Evaluation and molecular docking study of two flavonoids from Oroxylum indicum (L.) Kurz and their semi-synthetic derivatives as histone deacetylase inhibitors.

Chrysin (5,7-dihydroxyflavone, 6) and galangin 3-methyl ether (5,7-dihydroxy-3-methoxy flavone, 7) were obtained from the leaves of Oroxylum indicum (L.) Kurz in 4% and 6% yields, respectively. Both compounds could act as pan-histone deacetylase (HDAC) inhibitors. Structural modification of these lead compounds provided thirty-eight derivatives which were further tested as HDAC inhibitors. Compounds 6b, 6c, and 6q were the most potent derivatives with the IC50 values of 97.29 ± 0.63 µM, 91.71 ± 0.27 µM, and 96.87 ± 0.45 µM, respectively. Molecular docking study indicated the selectivity of these three compounds toward HDAC8 and the test against HDAC8 showed IC50 values in the same micromolar range. All three compounds were further evaluated for the anti-proliferative activity against HeLa and A549 cell lines. Compound 6q exhibited the best activity against HeLa cell line with the IC50 value of 13.91 ± 0.34 µM. Moreover, 6q was able to increase the acetylation level of histone H3. These promising HDAC inhibitors deserve investigation as chemotherapeutic agents for treating cancer.

Momordicae Semen inhibits migration and induces apoptotic cell death by regulating c-Myc and CNOT2 in human pancreatic cancer cells.

Pancreatic cancer(PC) is less common than other cancers; however, it has a poor prognosis. Therefore, studying novel target signaling and anticancer agents is necessary. Momordicae Semen (MS), the seed of Momordica sochinensis Spreng, mainly found in South-East Asia, including China and Bangladesh, is used to treat various diseases because of its anticancer, antioxidant, anti-inflammatory, and antibacterial properties. However, the effect of the MS extract on pancreatic cancer cells remains unknown. In this study investigated whether the MS extract exerted an anti-cancer effect by regulating c-Myc through CNOT2. Cytotoxicity and proliferation were investigated using MTT and colony formation assays. The levels of apoptotic, oncogenic, and migration-associated factors were confirmed using immunoblotting and immunofluorescence. Wound closure was analyzed using a wound healing assay. The chemical composition of the MS methanol extracts was analyzed using liquid chromatography-mass spectrometry. We confirmed that the MS extract regulated apoptotic factors and attenuated the stability of c-Myc and its sensitivity to fetal bovine serum. Furthermore, the MS extract increased apoptosis by regulating c-Myc and CNOT2 expression and enhanced the sensitivity of 5-FU in pancreatic cancer. This study showed that the MS extract is a promising new drug for PC.

WIF1 was downregulated in cervical cancer due to promoter methylation.

Wnt inhibitory factor 1 (WIF1) is frequently downregulated in a variety of cancer due to promoter methylation. However, the methylation status of the WIF1 promoter in cervical cancer remains unclear. This study aimed to elucidate the mechanism by which WIF1 promoter methylation contributes to cervical cancer development. The expression of WIF1 in cervical cancer tissues was examined by immunohistochemistry. The methylation status of the WIF1 promoter in cervical cancer cells was detected by methylation specific PCR. WIF1 mRNA levels and protein levels were detected by PCR and Western blot analysis. We found that WIF1 expression was low in cervical cancer tissues compared to adjacent normal cervical tissues. The WIF1 promoter was methylated in the cervical cancer SiHa cell line but not in the normal cervical epithelial cell line Ect1. Correspondingly, WIF1 mRNA levels and protein levels were significantly lower in SiHa cells than in Ect1 cells. Treatment with 5-aza-2-deoxycytidine (AZA) led to the upregulation of WIF1 mRNA and protein levels in SiHa cells, but the effects were abrogated by treatment with WIF1 siRNA. In addition, AZA treatment induced apoptosis and inhibited the invasion of SiHa cells, and the effects were abrogated by WIF1 siRNA. The protein levels of survivin, c-myc and cyclinD1 were significantly lower in SiHa cells treated with AZA, but their levels were upregulated after treatment with WIF1 siRNA. In conclusion, the methylation of the WIF1 promoter leads to the downregulation of WIF1 and the activation of Wnt/ß-catenin signaling in cervical cancer cells. WIF1 is a tumor suppressor that is inactivated in cervical cancer.

