Cell Differentiation Protein Research Articles

Identifying Mcl-1 protein dependencies using dimerization-specific antibody biomarker for predicting response to targeted apoptosis inducing therapies.

7038 Background: The anti-apoptotic Bcl-2 family proteins facilitate pro-survival and resistance to anti-cancer therapies. Measuring the function of these proteins has shown utility in predicting response to therapy. A method that expands the principle of BH3 profiling to solid tumors is presented. Measuring the occurrence of heterodimers of Myeloid Leukemia Cell Differentiation Protein (Mcl-1), and pro-apoptotic binding protein Bim is an indicator of cancer cell apoptotic priming state. The readout of Mcl-1 containing complex-specific biomarkers can identify survival dependencies in cancer cells potentially providing clinical utility in guiding cancer treatments. Methods: Engineered immunogens that recapitulate conformation-specific epitopes induced during binding of the Mcl-1/Bim protein complex were used to generate. monoclonal antibodies. One Heterodimer Specific Mcl-1 Bim (HsMcB) was chosen. The selective binding was confirmed using ELISA, fluorescence polarization, immunofluorescence microscopy and flow cytometry in Bim or Mcl-1 knockdown cells, and by immunohistochemistry in formalin-fixed paraffin-embedded patient tissue. Correlation of HsMcB measurements to BH3 profiling readouts from the Mcl-1 restricted Noxa peptide was explored. Results: HsMcB signal depends on both Mcl-1 and Bim protein levels. The ratio of the HsMcB to unbound protein (Mcl-1) signal ([HsMcB]/[Mcl-1]) was measured. Disruption of the complexes by BH3 mimetics targeted to Mcl-1 and depletion of Mcl-1 level using CDK9 inhibitors diminished the [HsMcB]/[Mcl-1] readout. We see correlations between these readouts and BH3 mimetic peptides readouts from the Mcl-1 specific Noxa and MS1 BH3 mimetic peptides in AML patient samples that have been treated with Mcl-1 inhibitors. Conclusions: Mcl-1 dependence is a predictive biomarker for venetoclax resistance and for response to Mcl-1 targeted therapies. Flow cytometric and IHC based measurements of a heterodimer complex offer a direct and simpler approach that harbors potential for use in clinical settings. Additional antibodies targeting Mcl-1/Bak and Mcl-1 Noxa complexes are being tested.

4-hydroxy-3-methoxy cinnamic acid accelerate myoblasts differentiation on C2C12 mouse skeletal muscle cells via AKT and ERK 1/2 activation

BackgroundThe dietary intake of plant-based supplements has a vital role in human health and development. However, the actions of secondary plant metabolites on cell growth, differentiation and their signaling mechanisms are still unclear. PurposeIn this study, we aim to investigate the C2C12 myoblast cells proliferation and differentiation by 4-hydroxy-3-methoxy cinnamic acid (=HMCA, ferulic acid) in a dose-dependent manner and to reveal its underlying mechanism of action. Methods: The effect of HMCA on C2C12 cell proliferation and differentiation were evaluated by expression of BMP's marker genes (-2, -4, -6, -7) and related myogenic proteins were analyzed by quantitative PCR and western blot techniques, respectively. ResultsThe in vitro findings confirmed that the HMCA upregulates BMPs (including BMP-2, -4, -6, and-7), gene expression in C2C12 skeletal muscle cells. Exposure to the lower dose of HMCA caused a significantly greater induction of myogenic differentiation than the higher dose during three- and six-day treatments. Further, the C2C12 myogenic differentiation signaling proteins MyoD, myogenin, JAK-1, -2, -3, STAT -2, -3, AMPK-α, ERK(1/2), and AKT were more preferentially activated by HMCA exposure cells than by untreated models. Thus, the experiment with inhibitors revealed that the HMCA induced muscle cell proliferation and differentiation through AKT and ERK (1/2) signaling cascades. Also, HMCA enhanced the C2C12 muscle cell differentiation protein markers such as myogenin, AKT and ERK (1/2) significantly (p ≤ 0.05) at day three in chemical inhibitors of LY 294002 and PD98056 treated samples. ConclusionThe HMCA has a significant effect on muscle cell differentiation through ERK(1/2) and AKT signaling activation. Also, the HMCA promotes C2C12 muscle cell proliferation and differentiation via activation of osteogenic genes and myogeneic protein markers. Therefore, this study suggests that the natural phenolic compound HMCA has a potent function in muscle cell proliferation, differentiation, and development.

