis.

PKMYT1: A Potential Target for CCNE1 Amplificated Colorectal Tumors.

Colorectal cancer is a malignant tumor with higher morbidity and mortality. The purpose of this study is to investigate whether inhibition of Protein Kinase, Membrane Associated Tyrosine/Threonine 1 (PKMYT1) affects tumor cell proliferation, survival and migration in colon tumors with high Cyclin E1 (CCNE1) expression. PcDNA3.1-CCNE1 vector and si-PKMYT1 were transfected in SW480 cells by Lipofectamine 2000. Q-PCR and western blot assay were processed to detect the expression. Transwell assay and Edu assay were undertaken to verify the migration and proliferation. CCNE1 promotes the proliferation and migration of SW480. Silencing of PKMYT1 inhibited the proliferation of tumor cells. Silencing the expression of PKMYT1 under the premise of overexpression of CCNE1, the level of Cyclin Dependent Kinase 1 (CDK1)-PT14 was reduced, indicating that the cell cycle was blocked. The expression of γH2AX increased significantly, indicating that the DDR pathway of tumor cells was activated and DNA damage accumulated. The results of immunofluorescence microscopy showed significantly increased expression of DNA damage-associated marker (γH2AX: H2AX Variant Histone). In CCNE1 amplificated colorectal tumor cells, knockdown of PKMYT1 reduced cells in S phase, inhibited cell proliferation and promoted cell apoptosis, confirming that PKMYT1 was a potential therapeutic target for colorectal tumor. This study may verify a potential therapeutic target and provide a new idea for the treatment of colorectal cancer in the future.

FBXL12 degrades FANCD2 to regulate replication recovery and promote cancer cell survival under conditions of replication stress.

Fanconi anemia (FA) signaling, a key genomic maintenance pathway, is activated in response to replication stress. Here, we report that phosphorylation of the pivotal pathway protein FANCD2 by CHK1 triggers its FBXL12-dependent proteasomal degradation, facilitating FANCD2 clearance at stalled replication forks. This promotes efficient DNA replication under conditions of CYCLIN E- and drug-induced replication stress. Reconstituting FANCD2-deficient fibroblasts with phosphodegron mutants failed to re-establish fork progression. In the absence of FBXL12, FANCD2 becomes trapped on chromatin, leading to replication stress and excessive DNA damage. In human cancers, FBXL12, CYCLIN E, and FA signaling are positively correlated, and FBXL12 upregulation is linked to reduced survival in patients with high CYCLIN E-expressing breast tumors. Finally, depletion of FBXL12 exacerbated oncogene-induced replication stress and sensitized cancer cells to drug-induced replication stress by WEE1 inhibition. Collectively, our results indicate that FBXL12 constitutes a vulnerability and a potential therapeutic target in CYCLIN E-overexpressing cancers.

Cyclin E/CDK2 and feedback from soluble histone protein regulate the S phase burst of histone biosynthesis.

Faithful DNA replication requires that cells fine-tune their histone pool in coordination with cell-cycle progression. Replication-dependent histone biosynthesis is initiated at a low level upon cell-cycle commitment, followed by a burst at the G1/S transition, but it remains unclear how exactly the cell regulates this burst in histone biosynthesis as DNA replication begins. Here, we use single-cell time-lapse imaging to elucidate the mechanisms by which cells modulate histone production during different phases of the cell cycle. We find that CDK2-mediated phosphorylation of NPAT at the restriction point triggers histone transcription, which results in a burst of histone mRNA precisely at the G1/S phase boundary. Excess soluble histone protein further modulates histone abundance by promoting the degradation of histone mRNA for the duration of S phase. Thus, cells regulate their histone production in strict coordination with cell-cycle progression by two distinct mechanisms acting in concert.

Pers reverse angiotensin II -induced vascular smooth muscle cell proliferation by targeting cyclin E expression via inhibition of the MAPK signaling pathway.