Discovery of novel polysubstituted N-alkyl acridone analogues as histone deacetylase isoform-selective inhibitors for cancer therapy.

Pan-histone deacetylase (HDAC) inhibitors often have some toxic side effects. In this study, three series of novel polysubstituted N-alkyl acridone analogous were designed and synthesised as HDAC isoform-selective inhibitors. Among them, 11b and 11c exhibited selective inhibition of HDAC1, HDAC3, and HDAC10, with IC50 values ranging from 87 nM to 418 nM. However, these compounds showed no inhibitory effect against HDAC6 and HDAC8. Moreover, 11b and 11c displayed potent antiproliferative activity against leukaemia HL-60 cells and colon cancer HCT-116 cells, with IC50 values ranging from 0.56 µM to 4.21 µM. Molecular docking and energy scoring functions further analysed the differences in the binding modes of 11c with HDAC1/6. In vitro anticancer studies revealed that the hit compounds 11b and 11c effectively induced histone H3 acetylation, S-phase cell cycle arrest, and apoptosis in HL-60 cells in a concentration-dependent manner.

BASP1 promotes high glucose-induced endothelial apoptosis in diabetes via activation of EGFR signaling.

AIMS: Diabetes mellitus is a common chronic disease of glucose metabolism. Endothelial dysfunction is an early event in diabetes complicated by cardiovascular disease. This study aimed to reveal the expression of BASP1 and its biological roles in endothelial cell dysfunction in diabetes complicated by cardiovascular disease. MATERIALS AND METHODS: By analyzing the databases related to diabetes complicated with coronary heart disease, BASP1 was screened out as an upregulated gene. Human umbilical vein endothelial cells (HUVECs) and primary mouse aortic endothelial cells were treated with high glucose to establish cell models of diabetes-related endothelial dysfunction, and the expression changes of BASP1 were verified by RT-qPCR, western blot, and immunofluorescence. BASP1 was silenced or overexpressed by siRNA or overexpression plasmid, and its effects on cell migration, apoptosis, tube formation, inflammatory response, and ROS were detected. The possible signaling pathway of BASP1 was found and the mechanism of BASP1 on promoting the progression of endothelial dysfunction was explored using the EGFR inhibitor, gefitinib. RESULTS: Bioinformatics analysis indicated that the expression of BASP1 in patients with diabetes mellitus and concomitant coronary heart disease was increased. High glucose induced the upregulation of BASP1 expression in endothelial cells, and showed a time-dependent relationship. Silencing of BASP1 alleviated the damage of high glucose to endothelial cells. BASP1 regulated EGFR positively. The promoting effect of BASP1 on endothelial cell apoptosis may be achieved by regulating the EGFR pathway. CONCLUSION: BASP1 promotes endothelial cell injury induced by high glucose in patients with diabetes, which may be activated by activating the EGFR pathway.

TRIM22 actives PI3K/Akt/mTOR pathway to promote psoriasis through enhancing cell proliferation and inflammation and inhibiting autophagy.

OBJECTIVE: To reveal the function and underlying mechanism of Tri-domain protein 22 (TRIM22) in psoriasis. METHODS: M5 cytokines were applied in HaCat cells to mimic psoriasis in vitro. The TRIM22-silencing viruses were established to knockdown TRIM22 in HaCat cells. Western blot and/or real-time PCR were used to detect the expression of TRIM22, KRT1, KRT6, p-P65, P65, LC3, Beclin 1, P62, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, and mTOR. ELISA kits were applied to assess levels of TNF-α, IL-1ß, IL-18, and HMGB1. RESULTS: TRIM22 expression levels were upregulated in M5-treated HaCat cells. M5 treatment enhanced cell proliferation and inflammation, and inhibited autophagy in HaCat cells which were effectively reversed by TRIM22 deficiency. Activation of PI3K/Akt/mTOR pathway is an essential promoter of cell proliferation and inflammation, and inhibitor of autophagy in psoriasis. TRIM22 deficiency blocked M5-induced activation of PI3K/Akt/mTOR pathway in HaCat cells. CONCLUSIONS: TRIM22 facilitates cell proliferation and inflammation, and suppresses autophagy in M5-treated HaCat cells through activating PI3K/Akt/mTOR pathway, and inhibition of TRIM22 can be a novel potential treatment for psoriasis.

miR-150 promotes progressive T cell differentiation via inhibiting FOXP1 and RC3H1.