Downregulation of protein disulfide‑isomerase A3 expression inhibits cell proliferation and induces apoptosis through STAT3 signaling in hepatocellular carcinoma.

Protein disulfide‑isomeraseA3 (PDIA3) is a chaperone protein that modulates folding of newly synthesized glycoproteins and responds to endoplasmic reticulum (ER) stress. Previous studies reported that increased expression of PDIA3 in hepatocellular carcinoma (HCC) is a marker for poor prognosis. However, the mechanism remains poorly understood. The aim of the present study, therefore, was to understand the role of PDIA3 in HCC development. First, immunohistochemical staining of tissues from 53HCC cases revealed that HCC tissues with high PDIA3 expression exhibited a higher proliferation index and contained fewer apoptotic cells than those with low expression. In addition, the knockdown of PDIA3 significantly inhibited cell proliferation and induced apoptosis in HCC cell lines. These results suggest that PDIA3 regulates cell proliferation and apoptosis in HCC. An examination of whether PDIA3 knockdown induced apoptosis through ER stress revealed that PDIA3 knockdown did not increase ER stress marker, 78kDa glucose‑regulated protein, in HCC cell lines. Furthermore, the association between PDIA3 and the signal transducer and activator of transcription3 (STAT3) signaling pathway were investigated invitro and invivo. Immunofluorescence staining and co‑immunoprecipitation experiments revealed colocalization and binding, respectively, of PDIA3 and STAT3 in HCC cell lines. The knockdown of PDIA3 decreased the levels of phosphorylated STAT3 (P‑STAT3; Tyr705) and downstream proteins of the STAT3 signaling pathway: The anti‑apoptotic proteins (Bcl‑2‑like protein 1, induced myeloid leukemia cell differentiation protein Mcl‑1, survivin and X‑linked inhibitor of apoptosis protein). In addition, PDIA3 knockdown provided little inhibitory effect on cell proliferation in HCC cell lines treated with AG490, a tyrosine‑protein kinase JAK/STAT3 signaling inhibitor. Finally, an association was demonstrated between PDIA3 and P‑STAT3 expression following immunostaining of 35 HCC samples. Together, the present data suggest that PDIA3 promotes HCC progression through the STAT3 signaling pathway.

Association of Leukemia Target Genes Tet2, Bcl2, and Slc23a2 in Vitamin C Pathways.

Vitamin C has been used in combination with several target genes in the treatment of leukemia. Tet methylcytosine dioxygenase (Tet2), B-cell lymphoma 2 (Bcl2), and solute carrier family 23 member 2 (Slc23a2) are the major target genes in the treatment of leukemia and are relevant to vitamin C. Using whole-genome expression profiles from mouse livers, the expression quantitative trait locus (eQTL), correlation matrix, and gene network graph were constructed with probes from each of these three genes and with their relative genes. The function of key genes was examined by their pathways and reported information. The results indicated that although direct correlations among their expression levels were not strong, alternative connecting pathways were discovered. By comparing the expression levels of one probe with known sequences from each of the three genes, we identified several key genes, induced myeloid leukemia cell differentiation protein (Mcl1), far upstream element-binding protein 1 (Fubp1), and tumor protein D52-like 2 (Tpd52l2), which play important roles in acute lymphocytic leukemia and acute myelocytic leukemia. In conclusion, Alternative pathways and key genes that connect Tet2, Bcl2, and Slc23a2 for their therapeutic applications with vitamin C were identified.

A Novel MCL1 Inhibitor Combined with Venetoclax Rescues Venetoclax-Resistant Acute Myelogenous Leukemia.