The circadian rhythm of blood pressure (BP) is believed to be regulated by the clock system, which is closely linked to levels of angiotensin II (Ang II). This study aimed to investigate whether Ang II mediates the proliferation of vascular smooth muscle cells (VSMCs) through the interaction between the clock system and the mitogen-activated protein kinase (MAPK) signaling pathway. Primary rat aortic VSMCs were treated with Ang II, with or without MAPK inhibitors. VSMC proliferation, expression of clock genes, CYCLIN E, and MAPK pathways were assessed. Ang II treatment resulted in increased VSMC proliferation and rapid upregulation of clock gene Periods (Pers) expression. Compared to the non-diseased control (NC) group, VSMCs incubated with Ang II displayed a noticeable delay in the G1/S phase transition and downregulation of CYCLIN E upon silencing of Per1 and Per2 genes. Importantly, silencing Per1 or Per2 in VSMCs led to decreased expression of key MAPK pathway proteins, including RAS, phosphorylated mitogen-activated protein kinase (P-MEK), and phosphorylated extracellular signal-regulated protein kinase (P-ERK). Moreover, the MEK and ERK inhibitors, U0126 and SCH772986, significantly attenuated the Ang II-induced proliferation of VSMCs, as evidenced by an increased G1/S phase transition and decreased CYCLIN E expression. The MAPK pathway plays a critical role in regulating VSMC proliferation in response to Ang II stimulation. This regulation is controlled by the expression of circadian clock genes involved in the cell cycle. These findings provide novel insights for further research on diseases associated with abnormal VSMC proliferation.

Therapeutic effects of hAMSCs secretome on proliferation of MDA-MB-231 breast cancer cells by the cell cycle arrest in G1/S phase

Background Cancer refers to a disease resulting from the uncontrolled division and growth of abnormal cells. Among different cancer types, breast cancer is considered as one of the most commonly diagnosed cancers. Herein, we explored the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) secretome on breast cancer cells (MDA-MB-231) through analyzing cell cycle progression. Methods We employed a co-culture system using 6-well Transwell plates and after 72 h, the cell cycle progression was evaluated in the hAMSCs-treated MDA-MB-231 cells through analyzing the expressions of RB, CDK4/6, cyclin D, CDK2, cyclin E, p16/INK4a, p21/WAF1/CIP1, and p27/KIP1 using quantitative real-time PCR (qRT-PCR) and western blot method. Cell proliferation, apoptosis, and cell cycle progression were checked using an MTT assay, DAPI staining, and flow cytometry. Results Our results indicated that elevation of RB, p21/WAF1/CIP1, and p27/KIP1 and suppression of RB hyperphosphorylation, p16/INK4a, cyclin E, cyclin D1, CDK2, and CDK4/6 may contribute to inhibiting the proliferation of hAMSCs-treated MDA-MB-231 cells through cell cycle arrest in G1/S phase followed by apoptosis. Conclusion hAMSCs secretome may be an effective approach on breast cancer therapy through the inhibition of cell cycle progression (AU)

Circular RNA circSEMA5A facilitates colorectal cancer development by regulating microRNA-195-5p to target CCNE1 axis.