T cells used in immune cell therapy, represented by T cell receptor therapy (TCR-T), are usually activated and proliferated in vitro and are induced to a terminally differentiated phenotype, with limited viability after transfusion back into the body. T cells exhibited a robust proliferative potential and in vivo viability in the early stages of progressive differentiation. In this study, we identified microRNAs that regulate T cell differentiation. After microRNA sequencing of the four subsets: Naïve T cells (TN), stem cell-like memory T cells (TSCM), central memory T cells (TCM）, and effector memory T cells (TEM), miR-150 was identified as the most highly expressed miRNA among the four subsets and was lowly expressed in the TSCM cells. We predicted the target genes of miR-150 miRNA and performed Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes analyses. We observed that the target genes of miR-150 were enriched in pathways associated with T-cell differentiation. FOXP1 and RC3H1 were identified as key target genes of miR-150 in the regulation of T-cell function. We examined the effects of miR-150 on the differentiation and function of healthy donor T-cells. We observed that miR-150 overexpression promoted T-cell differentiation to effector T-cells and effector memory T-cells, enhanced apoptosis, inhibited cell proliferation and increased secretion of pro-inflammatory cytokines such as IFN-γ and TNF-α. In addition, the expressions of early differentiation-related genes (ACTN1, CERS6, BCL2, and EOMES), advanced differentiation-related genes (KLRG1), and effector-function-related genes (PRF1 and GZMB) were significantly decreased after overexpression of miR-150. Collectively, our results suggested that miR-150 can promote progressive differentiation of T cells and the downmodulation of miR-150 expression while performing adoptive immunotherapy may inhibit T-cell differentiation and increase the proliferative potential of T cells.

ZNF300 enhances temozolomide resistance in gliomas by regulating lncRNA SNHG12.

Gliomas are the most common type of primary brain tumors, with high recurrence rate and mortality. In contrast to the high incidence, the prognosis of gliomas is dismal, and the therapeutic effect of temozolomide (TMZ), first-line chemotherapy drug for gliomas, is extremely limited by TMZ-resistance. ZNF300 is a member of zinc finger proteins, and promotes cell proliferation in several types of cancer. This study is designed to appraise the effects of ZNF300 on TMZ-resistance in gliomas. The TMZ-resistant glioma cell lines were established in U251 and A-172 cells. ZNF300 was significantly upregulated in TMZ-resistant glioma cells. ZNF300 depletion remodel TMZ sensitivity of glioma cells through inhibiting cell proliferation and promoting cell apoptosis. Meanwhile, ZNF300 depletion brought down-regulation of c-myc and SNHG12 which could be counteracted by overexpression of c-myc. Moreover, SNHG12 performed as the downstream of ZNF300 and c-myc, and overexpression of SNHG12 also could neutralize the effects of ZNF300 depletion on cell proliferation and cell apoptosis. Our data manifested that ZNF300 serves as a novel therapeutic target and biomarker of TMZ-resistant gliomas via mediating c-myc/SNHG12 pathway.

MiR-218 promotes oxidative stress and inflammatory response by inhibiting SPRED2-mediated autophagy in HG-induced HK-2 cells.