Suppression of apoptosis by expression of antiapoptotic BCL2 family members is a hallmark of acute myeloblastic leukemia (AML). Induced myeloid leukemia cell differentiation protein (MCL1), an antiapoptotic BCL2 family member, is commonly upregulated in AML cells and is often a primary mode of resistance to treatment with the BCL2 inhibitor venetoclax. Here, we describe VU661013, a novel, potent, selective MCL1 inhibitor that destabilizes BIM/MCL1 association, leads to apoptosis in AML, and is active in venetoclax-resistant cells and patient-derived xenografts. In addition, VU661013 was safely combined with venetoclax for synergy in murine models of AML. Importantly, BH3 profiling of patient samples and drug-sensitivity testing ex vivo accurately predicted cellular responses to selective inhibitors of MCL1 or BCL2 and showed benefit of the combination. Taken together, these data suggest a strategy of rationally using BCL2 and MCL1 inhibitors in sequence or in combination in AML clinical trials. SIGNIFICANCE: Targeting antiapoptotic proteins in AML is a key therapeutic strategy, and MCL1 is a critical antiapoptotic oncoprotein. Armed with novel MCL1 inhibitors and the potent BCL2 inhibitor venetoclax, it may be possible to selectively induce apoptosis by combining or thoughtfully sequencing these inhibitors based on a rational evaluation of AML.See related commentary by Leber et al., p. 1511.This article is highlighted in the In This Issue feature, p. 1494.

Pemetrexed exerts anticancer effects by inducing G0/G1‑phase cell cycle arrest and activating the NOXA/Mcl‑1 axis in human esophageal squamous cell carcinoma cells

Esophageal squamous cell carcinoma (ESCC) is a dominant histological subtype of esophageal cancer with notably high incidence and mortality rates. Pemetrexed is a clinical antifolate therapeutic agent with anticancer properties. The present study aimed to understand whether pemetrexed is able to exert anticancer effects on ESCC cells, and to determine the underlying molecular mechanism. ESCC cells were treated with pemetrexed and cell survival was assessed with MTT assays. The cell cycle and apoptosis were evaluated using flow cytometry analysis, and proteins were detected using western blotting. It was demonstrated that pemetrexed inhibited cell survival and induced G0/G1 cell cycle arrest and apoptosis in human ESCC cells. Furthermore, the results demonstrated that the phorbol-12-myristate-13-acetate-induced protein 1/induced myeloid leukemia cell differentiation protein Mcl-1 axis is involved in intrinsic apoptosis induced by pemetrexed. The protein expression of endoplasmic reticulum stress markers inositol-requiring enzyme 1α, binding immunoglobulin protein and CCAAT-enhancer-binding protein homologous protein were upregulated following treatment with pemetrexed. These results suggest that pemetrexed may induce an endoplasmic reticulum stress response while activating intrinsic apoptosis. The present study provided important mechanistic insights into potential cancer treatments involving pemetrexed and enhanced the understanding of human ESCC.

Phoyunnanin E Induces Apoptosis of Non-small Cell Lung Cancer Cells via p53 Activation and Down-regulation of Survivin

Lung cancer is by far the most common cause of cancer mortality, accounting for nearly 20% of all global cancer deaths. Therefore, potent and effective compounds for treatment of this cancer type are essential. Phoyunnanin E, isolated from Dendrobium venustum (Orchidaceae), has promising pharmacological activities; however, it is unknown if phoyunnanin E affects apoptosis of lung cancer cells. The apoptosis-inducing activity of phoyunnanin E on H460 lung cancer cells was investigated by Hoechst 33342, and annexin V-fluorescein isothiocyanate/propidium iodide staining. The underlying mechanism was determined via monitoring apoptosis-regulatory proteins by western blot analysis. The apoptotic effect of the compound was confirmed in H23 lung cancer cells. Phoyunnanin E significantly induced apoptotic cell death of H460 lung cancer cells, as indicated by condensed and fragmented nuclei with the activation of caspase-3 and -9 and poly (ADP-ribose) polymerase cleavage. Phoyunnanin E mediated apoptosis via a p53-dependent pathway by increasing the accumulation of cellular p53 protein. As a consequence, anti-apoptotic proteins including induced myeloid leukemia cell differentiation protein (MCL1) and B-cell lymphoma 2 (BCL2) were found to be significantly depleted, while pro-apoptotic BCL-2-associated X protein (BAX) protein was up-regulated. Furthermore, it was found that expression of an inhibitor of apoptosis, survivin, markedly reduced in response to phoyunnanin E treatment. The apoptosis-inducting effect was also found in phoyunnanin E-treated H23 lung cancer cells. These results indicate the promising effect of phoyunnanin E in induction of apoptosis, that may be useful for the development of novel anticancer agents.

