Exerts Growth Inhibitory Effects Research Articles

Morusin suppresses the stemness characteristics of gastric cancer cells induced by hypoxic microenvironment through inhibition of HIF-1α accumulation

Gastric cancer (GC) is a common, life-threatening malignancy that contributes to the global burden of cancer-related mortality, as conventional therapeutic modalities show limited effects on GC. Hence, it is critical to develop novel agents for GC therapy. Morusin, a typical prenylated flavonoid, possesses antitumor effects against various cancers. The present study aimed to demonstrate the inhibitory effect and mechanism of morusin on the stemness characteristics of human GC in vitro under hypoxia and to explore the potential molecular mechanisms. The effects of morusin on cell proliferation and cancer stem cell-like properties of the human GC cell lines SNU-1 and AGS were assessed by MTT assay, colony formation test, qRT-PCR, flow cytometry analysis, and sphere formation test under hypoxia or normoxia condition through in vitro assays. The potential molecular mechanisms underlying the effects of morusin on the stem-cell-like properties of human GC cells in vitro were investigated by qRT-PCR, western blotting assay, and immunofluorescence assay by evaluating the nuclear translocation and expression level of hypoxia-inducible factor-1α (HIF-1α). The results showed that morusin exerted growth inhibitory effects on SNU-1 and AGS cells under hypoxia in vitro. Moreover, the proportions of CD44+/CD24– cells and the sphere formation ability of SNU-1 and AGS reduced in a dose-dependent manner following morusin treatment. The expression levels of stem cell-related genes, namely Nanog, OCT4, SOX2, and HIF-1α, gradually decreased, and the nuclear translocation of the HIF-1α protein was apparently attenuated. HIF-1α overexpression partially reversed the abovementioned effects of morusin. Taken together, morusin could restrain stemness characteristics of GC cells by inhibiting HIF-1α accumulation and nuclear translocation and could serve as a promising compound for GC treatment.

UPR-Induced miR-616 Inhibits Human Breast Cancer Cell Growth and Migration by Targeting c-MYC.

C/EBP homologous protein (CHOP), also known as growth arrest and DNA damage-inducible protein 153 (GADD153), belongs to the CCAAT/enhancer-binding protein (C/EBP) family. CHOP expression is induced by unfolded protein response (UPR), and sustained CHOP activation acts as a pivotal trigger for ER stress-induced apoptosis. MicroRNA-616 is located within an intron of the CHOP gene. However, the regulation of miR-616 expression during UPR and its function in breast cancer is not clearly understood. Here we show that the expression of miR-616 and CHOP (host gene of miR-616) is downregulated in human breast cancer. Both miR-5p/-3p arms of miR-616 are expressed with levels of the 5p arm higher than the 3p arm. During conditions of ER stress, the expression of miR-616-5p and miR-616-3p arms was concordantly increased primarily through the PERK pathway. Our results show that ectopic expression of miR-616 significantly suppressed cell proliferation and colony formation, whereas knockout of miR-616 increased it. We found that miR-616 represses c-MYC expression via binding sites located in its protein coding region. Furthermore, we show that miR-616 exerted growth inhibitory effects on cells by suppressing c-MYC expression. Our results establish a new role for the CHOP locus by providing evidence that miR-616 can inhibit cell proliferation by targeting c-MYC. In summary, our results suggest a dual function for the CHOP locus, where CHOP protein and miR-616 can cooperate to inhibit cancer progression.

IFN-γ and IL-12 from Concentrated Ascites in Patients with Pancreatic Cancer Exerts Growth Inhibitory Effects against Pancreatic Cancer Cells