BACKGROUND: Diabetic nephropathy (DN) is one of the most common complications of diabetes mellitus (DM). MicroRNA (miR)-218 is associated with the development of diabetes. Besides, sprouty-related EVH1 domain containing 2 (SPRED2), the downstream target of miR-218, is involved in insulin resistance and inflammation. OBJECTIVES: Since inflammation plays a key role in DN, and SPRED2 is known to facilitate cell autophagy, the present study aimed to investigate the role and molecular mechanism of miR-218 and SPRED2-mediated autophagy in high glucose (HG)-induced renal tubular epithelial cells using an in vitro model. MATERIAL AND METHODS: The HK-2 cells were cultured in 5.5 mM or 30 mM D-glucose medium. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-218 and SPRED2. Western blotting was performed to calculate the levels of SPRED2, inflammatory cytokines, autophagy-related and apoptosis-related proteins. Reactive oxygen species (ROS) level was evaluated using cellular ROS assay kit, superoxide dismutase (SOD) activity was detected using SOD activity assay kit, and malondialdehyde (MDA) content was measured using lipid peroxidation. The levels of interleukin (IL)-1ß, IL-6, IL-4, and tumor necrosis factor alpha (TNF-α) were detected with enzyme-linked immunosorbent assay (ELISA). Cell apoptosis was evaluated using flow cytometry analysis. The targeting relationship between miR-218 and SPRED2 was identified with a luciferase reporter. The LC3-II expression was detected with immunofluorescence. RESULTS: The miR-218 expression was upregulated and SPRED2 expression was downregulated in HG-induced HK-2 cells. The miR-218 was proven to target SPRED2 and negatively regulate SPRED2 expression. Besides, downregulated miR-218 alleviated inflammatory response, oxidative stress and cell apoptosis, but aggravated autophagy. We also showed that downregulated SPRED2 reversed the effect of miR-218 on inflammation, cell apoptosis and autophagy in HG-induced HK-2 cells. CONCLUSIONS: The miR-218 can promote oxidative stress and inflammatory response in HG-induced renal tubular epithelial cells by inhibiting SPRED2-mediated autophagy. This study might bring novel understanding for molecular mechanism of DN.

Overcoming radio-resistance in esophageal squamous cell carcinoma via hypermethylation of PIK3C3 promoter region mediated by KDM5B loss.

Many patients with esophageal squamous cell carcinoma (ESCC) are inoperable because of old age or the advanced stage of the disease; thus radio- and chemotherapy are believed as the standard treatments for these patients. However, due to the radio-resistance of tumor cells that may develop during radiotherapy, results remain unsatisfactory. In this article, the possible relationship between the expression of lysine demethylase 5B (KDM5B) and ESCC radio-resistance is clarified, and the underlying mechanism is evaluated. Using the GSE75241 microarray, we identified KDM5B as a potential oncogene in ESCC. KDM5B was overexpressed in ESCC patients and cells. Inhibition of KDM5B enhanced the H3K4me3 methylation of phosphatidylinositol 3-kinase catalytic subunit type 3 (PIK3C3) promoter and induced the expression of PIK3C3. Knockdown of KDM5B or overexpression of PIK3C3 in KYSE-150 and TE-10 cells promoted apoptosis, cell cycle arrest, autophagy, and increased sensitivity to radiotherapy. Silencing of PIK3C3 attenuated the promoting effect of sh-KDM5B on the sensitivity of ESCC cells to radiotherapy. The inhibition of sh-KDM5B in radio-resistance of ESCC cells was also reproduced in vivo. Taken together, our findings provide evidence that reduced expression of KDM5B has a critical role in promoting ESCC radio-sensitivity by upregulating PIK3C3, suggesting KDM5B may function as an oncogene in ESCC.

Melatonin enhances osteoblastogenesis of senescent bone marrow stromal cells through NSD2-mediated chromatin remodelling.