Low‐molecular‐mass hyaluronan induces pulmonary inflammation by up‐regulation of Mcl‐1 to inhibit neutrophil apoptosis via PI3K/Akt1 pathway

Although low-molecular-mass hyaluronan (LMMHA) has been implicated in pulmonary inflammatory diseases, the signalling pathway of LMMHA (200 000 molecular weight) that initiates the inflammatory response in lung is still unknown. In this study, we evaluate the role of phosphoinositide 3-kinase (PI3K) and its downstream signalling pathway in LMMHA-induced lung inflammatory responses. Our results indicate that pharmacological inhibition of PI3K or genetic deletion of Akt1 enhances neutrophil apoptosis, attenuates neutrophil influx into the lungs of mice and diminishes the expression of pro-inflammatory factors such as interleukin-6, keratinocyte cell-derived chemokine and pro-matrix metalloproteinase-9 in bronchoalveolar lavage fluid after intratracheal administration of LMMHA. More importantly, we found that PI3K/Akt1 participates in LMMHA-induced inflammatory responses, which are mainly mediated by the myeloid leukaemia cell differentiation protein (Mcl-1). Our study suggests that LMMHA induced significantly increased levels of inflammatory factors in bronchoalveolar lavage fluid and activation of the PI3K/Akt1 pathway, which up-regulates the expression of the anti-apoptotic protein Mcl-1 and inhibits the activation of caspase-3, thereby suppressing neutrophil apoptosis to trigger lung inflammation. These findings reveal a novel molecular mechanism underlying sterile inflammation and provides a new potential target for the treatment of pulmonary disease.

Plumbagin, a natural naphthoquinone, inhibits the growth of esophageal squamous cell carcinoma cells through inactivation of STAT3.

Although plumbagin, a natural naphthoquinone, has exhibited antiproliferative activity in numerous types of cancer, its anticancer potential in esophageal squamous cell carcinoma (ESCC) remains unclear. In the present study, the effect of plumbagin on the growth of ESCC cells was investigated in vitro and invivo. ESCC cells were treated with plumbagin and tested for cell cycle distribution and apoptosis. The involvement of STAT3 signaling in the effect of plumbagin was examined. The results demonstrated that plumbagin treatment suppressed ESCC cell viability and proliferation, yet normal esophageal epithelial cell viability was not affected. Plumbagin treatment increased the proportion of cells in the G0/G1 phase of the cell cycle and decreased the proportion of cells in the S phase. Furthermore, plumbagin‑treated ESCC cells displayed a significantly greater % of apoptotic cells. Western blot analysis confirmed that plumbagin upregulated tumor protein p53 and cyclin‑dependent kinase inhibitor1A (also known as p21), while it downregulated cyclinD1, cyclin‑dependent kinase4, and induced myeloid leukemia cell differentiation protein Mcl‑1. Mechanistically, plumbagin inhibited STAT3 activation, and overexpression of constitutively active STAT3 reversed the plumbagin‑mediated growth suppression in ESCC cells. Invivo studies demonstrated that plumbagin delayed the growth of ESCC xenograft tumors and reduced STAT3 phosphorylation. Overall, plumbagin was demonstrated to target STAT3 signaling and to inhibit the growth of ESCC cells both invitro and invivo, suggesting that it may represent a potential anticancer agent for ESCC.

Selective targeting of antiapoptotic BCL-2 proteins in cancer.

Circumvention of apoptotic machinery is one of the distinctive properties of carcinogenesis. Extensively established key effectors of such apoptotic bypass mechanisms, the antiapoptotic BCL-2 (apoptosis regulator BCL-2) proteins, determine the response of cancer cells to chemotherapeutics. Within this background, research and development of antiapoptotic BCL-2 inhibitors were considered to have a tremendous amount of potential toward the discovery of novel pharmacological modulators in cancer. In this review, milestone achievements in the development of selective antiapoptotic BCL-2 proteins inhibitors for BCL-2, BCL-XL (BCL-2-like protein 1), and MCL-1 (induced myeloid leukemia cell differentiation protein MCL-1) were summarized and their future implications were discussed. In the first section, the design and development of BCL-2/BCL-XL dual inhibitor navitoclax, as well as the recent advances and clinical experience with selective BCL-2 inhibitor venetoclax, were synopsized. Preclinical data from selective BCL-XL inhibitors, which are currently undergoing extensive testing as a single agent or in combination with other therapeutic agents, were further summarized. In the second section, MCL-1 inhibitors developed as potential anticancer agents were reviewed regarding their specificity toward MCL-1. Explicitly, studies leading to the identification of MCL-1, nonselective and selective targeting of MCL-1, and recently initiated clinical trials were compiled in chronological order. Based on these concepts, future directions were further discussed for increasing selectivity in the design of prosurvival BCL-2 member inhibitors.