Colorectal cancer (CRC) is one of the most prevalent malignancies with a high mortality rate worldwide. Circular RNAs (circRNAs) have lately emerged as key molecules involved in cancer development and metastasis. CircSEMA5 is reported to be oncogenic in some cancers, yet its role in the pathogenesis of CRC remains unknown. Herein, we attempted to investigate the functional role and molecular mechanism of circSEMA5A underlying CRC progression. RT-qPCR and RNase R digestion assays were used to evaluate circSEMA5A expression characteristics in CRC cells. Loss-of-function assays were performed to clarify circSEMA5A role in CRC biological processes. Bioinformatics and mechanism experiments were conducted to assess the association of circSEMA5A or CCNE1 with miR-195-5p in CRC cells. Rescue assays were conducted to explore the regulatory function of circSEMA5A-miR-195-5p-CCNE1 in CRC cellular processes. Through bioinformatics and functional screening, we found that circSEMA5A was highly expressed in CRC cells and was mainly localized in the nucleus. CircSEMA5A promoted CRC proliferative, migratory, and invasive capabilities in cultured cells and facilitated the tumorigenic process in xenografts; however, circSEMA5A silencing repressed tumor metastasis in CRC cells. Mechanistically, circSEMA5A was competitively bound with miR-195-5p to upregulate CCNE1 expression. Moreover, the impact of circSEMA5A knockdown on CRC cell proliferative, migratory, and invasive capabilities was countervailed by miR-195-5p inhibitor or CCNE1 overexpression. To summarize, circSEMA5A is a novel circRNA that serves as an oncogene in CRC progression. CircSEMA5A facilitates CRC cell malignancy and tumor growth through sponging miR-195-5p to upregulate CCNE1, thus providing a new direction for CRC diagnosis and targeted therapy.

Structural basis of CDK3 activation by cyclin E1 and inhibition by dinaciclib.

Cell cycle transitions are controlled by multiple cell cycle regulators, especially CDKs. Several CDKs, including CDK1-4 and CDK6, promote cell cycle progression directly. Among them, CDK3 is critically important because it triggers the transitions of G0 to G1 and G1 to S phase through binding to cyclin C and cyclin E1, respectively. In contrast to its highly related homologs, the molecular basis of CDK3 activation remains elusive due to the lack of structural information of CDK3, particularly in cyclin bound form. Here we report the crystal structure of CDK3 in complex with cyclin E1 at 2.25 Å resolution. CDK3 resembles CDK2 in that both adopt a similar fold and bind cyclin E1 in a similar way. The structural discrepancy between CDK3 and CDK2 may reflect their substrate specificity. Profiling a panel of CDK inhibitors reveals that dinaciclib inhibits CDK3-cyclin E1 potently and specifically. The structure of CDK3-cyclin E1 bound to dinaciclib reveals the inhibitory mechanism. The structural and biochemical results uncover the mechanism of CDK3 activation by cyclin E1 and lays a foundation for structural-based drug design.

Spatiotemporal Epigenetic Control of the Histone Gene Chromatin Landscape during the Cell Cycle.

Higher-order genomic organization supports the activation of histone genes in response to cell cycle regulatory cues that epigenetically mediates stringent control of transcription at the G1/S-phase transition. Histone locus bodies (HLBs) are dynamic, non-membranous, phase-separated nuclear domains where the regulatory machinery for histone gene expression is organized and assembled to support spatiotemporal epigenetic control of histone genes. HLBs provide molecular hubs that support synthesis and processing of DNA replication-dependent histone mRNAs. These regulatory microenvironments support long-range genomic interactions among non-contiguous histone genes within a single topologically associating domain (TAD). HLBs respond to activation of the cyclin E/CDK2/NPAT/HINFP pathway at the G1/S transition. HINFP and its coactivator NPAT form a complex within HLBs that controls histone mRNA transcription to support histone protein synthesis and packaging of newly replicated DNA. Loss of HINFP compromises H4 gene expression and chromatin formation, which may result in DNA damage and impede cell cycle progression. HLBs provide a paradigm for higher-order genomic organization of a subnuclear domain that executes an obligatory cell cycle-controlled function in response to cyclin E/CDK2 signaling. Understanding the coordinately and spatiotemporally organized regulatory programs in focally defined nuclear domains provides insight into molecular infrastructure for responsiveness to cell signaling pathways that mediate biological control of growth, differentiation phenotype, and are compromised in cancer.

RelB represses miR-193a-5p expression to promote the phenotypic transformation of vascular smooth muscle cells in aortic aneurysm.