Patients with pancreatic cancer (PC) often suffer from refractory ascites associated with peritoneal metastasis. This severely impairs activities of daily living and leads to an unfavorable prognosis. Cell-free and concentrated ascites reinfusion therapy (CART) has attracted attention as a promising therapy for relieving the symptoms of malignant ascites. Accumulating evidence suggests that malignant ascites contains a variety of soluble factors, such as cytokines, that can be beneficial or detrimental in the prognosis of patients with refractory ascites. However, the expression profiles of these cytokines in the ascites before and after CART remain unknown. In this study, we used a comprehensive cytokine array to measure the expression levels of 102 cytokines in ascites derived from patients with PC before and after CART. The assay results revealed that the concentrations of several cytokines exacerbating tumor angiogenesis and tumor-suppressive interferon-gamma (IFN-γ) and interleukin-12 (IL-12) were higher in ascites after CART than before CART. Interestingly, growth of KP-2 human PC cells following exposure to ascites after CART decreased considerably compared to that before CART. Concomitant treatment of neutralizing antibodies against IFN-γ or IL-12 with ascites after CART restored the growth of KP-2 cells to the control level. These findings indicate that IFN-γ and IL-12 in ascites after CART may contribute to the inhibited growth of PC cells, highlighting their potential as biomarkers for assessing the clinical efficacy of CART procedures in patients with PC.

CNSC-17. GLIOMA SYNAPSES RECRUIT MECHANISMS OF ADAPTIVE PLASTICITY

Abstract The nervous system plays an increasingly appreciated role in the regulation of cancer. In malignant gliomas, neuronal activity drives tumor progression not only through paracrine signaling factors such as neuroligin-3 and brain-derived neurotrophic factor (BDNF), but also through electrophysiologically functional neuron-to-glioma synapses. Malignant synapses are mediated by calcium-permeable AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors in both pediatric and adult high-grade gliomas, and consequent depolarization of the glioma cell membrane drives tumor proliferation. The nervous system exhibits plasticity of both synaptic connectivity and synaptic strength, contributing to neural circuit form and functions. In health, one factor that promotes plasticity of synaptic connectivity and strength is activity-regulated secretion of the neurotrophin BDNF. Here, we show that malignant synapses exhibit similar plasticity regulated by BDNF-TrkB (tropomyosin receptor kinase B) signaling. Signaling through the receptor TrkB, BDNF promotes AMPA receptor trafficking to the glioma cell membrane, resulting in increased amplitude of glutamate-evoked currents in the malignant cells. This potentiation of malignant synaptic strength shares mechanistic features with the long-term potentiation (LTP) that is thought to contribute to memory and learning in the healthy brain. BDNF-TrkB signaling also regulates the number of neuron-to-glioma synapses. Abrogation of activity-regulated BDNF secretion from the brain microenvironment or loss of TrkB in human glioma cells exerts growth inhibitory effects in vivo and in neuron:glioma co-cultures that cannot be explained by classical growth factor signaling alone. Blocking TrkB genetically or pharmacologically abrogates these effects of BDNF on glioma synapses and substantially prolongs survival in xenograft models of pediatric glioblastoma and diffuse intrinsic pontine glioma (DIPG). Taken together, these findings indicate that BDNF-TrkB signaling promotes malignant synaptic plasticity and augments tumor progression. While targeting Trk signaling is presently being explored for NTRK-fusion brain tumors, these findings indicate that BDNF-TrkB signaling may also represent an important therapeutic target for NTRK2 wildtype gliomas.

MicroRNA-15 suppresses viability, migration and invasion of the human MG-63 osteosarcoma cells via inhibition of cyclin dependent kinase 6 (CDK6).

MicroRNA-15a-3p (miR-15) acts as tumor-suppressor in different human cancers including osteosarcoma. Nonetheless, the molecular function of miR-15 in osteosarcoma via suppression of cyclin dependent kinase 6 (CDK6) is yet to be studied. The results showed significant downregulation of miR-15 in osteosarcoma tissues and cell lines. Overexpression of miR-15 inhibited the proliferation and colony formation of the MG-63 osteosarcoma cells via induction of apoptosis. Moreover, miR-15 inhibited the migration and invasion of MG-63 osteosarcoma cells. The tumor-suppressive functional role of miR-15 was shown to be exerted via suppression of CDK6. The expression of CDK6 was upregulated in osteosarcoma and its silencing could exert growth inhibitory effects on human osteosarcoma cells. However, overexpression of CDK6 could nullify the tumor-suppressive effects of miR-15 on the MG-63 osteosarcoma cells. Taken together, miR-15 negatively regulates growth, migration and invasion of osteosarcoma cells by targeting CDK6 at post-transcriptional level. These findings suggest the therapeutic potential of miR-15/CDK6 in human osteosarcoma.