Combined treatment with sorafenib and silibinin synergistically targets both HCC cells and cancer stem cells by enhanced inhibition of the phosphorylation of STAT3/ERK/AKT

Silibinin, a nontoxic bioactive component in milk thistle, is used as a liver-protective drug in the clinic mainly because of its antioxidant and anti-inflammation activities. In this study, we studied the cytotoxic effects of silibinin combined with sorafenib on hepatocellular carcinoma (HCC). The results indicated that silibinin combined with sorafenib potently inhibited the proliferation of various HCC cells and induced significant apoptosis. In an HCC subcutaneous transplantation tumor model, the combination of silibinin and sorafenib significantly suppressed tumor growth compared with monotherapy. As determined by fluorescence staining and Western blots, the combination of the two drugs inhibited the phosphorylation of RAC-alpha serine/threonine-protein kinase (AKT) and signal transducer and activator of transcription 3 (STAT3) together with the expression of antiapoptotic proteins including myeloid leukemia cell differentiation protein Mcl-1 (Mcl-1) and apoptosis regulator Bcl-2 (Bcl-2), resulting in the death of cancer cells. We also found that the combination inhibited the formation and self-renewal of HCC stem cells by down-regulating the expression of stemness-related proteins, such as Homeobox protein NANOG (Nanog) and Krueppel-like factor 4 (Klf4). These results suggested that silibinin improved the efficacy of sorafenib in HCC therapy, indicating a clinical promising therapeutic strategy for HCC patients.

Apoptosis induced by 9,11‑dehydroergosterol peroxide from GanodermaLucidum mycelium in human malignant melanoma cells is Mcl‑1 dependent.

9,11-Dehydroergosterol peroxide [9(11)-DHEP] is an important steroid from medicinal mushroom, which has been reported to exert antitumor activity in several tumor types. However, the role of 9(11)-DHEP toward the malignant melanoma cells has not been investigated. In the present study, the steroid from Ganoderma lucidum was purified on a submerged culture, and its antitumor mechanisms on A375 human malignant melanoma cells was investigated by MTT, flow cytometry and western blotting. The studies demonstrated that apoptotic mechanisms of the steroid were caspase-dependent and mediated via the mitochondrial pathway. The steroid did not cause significant changes in the expression levels of B-cell lymphoma 2 (Bcl-2) family proteins, Bcl-2-like protein 11, p53 upregulated modulator of apoptosis, Bcl-2-associated X protein, BH3 interacting-domain death agonist, Bcl-2-associated death promoter and Bcl-2, but it significantly downregulated induced myeloid leukemia cell differentiation protein Mcl-1 (Mcl-1) in melanoma cells, suggesting the key role of Mcl-1 in regulating apoptosis of melanoma cells induced by the steroid. These properties of 9(11)-DHEP advocate its usage as supplements in human malignant melanoma chemoprevention. The present study also suggests that mycelium of G. lucidum has a potential for producing bioactive substances and extracts with applications in medicine.

SPAG6 silencing induces apoptosis in the myelodysplastic syndrome cell line SKM‑1 via the PTEN/PI3K/AKT signaling pathway in vitro and in vivo.