Aortic aneurysm (AA) is a potentially fatal disease with the possibility of rupture, causing high mortality rates with no effective drugs for the treatment of AA. The mechanism of AA, as well as its therapeutic potential to inhibit aneurysm expansion, has been minimally explored. Small non-coding RNA (miRNAs and miRs) is emerging as a new fundamental regulator of gene expression. This study aimed to explore the role and mechanism of miR-193a-5p in abdominal aortic aneurysms (AAA). In AAA vascular tissue and Angiotensin II (Ang II)-treated vascular smooth muscle cells (VSMCs), the expression of miR-193a-5 was determined using real-time quantitative PCR (RT-qPCR). Western blotting was used to detect the effects of miR-193a-5p on PCNA, CCND1, CCNE1, and CXCR4. To detect the effect of miR-193a-5p on the proliferation and migration of VSMCs, CCK-8, and EdU immunostaining, flow cytometry, wound healing, and Transwell Chamber analysis were performed. In vitro results suggest that overexpression of miR-193a-5p inhibited the proliferation and migration of VSMCs, and its inhibition aggravated their proliferation and migration. In VSMCs, miR-193a-5p mediated proliferation by regulating CCNE1 and CCND1 genes and migration by regulating CXCR4. Further, in the Ang II-induced abdominal aorta of mice, the expression of miR-193a-5p was reduced and significantly downregulated in the serum of patients with aortic aneurysm (AA). In vitro studies confirmed that Ang II-induced downregulation of miR-193a-5p in VSMCs by upregulation of the expression of the transcriptional repressor RelB in the promoter region. This study may provide new intervention targets for the prevention and treatment of AA.

Cyclin E-induced replicative stress drives p53-dependent whole-genome duplication.

Whole-genome duplication (WGD) is a frequent event in cancer evolution and an important driver of aneuploidy. The role of the p53 tumor suppressor in WGD has been enigmatic: p53 can block the proliferation of tetraploid cells, acting as a barrier to WGD, but can also promote mitotic bypass, a key step in WGD via endoreduplication. In wild-type (WT) p53 tumors, WGD is frequently associated with activation of the E2F pathway, especially amplification of CCNE1, encoding cyclin E1. Here, we show that elevated cyclin E1 expression causes replicative stress, which activates ATR- and Chk1-dependent G2 phase arrest. p53, via its downstream target p21, together with Wee1, then inhibits mitotic cyclin-dependent kinase activity sufficiently to activate APC/CCdh1 and promote mitotic bypass. Cyclin E expression suppresses p53-dependent senescence after mitotic bypass, allowing cells to complete endoreduplication. Our results indicate that p53 can contribute to cancer evolution through the promotion of WGD.

Therapeutic effects of hAMSCs secretome on proliferation of MDA-MB-231 breast cancer cells by the cell cycle arrest in G1/S phase.

BACKGROUND: Cancer refers to a disease resulting from the uncontrolled division and growth of abnormal cells. Among different cancer types, breast cancer is considered as one of the most commonly diagnosed cancers. Herein, we explored the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) secretome on breast cancer cells (MDA-MB-231) through analyzing cell cycle progression. METHODS: We employed a co-culture system using 6-well Transwell plates and after 72 h, the cell cycle progression was evaluated in the hAMSCs-treated MDA-MB-231 cells through analyzing the expressions of RB, CDK4/6, cyclin D, CDK2, cyclin E, p16/INK4a, p21/WAF1/CIP1, and p27/KIP1 using quantitative real-time PCR (qRT-PCR) and western blot method. Cell proliferation, apoptosis, and cell cycle progression were checked using an MTT assay, DAPI staining, and flow cytometry. RESULTS: Our results indicated that elevation of RB, p21/WAF1/CIP1, and p27/KIP1 and suppression of RB hyperphosphorylation, p16/INK4a, cyclin E, cyclin D1, CDK2, and CDK4/6 may contribute to inhibiting the proliferation of hAMSCs-treated MDA-MB-231 cells through cell cycle arrest in G1/S phase followed by apoptosis. CONCLUSION: hAMSCs secretome may be an effective approach on breast cancer therapy through the inhibition of cell cycle progression.