Biosynthesis, Characterization and Evaluation of Biological Properties of Biogenic Gold Nanoparticles Synthesized Using Nothapodytes foetida Leaf Extract

Background and Objectives: Biogenic nanomaterials are recently being researched in biomedicine for their antioxidant, antimicrobial, anti-inflammatory and antitumor performance. In this study, gold nanoparticles (AuNPs) were biologically synthesized using Nothapodytes foetida (N. foetida), a camptothecin producing plant, to evaluate its potent cytotoxic activity against cancer cells and antimicrobial activities against human pathogenic bacteria. Methods: The AuNPs were characterized by UV-visible spectroscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM), fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and dynamic light scattering (DLS). The antimicrobial activity of AuNPs was determined by conducting the minimum inhibitory concentration (MIC) assay on gram positive and gram negative bacteria, whereas cytotoxicity against cancer cells was assessed through the MTT assay and genotoxicity was assayed by DNA fragmentation. Results: TEM and DLS results showed the synthesis of 20-200 nm sized M. foetida AuNPs (NFAuNPs). The NFAuNPs exerted growth inhibitory effects on gram negative bacteria with the MIC for the inhibition of Eschericia coli, Pseudomonas aeruginosa being 0.15 mg/mL and 0.25 mg/mL respectively. The cytotoxicity results indicated that the sublethal concentrations of NFAuNPs for inhibiting 50% of cell growth (IC50) of cancer cells including HeLa, MCF-7 and HCT-15 after 48 h of exposure were 7.2, 9.67 and 5.28 μg/mL respectively. DNA fragmentation in cells exposed to 10-75 μg/mL of NFAuNP concentrations revealed their genotoxic effects. Conclusion: The study revealed the biogenic synthesis of AuNPs using the N. foetida plant extract and its biological potential as an antimicrobial and anticancer agent.

Indirubin exerts anticancer effects on human glioma cells by inducing apoptosis and autophagy

Glioma causes significant mortality across the world and the most aggressive type of brain cancer. The incidence of glioma is believed to increase in the next few decades and hence more efficient treatment strategies need to be developed for management of glioma. Herein, we examined the anticancer effects of Indirubin against a panel of human glioma cells and attempted to explore the underlying mechanisms. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay showed that Indirubin could inhibit the growth of all the glioma cells but the lowest IC50 of 12.5 µM was observed against the U87 and U118 glioma cells. Additionally, the cytotoxic effects of Indirubin were comparatively negligible against the normal astrocytes with an IC50 of > 100 µM. Investigation of mechanism of action, revealed that Indirubin exerts growth inhibitory effects on the U87 and U118 glioma cells by autophagic and apoptotic cell death. Annexin V/PI staining assay showed that apoptotic cell percentage increased dose dependently. Apoptosis was associated with increase in Bax decrease in Bcl-2 expressions. Additionally, the expression of autophagic proteins such as LC3II, ATG12, ATG15 and Beclin 1 was also increased. Wound heal assay showed that Indirubin caused remarkable decrease in the migration of the U87 and U118 cells indicative of anti-metastatic potential of Indirubin. Taken together, these results suggest that Indirubin exerts potent anticancer effects on glioma cells and may prove essential in the management of glioma.

Long non-coding RNA GAS5 regulates the growth and metastasis of human cervical cancer cells via induction of apoptosis and cell cycle arrest