Apoptosis is a multi-step mechanism of cell self‑destruction for maintaining cellular homeostatic balance. Accumulating evidence indicates that abnormal apoptosis promotes the evolution and progression of myelodysplastic syndromes (MDS). As a novel cancer-testis antigen, sperm‑associated antigen6 (SPAG6) has been reported to regulate apoptosis through the tumor necrosis factor-related apoptosis-inducing ligand signaling pathway in the MDS cell line SKM‑1. However, the mechanism of the intrinsic cell death pathway for apoptosis induction by SPAG6 silencing is unclear. In the present study, the invitro effects of SPAG6 silencing were investigated in SKM‑1 cells through extensive biochemical and molecular approaches. Western blotting and reverse transcription-quantitative polymerase chain reaction were used to detect the expression of SPAG6 and activation of PTEN/PI3K/AKT signal pathway. Additionally, SKM‑1 cells transduced with SPAG6 short hairpin RNA (shRNA) lentivirus were treated with the phosphatidylionositol 3-kinase (PI3K) inhibitor LY294002 or pan caspase inhibitor z‑VAD‑fmk and the apoptosis rates were measured by flow cytometry, and the expressions of associated proteins were examined by western blot analysis. A mouse xenograft model was also used to further evaluate the effects of SPAG6 knockdown on inducing tumor apoptosis invivo. Lentivirus-mediated knockdown of SPAG6 in SKM‑1 cells increased phosphatase and tensin homolog (PTEN) expression and reduced protein kinaseB (AKT) phosphorylation, which in turn resulted in cell apoptosis as evidenced by induced myeloid leukaemia cell differentiation protein Mcl‑1 downregulation, cytochromec release and increased caspase‑9 expression. Consistently, the PI3K inhibitor LY294002 synergistically enhanced apoptosis of SKM‑1 cells when co-administered with SPAG6 shRNA lentivirus. Furthermore, treatment with the pan caspase inhibitor z‑VAD‑fmk failed to prevent PTEN activation upon SPAG6 knockdown, suggesting that SPAG6-regulated PTEN expression was caspase activation-independent. In addition, SPAG6 knockdown was associated with DNMT1 downregulation, implying that SPAG6 may indirectly control PTEN expression via DNA methylation. Furthermore, tumor tissues from nonobese diabetic/severe combined immunodeficient mice inoculated with SPAG6-shRNA lentivirus pre-infected SKM‑1 cells exhibited significantly elevated apoptosis in the extrinsic and intrinsic pathways. These results demonstrate that SPAG6 silencing induces PTEN expression to regulate apoptosis though the PI3K/AKT pathway, indicating that SPAG6 may be a potential therapeutic target for MDS.

Secoisolariciresinol diglucoside prevents the oxidative stress-induced apoptosis of myocardial cells through activation of the JAK2/STAT3 signaling pathway.

Myocardial cell apoptosis mediated by oxidative stress has previously been identified as a key process in ischemic heart disease. Secoisolariciresinol diglucoside(SDG), a polyphenolic plant lignan primarily found in flaxseed, has been demonstrated to effectively protect myocardial cells from apoptosis. In the present study, the role of the Janus kinase2(JAK2)/signal transducer and activator of transcription3(STAT3) was investigated in mediating the protective effect of SDG. Findings of the present study revealed that treatment with H2O2 reduced cell viability and induced apoptosis in H9C2 rat cardiomyocytes. However, SDG was able to reduce the effect of H2O2 in a dose‑dependent manner. H2O2 reduced the expression level of phosphorylated STAT3 and inhibited the levels of B‑cell lymphoma‑extra‑large and induced myeloid leukemia cell differentiation protein, which are the STAT3 target genes. Conversely, SDG rescued phosphorylation of STAT3 and increased the levels of STAT3 target genes. Treatment with SDG alone led to a dose‑dependent increased phosphorylation of JAK2 and STAT3, without activating Src. Furthermore, the anti‑apoptotic effects of SDG were partially abolished by a JAK2/STAT3 inhibitor. In addition, molecular docking revealed that SDG may bind to the protein kinase domain of JAK2, at a binding energy of ‑8.258kcal/mol. Molecular dynamics simulations revealed that JAK2‑SDG binding was stable. In conclusion, activation of the JAK2/STAT3 signaling pathway contributed to the anti‑apoptotic activity of SDG, which may be a potential JAK2 activator.