Long Non-coding RNA LINC00473 Promotes Breast Cancer Progression via miR-424-5p/CCNE1 Pathway.

BACKGROUND: There has been a large increase in the incidence of breast cancer (BC) among women. LINC00473 is a cancer-related lncRNA, participating in the progression of many cancers, but its role in the progression of BC awaits more elaboration. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to quantify LINC00473, miR-424-5p, and cyclin E1 (CCNE1) mRNA expression levels in BC tissues and cells. Cell counting kit-8 (CCK-8) assay was employed to detect the cell viability; the cell migration and invasion abilities were evaluated by the Transwell assay. Western blot and immunohistochemistry (IHC) were adopted to study CCNE1 protein expression; dual-luciferase reporter assay was performed to clarify the targeting relationships among LINC00473, miR-424-5p, and CCNE1. RESULTS: LINC00473 expression was elevated in BC tissues and cell lines, which was associated with lymph node metastasis and higher clinical stage of the patients with BC. LINC00473 proved to be a molecular sponge for miR-424-5p; LINC00473 knockdown impeded the growth, migration, invasion, and epithelial-mesenchymal transition of BC cells, while these effects were abolished by miR-424-5p inhibitors; miR-424-5p targeted CCNE1 to restrain its expression. LINC00473 positively regulated CCNE1 expression, and CCNE1 restoration counteracted the effects induced by LINC00473 knockdown in BC cells. CONCLUSION: LINC00473 facilitates the progression of BC through miR-424-5p/CCNE1 axis.

Effects of Imatinib Combined With Everolimus on Mouse Pituitary Tumor Cell AtT-20.

Context: Pituitary adenoma is a clinical syndrome in which excessive production of pituitary corticotropin (ACTH). For ACTH tumor cells, researchers know little about the influence of the cell-cycle process on ACTH production and cell proliferation. Some research has shown that imatinib can induce apoptosis of tumor cells. Objective: The study intended to explore the effects and molecular mechanisms of imatinib combined with everolimus on AtT-20 cells in AtT-20 mouse pituitary tumors. Design: The research team performed a laboratory study using murine corticotropin tumor AtT-20 cells. Setting: The study took place at the Department of Neurosurgery at Renmin Hospital of the Hubei University of Medicine in Shiyan, Hubei, China. Intervention: The research team cultured the cells in AtT-20-cell-specific medium containing 100 µg/mL of streptomycin, 100 U/mL of penicillin, and 10% fetal bovine serum at 37°C and 5% CO2. The team divided the cells into a control group, a normal culture without the drug, and an intervention group, incubated for 24 hours with 1 µM of imatinib and 3 µM of everolimus when the cells grew to 40% confluence. Outcome Measures: The research team: (1) determined the effects of the combined drugs on cell viability using a methyl thiazolyl tetrazolium (MTT) assay; (2) detected the cell's mitochondrial membrane potential and LDH leakage using "sytox blue, 5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide," CBIC2(3) or JC-1, and lactate dehydrogenase (LDH) assay kits, respectively; (3) detected AtT-20 cell apoptosis using a "terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labeling" (TUNEL) kit; (4) analyzed the expression of protein kinase B (p-Akt), cAMP-response element binding protein (p-CREB), p27, p53, and cyclin E using a Western blot test; (5) detected the mRNA expression of opioid melanin procorticotropin (POMC)), caspase-3, and pituitary tumor transforming gene 1 (PTTG1) using reverse transcription-polymerase chain reaction (RT-PCR); (6) measure the concentration of adreno-cortico-tropic-hormone (ACTH) in the supernatant using an enzyme-linked immunoassay (ELISA) kit; and (7) assessed the cell cycle distribution using flow cytometry. Results: No differences existed in cell viability between the groups at the baseline (0 h) of the culture period (P > .05). Compared to the control group, the intervention group's: (1) cell viability was significantly lower at 4, 8, and 12 hours and postintervention at 16 hours (P < .001); (2) LDH concentration was significantly higher (P < .001); (3) mitochondrial membrane potential was significantly lower (P < .001); (4) apoptosis rate of TUNEL was significantly higher (P < .001 ); (5) expression of p-Akt, p-CREB phosphorylation, and cyclin E was significantly lower (P < .001), (6) expression of p27 and p53 protein was significantly higher (P < .001); (7) mRNA expression of POMC and PTTG1 were significantly lower (P < .001); (8) mRNA expression of caspase-3 was significantly higher (P < .001); (9) concentration of ACTH was lower (P < .001); and (10) percentage of cells in the G0/G1 phase was significantly higher, while the percentage of cells in the S phase was significantly lower (P < .05). Conclusions: Imatinib combined with everolimus can affect the AtT-20 cell cycle through the signaling pathway of the phosphatidylin-ositol-3-kinase (PI3K)/Akt/ protein kinase A (PKA) system and can inhibit cell proliferation and induce cell apoptosis. Therefore, Imatinib and everolimus may be an effective combination of candidates for drugs for mouse pituitary tumor.