Studies have proved the role of GAS5 in the development of different cancers. This study was undertaken to investigate the role and explore therapeutic implications of GAS5 in human cervical cancer. The results showed that GAS5 was significantly (p < 0.05) downregulated in human cervical cancer tissues. The results also showed that cervical cancer progresses with the suppression of GAS5 expression levels. Additionally, the expression of GAS5 was also significantly (p < 0.05) downregulated in human cervical cancer cell lines. Nonetheless, overexpression of GAS5 caused a remarkable decrease in the proliferation of C33A and HeLa cervical cancer cells. The decrease in the proliferation rate was attributed to the induction of apoptosis of C33A and HeLa cells which was accompanied with upregulation of Bax and suppression of Bcl-2. Additionally, GAS5 overexpression also promoted the arrest of C33A and HeLa cells at the G2/M check point of cell cycle via suppression of cyclin B1 and CDK1 expression. The transwell assays showed that GAS5 overexpression significantly (p < 0.05) inhibited the migration and invasion of the C33A and HeLa cervical cancer cells. The bioinformatics analysis as well as the dual luciferase assay showed GAS5 acts as a target of miR-135a. Interestingly, the expression of miR-135a was upregulated in the human cervical cancer cells and its suppression exerted growth inhibitory effects on the C33A and HeLa cells. However, silencing of GAS5 could nullify the effects of miR-135a suppression on the proliferation of C33A and HeLa cells. Taken together, the results of this study point towards the therapeutic implications of GAS5 in the treatment of cervical cancer.

Dealing with saturated and unsaturated fatty acid metabolism for anticancer therapy.

Although saturated fatty acid (FA) (SFA) and monounsaturated FA (MUFA) are synthesized in cancer cells from acetyl-CoA, polyunsaturated FAs (PUFAs) are necessarily obtained from diet. Depending on concentrations and metabolism, these different FAs may support tumor proliferation but also exert growth inhibitory effects. The mutual interplay between them also requires to integrate the FA oxidation component that may be concomitant with FA synthesis is cancer cells. New molecular mechanisms driving FA synthesis, lipotoxicity and anti-inflammatory activity of eicosanoids in mouse and human cancers were recently elicited. To block or take advantage of the above represent attractive perspectives of treatments to fight cancer progression. The various enzymatic reactions leading to SFA synthesis represent as many targets to prevent tumor growth. Ironically excess SFAs are per-se toxic for cancer cells and the introduction of a double bound to form MUFA is actually limiting lipotoxicity in cancer cells. Blocking stearoyl-CoA desaturase therefore represents another attractive modality. By contrast, dietary PUFAs may exert direct cytotoxic effects by promoting apoptosis or by generating anti-inflammatory eicosanoids. Altogether, these data point out the intricate relationship between SFA, MUFA and PUFA at the heart of the metabolism of proliferating cancer cells.

Growth inhibition and transcriptional effects of ribavirin in lymphoma.

Ribavirin exhibits inhibitory effects on the epigenetic enzyme enhancer of zeste homolog2 (EZH2), which participates in lymphomagenesis. Additionally, preclinical and clinical studies have demonstrated the anti‑lymphoma activity of this drug. To further investigate the potential of ribavirin as an anticancer treatment for lymphoma, the tumor‑suppressive effects of ribavirin were analyzed in lymphoma cell lines. The effects of ribavirin on the viability and clonogenicity of the B‑cell lymphoma cell line Pfeiffer (EZH2‑mutant), Toledo (EZH2 wild‑type) and cutaneous T‑cell lymphoma Hut78 cell line were assessed. Expression of EZH2 and trimethylation status of histone3, lysine27 trimethylated (H3K27m3) was also determined in response to ribavirin. The transcriptional effects of ribavirin on Hut78 cells were analyzed by microarray expression and the results were validated by reverse transcription‑quantitative polymerase chain reaction, western blotting and knockout of signal transducer and activator of transcription1 (STAT1). The results of the present study demonstrated that ribavirin suppressed the growth and clonogenicity of cells in a dose‑dependent manner. Ribavirin did not affect the expression of EZH2 nor altered its activity as evaluated by H3K27 trimethylation status. Furthermore, the results of transcriptome analysis indicated that the majority of the canonical pathways affected by ribavirin were associated with the immune system, including 'antigen presentation', 'communication between innate and adaptive immune cells' and 'cross‑talk between dendritic and natural killer cells'. The results of gene expression analysis were confirmed, by demonstrating at the RNA and protein levels, downregulation of stearoyl‑CoA desaturase and upregulation of STAT1. Depletion of STAT1, which was proposed as a key regulator of the aforementioned pathways, exerted growth inhibitory effects almost to the same extent as ribavirin. In conclusion, ribavirin was proposed to exert growth inhibitory effects on lymphoma cell lines, particularly Hut78 cells, a cutaneous T‑cell lymphoma cell line. Of note, these effects may depend on, at least in part, the activation of canonical immune pathways regulated by the key factors STAT1 and interferon‑γ. Our results provide insight into the anti‑lymphoma potential of ribavirin; however, further investigations in preclinical and clinical studies are required to determine the effectiveness of ribavirin as a therapeutic agent for treating lymphoma.