Identification of novel prognosis-related genes associated with cancer using integrative network analysis

Prognosis identifies the seriousness and the chances of survival of a cancer patient. However, it remains a challenge to identify the key cancer genes in prognostic studies. In this study, we collected 2064 genes that were related to prognostic studies by using gene expression measurements curated from published literatures. Among them, 1820 genes were associated with copy number variations (CNVs). The further functional enrichment on 889 genes with frequent copy number gains (CNGs) revealed that these genes were significantly associated with cancer pathways including regulation of cell cycle, cell differentiation and mitogen-activated protein kinase (MAPK) cascade. We further conducted integrative analyses of CNV and their target genes expression using the data from matched tumour samples of The Cancer Genome Atlas (TCGA). Ultimately, 95 key prognosis-related genes were extracted, with concordant CNG events and increased up-regulation in at least 300 tumour samples. These genes, and the number of samples in which they were found, included: ACTL6A (399), ATP6V1C1 (425), EBAG9 (412), FADD (308), MTDH (377), and SENP5 (304). This study provides the first observation of CNV in prognosis-related genes across pan-cancer. The systematic concordance between CNG and up-regulation of gene expression in these novel prognosis-related genes may indicate their prognostic significance.

Anthelmintic pyrvinium pamoate blocks Wnt/β-catenin and induces apoptosis in multiple myeloma cells.

Multiple myeloma (MM) is a malignancy of the bone marrow. The median survival time of patients with MM is only 5 years, with patients frequently experiencing relapse. Currently, there is no effective therapy for recurrent MM. The results of the present study indicated that pyrvinium pamoate (PP), a US Food and Drug Administration-approved oral anthelmintic drug, exhibited potent antitumor activity in MM cells in vitro. It is demonstrated that PP inhibited MM cell proliferation and mediated apoptosis. Notably, PP markedly promoted the degradation of β-catenin and abrogated its phosphorylation. PP triggered apoptosis in MM cells by inducing the release of cytochrome c and downregulating the expression of myeloid leukemia cell differentiation protein. In addition, PP effectively induced cell death in primary MM cells. In conclusion, PP may be a promising agent for the clinical treatment of MM.

Mucin 1 promotes radioresistance in hepatocellular carcinoma cells through activation of JAK2/STAT3 signaling.

Mucin 1 (MUC1) is aberrantly overexpressed in numerous human cancer types, including hepatocellular carcinoma (HCC) and contributes to chemoresistance of tumor cells. The aim of the present study was to evaluate the possible implication of MUC1 in radioresistance of HCC cells and the underlying mechanisms. It was demonstrated that MUC1 was significantly upregulated in HCC cells following irradiation exposure, which was coupled with increased phosphorylation of signal transducer and activator of transcription 3 (STAT3). Enforced expression of MUC1 significantly (P<0.05) promoted the clonogenic survival of HCC cells following irradiation compared with empty vector-transfected cells. MUC1 overexpression resulted in >60% reduction in apoptosis induced by irradiation, as determined by Annexin-V/propidium iodide double staining and flow cytometry analysis. Furthermore, overexpression of MUC1 significantly (P<0.05) attenuated the activation of caspase-3 and poly (ADP-ribose) polymerase in response to irradiation exposure. Mechanistically, MUC1 inhibited irradiation-induced apoptosis through activation of janus kinase 2 (JAK2) and STAT3, and induction of anti-apoptotic proteins induced myeloid leukemia cell differentiation protein Mcl-1 (Mcl-1) and BCL2 like 1 (Bcl-xL). Small hairpin RNA-mediated knockdown of STAT3 or MUC1 resensitized MUC1-overexpressing cells to irradiation-induced apoptosis, which was accompanied by reduced expression of Bcl-xL and Mcl-1. Collectively, MUC1 contributes to radioresistance of HCC cells likely through activation of the JAK2/STAT3 signaling pathway and thus represents a potential target for improving radiotherapy against HCC.

The Circular RNA Landscape of Periodontal Ligament Stem Cells During Osteogenesis.