Low-molecular-weight cyclin E deregulates DNA replication and damage repair to promote genomic instability in breast cancer.

Low-molecular-weight cyclin E (LMW-E) is an N-terminus deleted (40 amino acid) form of cyclin E detected in breast cancer, but not in normal cells or tissues. LMW-E overexpression predicts poor survival in breast cancer patients independent of tumor proliferation rate, but the oncogenic mechanism of LMW-E and its unique function(s) independent of full-length cyclin E (FL-cycE) remain unclear. In the current study, we found LMW-E was associated with genomic instability in early-stage breast tumors (n = 725) and promoted genomic instability in human mammary epithelial cells (hMECs). Mechanistically, FL-cycE overexpression inhibited the proliferation of hMECs by replication stress and DNA damage accumulation, but LMW-E facilitated replication stress tolerance by upregulating DNA replication and damage repair. Specifically, LMW-E interacted with chromatin and upregulated the loading of minichromosome maintenance complex proteins (MCMs) in a CDC6 dependent manner and promoted DNA repair in a RAD51- and C17orf53-dependent manner. Targeting the ATR-CHK1-RAD51 pathway with ATR inhibitor (ceralasertib), CHK1 inhibitor (rabusertib), or RAD51 inhibitor (B02) significantly decreased the viability of LMW-E-overexpressing hMECs and breast cancer cells. Collectively, our findings delineate a novel role for LMW-E in tumorigenesis mediated by replication stress tolerance and genomic instability, providing novel therapeutic strategies for LMW-E-overexpressing breast cancers.

Temporal changes in cyclinD-CDK4/CDK6 and cyclinE-CDK2 pathways: implications for the mechanism of deficient decidualization in an immune-based mouse model of unexplained recurrent spontaneous abortion.

BACKGROUND: Deficient endometrial decidualization has been associated with URSA. However, the underlying mechanism is poorly understood. This study aimed to investigate the temporal cytokine changes and the involvement of CyclinD-CDK4/6 and CyclinE-CDK2 pathways in the regulation of the G1 phase of the cell cycle during decidualization in a murine model of URSA. METHODS: Serum and decidual tissues of mice were collected from GD4 to GD8. The embryo resorption and abortion rates were observed on GD8 and the decidual tissue status was assessed. In addition, PRL, Cyclin D, CDK6, CDK4, Cyclin E, CDK2 expression in mice were measured. RESULTS: URSA mice showed high embryo resorption rate and PRL, Cyclin D, Cyclin E CDK2, CDK4, CDK6 down-regulation during decidualization. The hyperactivated Cyclin D-CDK4/CDK6 and cyclin E/CDK2 pathways inhibit the decidualization process and leading to deficient decidualization. CONCLUSION: Insufficient decidualization is an important mechanism of URSA. which is related to the decrease of Cyclin D、Cyclin E、 CDK2、CDK4 and CDK6 in decidualization process of URSA.