Administration of metformin to increase the efficacy of chemotherapy regimen in cancers; a new look to an old drug

Chemotherapeutic agents are still the major and most common therapy to prevent tumor growth. These drugs have various adverse effects in different organs in addition to systemic effects. Finding more specific and effective drugs or new adjuvant therapeutic substance is needed to improve the success rate. Several studies have proposed the possible mechanisms of anti-neoplastic of metformin, however, its exact mechanism is still obscure. The suggested mechanisms are; alteration in the host metabolic environment after administration of metformin, such as decreases in insulin-dependent stimulation of tumor growth or direct effect on cancer cells, such as the impact on adenosine monophosphate (AMP)-activated protein kinase signaling pathway. Metformin, as an adjuvant therapy, synergistically exerts growth inhibitory effects against cell growth and can induce cell apoptosis in an animal model. Adding metformin to the chemotherapy regimen may reduce resistance and enhance therapeutic efficacy.

Metformin Induces Apoptosis and Alters Cellular Responses to Oxidative Stress in Ht29 Colon Cancer Cells: Preliminary Findings.

Accumulating evidence suggests that metformin, used as an antidiabetic drug, possesses anti-cancer properties. Metformin reduced the incidence and growth of experimental tumors in vivo. In a randomized clinical trial among nondiabetic patients, metformin treatment significantly decreased the number of aberrant crypt foci compared to the untreated group with a follow-up of 1 month. In our study, HT29 cells were treated with graded concentrations of metformin, 10 mM/25 mM/50 mM for 24/48 h. We performed immunofluorescence experiments by means of confocal microscopy and western blot analysis to evaluate a panel of factors involved in apoptotic/autophagic processes and oxidative stress response. Moreover, HT29 cells treated with metformin were analyzed by a flow cytometry assay to detect the cell apoptotic rate. The results demonstrate that metformin exerts growth inhibitory effects on cultured HT29 cells by increasing both apoptosis and autophagy; moreover, it affects the survival of cultured cells inhibiting the transcriptional activation of Nuclear factor E2-related factor 2 (NRF-2) and nuclear factor-kappa B (NF-κB). The effects of metformin on HT29 cells were dose- and time-dependent. These results are very intriguing since metformin is emerging as a multi-faceted drug: It has a good safety profile and is associated with low cost and might be a promising candidate for the prevention or the treatment of colorectal cancer.

Lactobacillus raises in vitro anticancer effect of geniposide in HSC-3 human oral squamous cell carcinoma cells.

The present study determined the ability of the Lactobacillus rhamnosus GG strain (LGG) to enhance the anticancer effects of geniposide on HSC-3 human oral squamous carcinoma cells. LGG (1.0×103 CFU/ml) on its own had no impact on human oral keratinocytes and HSC-3 cancer cells. Geniposide (25 or 50 µg/ml) had no impact on human oral keratinocytes, but exerted growth inhibitory effects on HSC-3 cancer cells, which were increased in the presence of LGG. Flow cytometric analysis and a nuclear staining assay with DAPI revealed that HSC-3 cancer cells treated with LGG-geniposide (1.0×103 CFU/ml LGG and 50 µg/ml geniposide) had a higher apoptotic rate than cells in other treatment groups, particularly that treated with geniposide (50 µg/ml) only. Geniposide also increased the mRNA and protein expression of caspase-3, -8 and -9 as well as B-cell lymphoma 2 (Bcl-2)-associated X protein, p53, p21, inhibitor of nuclear factor-κB (NF-κB) α, Fas and Fas ligand, while decreasing Bcl-2, Bcl extra large protein, inhibitor of apoptosis-1 and -2, NF-κB, cyclooxigenase-2 and inducible nitric oxide synthase in HSC-3 cells, which was increased in the presence of LGG. These results indicated that LGG enhanced the anticancer effects of geniposide in HSC-3 cells.