The present study aims to investigate the distinct expression pattern of circular RNAs (circRNAs) in periodontal ligament stem cells (PDLSCs) during osteogenesis. PDLSCs were isolated and cultured in osteogenic medium. Total RNA was extracted from cells at day 0 (D0), day 3 (D3), day 7 (D7), and day 14 (D14) and submitted to RNA-sequencing to detect expression profiles of circRNAs, messenger RNAs (mRNAs), and microRNAs (miRNAs). Real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was performed to validate expression of circRNAs and miRNAs. Differential expression analysis and gene ontology analysis were performed. A circRNA-miRNA-mRNA network was constructed to reveal the potential regulatory role of circRNAs. A total of 12,693 circRNA transcripts were detected, and circRNAs displayed stage-specific expression. Expression of four well-known circRNAs was validated by qRT-PCR. In total, 118 circRNAs were differentially expressed at D3, 128 circRNAs were differentially expressed at D7, and 139 circRNAs were differentially expressed at D14 compared with D0. Host genes of differentially expressed circRNAs were enriched in cytoplasmic or membrane-bound vesicles and extracellular matrix, indicating their potential roles in modulating biogenesis of extracellular vesicles. Moreover, mRNAs that were potentially regulated by circRNAs were enriched in bone-formation-associated processes, including extracellular matrix organization, cell differentiation, and bone morphogenetic protein signaling pathway. Expression profiles of circRNAs were significantly altered during osteogenic differentiation of PDLSCs, providing a clue for future studies on the role of circRNAs in osteoblast differentiation.

LCMR1 interacts with DEK to suppress apoptosis in lung cancer cells.

To win the war against lung cancer, the molecular mechanisms underlying its oncogenesis and metastasis must be identified in order to develop novel diagnosis and treatment strategies. We previously identified a novel gene, namely lung cancer metastasis related protein 1 (LCMR1; GenBank accession no.AY148462), which was demonstrated to be overexpressed in non‑small‑cell lung cancer. LCMR1 expression was significantly associated with clinical stage. To further understand the mechanism of LCMR1 in lung cancer, the present study screened a cDNA library from the lung cancer cell line 95D for proteins interacting with LCMR1 by yeast two‑hybrid assay, and the protein DEK was identified. Co‑immunoprecipitation and glutathione S‑transferase pull‑down assays were performed to confirm the interaction between LCMR1 and DEK in vivo and invitro. The results demonstrated that the interaction was mediated primarily by the N‑terminal region of DEK, suggesting that LCMR1 may be involved in the regulation of cell apoptosis. Using RNA interference, DEK and LCMR1 were demonstrated to cooperate in the inhibition of apoptosis in lung cancer cells, and this effect was associated with the induced myeloid leukemia protein cell differentiation protein 1 pathway. The present findings suggest that LCMR1 might serve as a potential molecular target for lung cancer therapy.

Impaired Mitochondrial Microbicidal Responses in Chronic Obstructive Pulmonary Disease Macrophages.

Chronic obstructive pulmonary disease (COPD) is characterized by impaired clearance of pulmonary bacteria. The effect of COPD on alveolar macrophage (AM) microbicidal responses was investigated. AMs were obtained from bronchoalveolar lavage from healthy donors or patients with COPD and challenged with opsonized serotype 14 Streptococcus pneumoniae. Cells were assessed for apoptosis, bactericidal activity, and mitochondrial reactive oxygen species (mROS) production. A transgenic mouse line in which the CD68 promoter ensures macrophage-specific expression of human induced myeloid leukemia cell differentiation protein Mcl-1 (CD68.hMcl-1) was used to model the molecular aspects of COPD. COPD AMs had elevated levels of Mcl-1, an antiapoptotic B-cell lymphoma 2 family member, with selective reduction of delayed intracellular bacterial killing. CD68.hMcl-1 AMs phenocopied the microbicidal defect because transgenic mice demonstrated impaired clearance of pulmonary bacteria and increased neutrophilic inflammation. Murine bone marrow-derived macrophages and human monocyte-derived macrophages generated mROS in response to pneumococci, which colocalized with bacteria and phagolysosomes to enhance bacterial killing. The Mcl-1 transgene increased oxygen consumption rates and mROS expression in mock-infected bone marrow-derived macrophages but reduced caspase-dependent mROS production after pneumococcal challenge. COPD AMs also increased basal mROS expression, but they failed to increase production after pneumococcal challenge, in keeping with reduced intracellular bacterial killing. The defect in COPD AM intracellular killing was associated with a reduced ratio of mROS/superoxide dismutase 2. Up-regulation of Mcl-1 and chronic adaption to oxidative stress alter mitochondrial metabolism and microbicidal function, reducing the delayed phase of intracellular bacterial clearance in COPD.