Condensed tannins from Ulmus pumila L. leaves induce G2/M phase arrest and apoptosis via caspase-cascade activation in TFK-1 cholangiocarcinoma cells.

Condensed tannins the polyphenolic compounds that are widespread in plants have been proved to have antitumor potential. Here, we purified the bioactive condensed tannins from leaves of Ulmus pumila L. and explored their structural characteristics, antitumor effect on TFK-1 cholangiocarcinoma cells as well as the related potential mechanism. The UV-Vis, FT-IR spectroscopy, ESI-Full-MS, and thiolysis-HPLC-ESI-MS demonstrated that U. pumila condensed tannins (UCTs) consisted essentially of procyanidins with epicatechin as the main flavan-3-ol extension unit. The UCTs could significantly reduce the survival rate of human cholangiocarcinoma TFK-1, SK-CHA-1, and MZ-CHA-1 cells with the better inhibitory effect on TFK-1 cell proliferation. Flow cytometric assay showed that UCTs affected TFK-1 survival by G2/M phase arrest and inducing apoptosis in a dose-dependent manner. In addition, a total of 6592 differentially expressed genes (DEGs), consisting of 94 upregulated and 6498 downregulated DEGs, were identified between untreated and UCTs-treated TFK-1 cells using RNA-seq technology. Enrichment analysis based on the KEGG database revealed that these DEGs were closely associated with cell cycle and p53 apoptotic signaling pathways. Furthermore, qRT-PCR confirmed that treatment of UCTs to TFK-1 cells caused significant changes in the expression of cyclin E, cdc25 A, cytochrome c, caspase-3, and caspase-8. These results indicated that UCTs exhibited the growth inhibition effect on TFK-1 cells possibly via G2/M cell cycle arrest and activation of caspase-cascade to induce apoptosis, and had potential as an anti-cholangiocarcinoma drug for further development. PRACTICAL APPLICATIONS: Ulmus pumila L. as a valuable tree species has been widely used in fields of medicine and food. Condensed tannins, the polyphenolic compounds widespread in plants, have been proved to have antitumor potential and be safe to normal cells. In this study, the condensed tannins from leaves of U. pumila (UCTs) remarkably suppressed cholangiocarcinoma (CCA) cell viability possibly via G2/M cell cycle arrest and activation of caspase-cascade to induce apoptosis. The results provided evidence for the application of UCTs as a potential therapeutic drug for CCA tumor.

DFNA5 inhibits colorectal cancer proliferation by suppressing the mTORC1/2 signaling pathways via upregulation of DEPTOR.

The human deafness, autosomal dominant 5 gene (DFNA5), a newly discovered executor of pyroptosis, has been strongly implicated in the tumorigenesis of several human cancers. However, an understanding of the functional role of DFNA5 in the development and progression of colorectal cancer (CRC) is limited. In this study, we demonstrated that DFNA5 was downregulated in CRC tissues. Ectopic expression of DFNA5 inhibited tumor cell growth in vitro, retarded tumor formation in vivo, and blocked a cell-cycle transition from the G0/G1 to the S phase, whereas a DFNA5 knockdown promoted cell proliferation. Western blotting showed that the levels of cell cycle-related proteins, including cyclin D1, cyclin E, CDK2, and p21, were accordingly altered upon DFNA5 overexpression or DFNA5 knockdown. Mechanistic studies indicated that DFNA5 exerted its tumor suppressor functions by antagonizing mTORC1/2 signaling via upregulation of DEPTOR. In addition, blockage of mTORC1/2 signaling by Torin-1 abolished the accelerative proliferation by DFNA5 knockdown. In conclusion, these results indicated that DFNA5 inhibits the proliferation and tumor formation of colon cancer cells by suppressing mTORC1/2 signaling.