Dietary mastic oil extracted from Pistacia lentiscus var. chia suppresses tumor growth in experimental colon cancer models

Plant-derived bioactive compounds attract considerable interest as potential chemopreventive anticancer agents. We analyzed the volatile dietary phytochemicals (terpenes) present in mastic oil extracted from the resin of Pistacia lentiscus var. chia and comparatively investigated their effects on colon carcinoma proliferation, a) in vitro against colon cancer cell lines and b) in vivo on tumor growth in mice following oral administration. Mastic oil inhibited - more effectively than its major constituents- proliferation of colon cancer cells in vitro, attenuated migration and downregulated transcriptional expression of survivin (BIRC5a). When administered orally, mastic oil inhibited the growth of colon carcinoma tumors in mice. A reduced expression of Ki-67 and survivin in tumor tissues accompanied the observed effects. Notably, only mastic oil -which is comprised of 67.7% α-pinene and 18.8% myrcene- induced a statistically significant anti-tumor effect in mice but not α-pinene, myrcene or a combination thereof. Thus, mastic oil, as a combination of terpenes, exerts growth inhibitory effects against colon carcinoma, suggesting a nutraceutical potential in the fight against colon cancer. To our knowledge, this is the first report showing that orally administered mastic oil induces tumor-suppressing effects against experimental colon cancer.

Impact of sesquiterpenes from Inula racemosa (Asteraceae) on growth, development and nutrition of Spodoptera litura (Lepidoptera: Noctuidae).

The use of botanical pesticides for protecting crops from insect pests has assumed greater importance all over the world owing to growing awareness of harmful effects of indiscriminate use of synthetic pesticides. Inula racemosa Hook. f. (Asteraceae), a medicinally important perennial herb, is rich in sesquiterpenes with many biological activities. The present studies were conducted with the objective to evaluate the sesquiterpenes isolated from I. racemosa for insecticidal activity against Spodoptera litura (F.). Alantolactone and isoalantolactone isolated from I. racemosa exerted growth inhibitory effects on S. litura. Addition of both the sesquiterpenes to larval diet extended the development period and reduced pupation as well as adult emergence. The dietary utilisation experiments on third-instar larvae of S. litura revealed reduction in consumption and growth rates of larvae as well as efficiency of conversion of ingested and digested food owing to alantolactone and isoalantolactone. The root extract of I. racemosa, which is rich in two sesquiterpenes, i.e. alantolactone and isoalantolactone, has the potential for management of S. litura. However, there is a need to understand the specific mechanism of action of these compounds. © 2016 Society of Chemical Industry.

3-(Dipropylamino)-5-hydroxybenzofuro[2,3-f]quinazolin-1(2H)-one (DPA-HBFQ-1) plays an inhibitory role on breast cancer cell growth and progression

A series of unknown 3-(alkyl(dialkyl)amino)benzofuro[2,3-f]quinazolin-1(2H)-ones 4–17 has been synthesized as new ellipticine analogs, in which the carbazole moiety and the pyridine ring were replaced by a dibenzofuran residue and a pyrimidine ring, respectively. The synthesis of these benzofuroquinazolinones 4–17 was performed in a simple one-pot reaction using 3-aminodibenzofuran or its 2-methoxy derivative, as starting materials. From 3-(dipropylamino)-5-methoxybenzofuro[2,3-f] quinazolin-1(2H)-one (13), we prepared 3-(dipropylamino)-5-hydroxybenzofuro[2,3-f]quinazolin-1(2H)-one (18), referred to as DPA-HBFQ-1. The cytotoxic activities of all the synthesized compounds, tested in different human breast cancer cell lines, revealed that DPA-HBFQ-1 was the most active compound. In particular, the latter was able to inhibit anchorage-dependent and -independent cell growth and to induce apoptosis in estrogen receptor alpha (ERα)-positive and -negative breast cancer cells. It did not affect proliferation and apoptotic responses in MCF-10A normal breast epithelial cells. The observed effects have been ascribed to an enhanced p21Cip1/WAF1 expression in a p53-dependent manner of tumor suppressor and to a selective inhibition of human topoisomerase II. In addition, DPA-HBFQ-1 exerted growth inhibitory effects also in other cancer cell lines, even though with a lower cytotoxic activity. Our results indicate DPA-HBFQ-1 as a good candidate to be useful as cancer therapeutic agent, particularly for breast cancer.

NBPF1, a tumor suppressor candidate in neuroblastoma, exerts growth inhibitory effects by inducing a G1 cell cycle arrest.

BackgroundNBPF1 (Neuroblastoma Breakpoint Family, member 1) was originally identified in a neuroblastoma patient on the basis of its disruption by a chromosomal translocation t(1;17)(p36.2;q11.2). Considering this genetic defect and the frequent genomic alterations of the NBPF1 locus in several cancer types, we hypothesized that NBPF1 is a tumor suppressor. Decreased expression of NBPF1 in neuroblastoma cell lines with loss of 1p36 heterozygosity and the marked decrease of anchorage-independent clonal growth of DLD1 colorectal carcinoma cells with induced NBPF1 expression further suggest that NBPF1 functions as tumor suppressor. However, little is known about the mechanisms involved.MethodsExpression of NBPF was analyzed in human skin and human cervix by immunohistochemistry. The effects of NBPF1 on the cell cycle were evaluated by flow cytometry. We investigated by real-time quantitative RT-PCR the expression profile of a panel of genes important in cell cycle regulation. Protein levels of CDKN1A-encoded p21CIP1/WAF1 were determined by western blotting and the importance of p53 was shown by immunofluorescence and by a loss-of-function approach. LC-MS/MS analysis was used to investigate the proteome of DLD1 colon cancer cells with induced NBPF1 expression. Possible biological interactions between the differentially regulated proteins were investigated with the Ingenuity Pathway Analysis tool.ResultsWe show that NBPF is expressed in the non-proliferative suprabasal layers of squamous stratified epithelia of human skin and cervix. Forced expression of NBPF1 in HEK293T cells resulted in a G1 cell cycle arrest that was accompanied by upregulation of the cyclin-dependent kinase inhibitor p21CIP1/WAF1 in a p53-dependent manner. Additionally, forced expression of NBPF1 in two p53-mutant neuroblastoma cell lines also resulted in a G1 cell cycle arrest and CDKN1A upregulation. However, CDKN1A upregulation by NBPF1 was not observed in the DLD1 cells, which demonstrates that NBPF1 exerts cell-specific effects. In addition, proteome analysis of NBPF1-overexpressing DLD1 cells identified 32 differentially expressed proteins, of which several are implicated in carcinogenesis.ConclusionsWe demonstrated that NBPF1 exerts different tumor suppressive effects, depending on the cell line analyzed, and provide new clues into the molecular mechanism of the enigmatic NBPF proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1408-5) contains supplementary material, which is available to authorized users.

Novel Pactamycin Analogs Induce p53 Dependent Cell-Cycle Arrest at S-Phase in Human Head and Neck Squamous Cell Carcinoma (HNSCC) Cells.

Pactamycin, although putatively touted as a potent antitumor agent, has never been used as an anticancer drug due to its high cytotoxicity. In this study, we characterized the effects of two novel biosynthetically engineered analogs of pactamycin, de-6MSA-7-demethyl-7-deoxypactamycin (TM-025) and 7-demethyl-7-deoxypactamycin (TM-026), in head and neck squamous cell carcinoma (HNSCC) cell lines SCC25 and SCC104. Both TM-025 and TM-026 exert growth inhibitory effects on HNSCC cells by inhibiting cell proliferation. Interestingly, unlike their parent compound pactamycin, the analogs do not inhibit synthesis of nascent protein in a cell-based assay. Furthermore, they do not induce apoptosis or autophagy in a dose- or a time-dependent manner, but induce mild senescence in the tested cell lines. Cell cycle analysis demonstrated that both analogs significantly induce cell cycle arrest of the HNSCC cells at S-phase resulting in reduced accumulation of G2/M-phase cells. The pactamycin analogs induce expression of cell cycle regulatory proteins including master regulator p53, its downstream target p21Cip1/WAF1, p27kip21, p19, cyclin E, total and phospho Cdc2 (Tyr15) and Cdc25C. Besides, the analogs mildly reduce cyclin D1 expression without affecting expression of cyclin B, Cdk2 and Cdk4. Specific inhibition of p53 by pifithrin-α reduces the percentage of cells accumulated in S-phase, suggesting contribution of p53 to S-phase increase. Altogether, our results demonstrate that Pactamycin analogs TM-025 and TM-026 induce senescence and inhibit proliferation of HNSCC cells via accumulation in S-phase through possible contribution of p53. The two PCT analogs can be widely used as research tools for cell cycle inhibition studies in proliferating cancer cells with specific mechanisms of action.

Rigosertib induces cell death of a myelodysplastic syndrome-derived cell line by DNA damage-induced G2/M arrest.

A multi-kinase inhibitor, rigosertib (ON 01910.Na) has recently been highlighted as a novel type of anti-cancer agent for the treatment of the myelodysplastic syndromes (MDS), but its action mechanisms remain to be clarified. We investigated the in vitro effects of rigosertib on an MDS-derived cell line MDS-L and a myeloid leukemia cell line HL-60. Rigosertib suppressed the proliferation of both HL-60 and MDS-L cells and induced apoptosis by inhibition of the PI3 kinase/Akt pathway. As the effects on cell cycle, rigosertib treatment promoted the phosphorylation of histone H2AX and led to the DNA damage-induced G2/M arrest. In addition, an immunofluorescence staining study demonstrated the abnormal localization of aurora A kinase, suggesting that rigosertib causes perturbation of spindle assembly and deregulated mitotic patterns towards cell cycle arrest and apoptosis. We also found that rigosertib exerted growth inhibitory effects on two lymphoid cell lines, Jurkat and Ramos. We further examined the molecular pathways influenced by rigosertib from the gene expression profiling data of MDS-L cells and found a possible involvement of rigosertib treatment in the upregulation of the genes related to microtubule kinetics and the downregulation of the mRNA degradation system. The gene set enrichment analysis showed the suppression of “nonsense-mediated mRNA decay (NMD)” as the most significantly affected gene set. These data provide a new aspect and a potential utility of rigosertib for the treatment of refractory hematopoietic malignancies.

Resveratrol exerts growth inhibitory effects on human SZ95 sebocytes through the inactivation of the PI3-K/Akt pathway

Resveratrol is known to be a naturally produced polyphenol that is able to reduce cell proliferation in several types of cancer cells and adipocytes. However, the antiproliferative effects of resveratrol on the growth of human sebocytes are not yet clear. In the present study, we investigated possible cellular pathways associated with these growth inhibitory effects on human SZ95 sebocytes. Our results revealed that resveratrol inhibited the proliferation of sebocytes, and that this resulted in disruption of the cell cycle. The inactivation of extracellular signal-regulated protein kinase (ERK), Akt and peroxisome proliferator-activated receptor (PPAR)-γ was also shown to be involved in the inhibition of sebocyte growth by resveratrol. To examine the antiproliferative effects of resveratrol, we determined the levels of cell cycle control proteins. Resveratrol inhibited cyclin D1 synthesis, whereas it stimulated p21WAF1/CIP1 (p21) and p27KIP1 (p27) synthesis. In addition, we demonstrated that the resveratrol-mediated cell cycle arrest resulted in an increase in the proportion of cells in the sub-G0/G1 phase. Moreover, we found that the growth inhibitory effects of resveratrol were enhanced by treatment with LY294002 [a phosphatidylinositol 3-kinase (PI3-K) inhibitor] more so than by treatment with PD98059 (a MEK inhibitor), which indicates that resveratrol exerts its inhibitory effects on sebocyte proliferation through the inhibition of Akt. Linoleic acid (LA) is a well-established lipid inducer in sebocytes and is known to stimulate sebocyte differentiation through the upregulation of PPAR-γ. In this study, resveratrol was found to decrease the lipid content and PPAR-γ expression during LA-stimulated lipogenesis. Our results indicate that resveratrol plays a critical role in the inhibition of sebocyte growth through the inactivation of the Akt pathway. The present data suggest that resveratrol may be used as a therapeutic agent for the treatment of acne vulgaris.