BACKGROUND: Aging-associated osteoporosis is frequently seen in the elderly in clinic, but efficient managements are limited because of unclear nosogenesis. The current study aims to investigate the role of melatonin on senescent bone marrow stromal cells (BMSCs) and the underlying regulating mechanism. METHODS: Melatonin levels were tested by ELISA. Gene expression profiles were performed by RNA-sequencing, enrichment of H3K36me2 on gene promoters was analyzed by Chromatin Immunoprecipitation Sequencing (ChIP-seq), and chromatin accessibility was determined by Assay for Transposase-Accessible Chromatin with high-throughput sequencing (ATAC-seq). Osteogenesis of BMSCs in vitro was measured by Alizarin Red and Alkaline Phosphatase staining, and in vivo effects of melatonin was assessed by histological staining and micro computed tomography (micro-CT) scan. Correlation of NSD2 expression and severity of senile osteoporosis patients were analyzed by Pearson correlation. RESULTS: Melatonin levels were decreased during aging in human bone marrow, accompanied by downregulation of the histone methyltransferase nuclear receptor binding SET domain protein 2 (NSD2) expression in the senescent BMSCs. Melatonin stimulated the expression of NSD2 through MT1/2-mediated signaling pathways, resulting in the rebalancing of H3K36me2 and H3K27me3 modifications to increase chromatin accessibility of the osteogenic genes, runt-related transcription factor 2 (RUNX2) and bone gamma-carboxyglutamate protein (BGLAP). Melatonin promoted osteogenesis of BMSCs in vitro, and alleviates osteoporosis progression in the aging mice. In clinic, severity of senile osteoporosis (SOP) was negatively correlated with melatonin level in bone marrow, as well as NSD2 expression in BMSCs. Similarly, melatonin remarkably enhanced osteogenic differentiation of BMSCs derived from SOP patients in vitro. CONCLUSIONS: Collectively, our study dissects previously unreported mechanistic insights into the epigenetic regulating machinery of melatonin in meliorating osteogenic differentiation of senescent BMSC, and provides evidence for application of melatonin in preventing aging-associated bone loss.

GRAMD4 inhibits tumour metastasis by recruiting the E3 ligase ITCH to target TAK1 for degradation in hepatocellular carcinoma.

BACKGROUND: Aberrant TAK1 (transforming growth factor ß-activated kinase 1) activity is known to be involved in a variety of malignancies, but the regulatory mechanisms of TAK1 remain poorly understood. GRAMD4 (glucosyltransferase Rab-like GTPase activator and myotubularin domain containing 4) is a newly discovered p53-independent proapoptotic protein with an unclear role in HCC (hepatocellular carcinoma). RESULTS: In this research, we found that GRAMD4 expression was lower in HCC samples, and its downregulation predicted worse prognosis for patients after surgical resection. Functionally, GRAMD4 inhibited HCC migration, invasion and metastasis. Mechanistically, GRAMD4 interacted with TAK1 to promote its protein degradation, thus, resulting in the inactivation of MAPK (Mitogen-activated protein kinase) and NF-κB pathways. Furthermore, GRAMD4 was proved to recruit ITCH (itchy E3 ubiquitin protein ligase) to promote the ubiquitination of TAK1. Moreover, high expression of TAK1 was correlated with low expression of GRAMD4 in HCC patients. CONCLUSIONS: GRAMD4 inhibits the migration and metastasis of HCC, mainly by recruiting ITCH to promote the degradation of TAK1, which leads to the inactivation of MAPK and NF-κB signalling pathways.

Potent Inhibition of HIF1α and p300 Interaction by a Constrained Peptide Derived from CITED2.

Disrupting the interaction between HIF1α and p300 is a promising strategy to modulate the hypoxia response of tumor cells. Herein, we designed a constrained peptide inhibitor derived from the CITED2/p300 complex to disturb the HIF1α/p300 interaction. Through truncation/mutation screening and a terminal aspartic acid-stabilized strategy, a constrained peptide was constructed with outstanding biochemical/biophysical properties, especially in binding affinity, cell penetration, and serum stability. To date, our study was the first one to showcase that stabilized peptides derived from CITED2 using helix-stabilizing methods acted as a promising candidate for modulating hypoxia-inducible signaling.

circSAMD4A participates in the apoptosis and autophagy of dopaminergic neurons via the miR­29c­3p­mediated AMPK/mTOR pathway in Parkinson's disease.

Parkinson's disease (PD) can lead to movement injury and cognitive dysfunction. Although advances have been made in attenuating PD, the effect of inhibiting the development of PD remains disappointing. Therefore, the present study aimed at investigating the etiology of Parkinson's disease and developing an alternative therapeutic strategy for patients with PD. A PD mouse model was established using an intraperitoneal injection of 1­methyl­4­phenyl­1,2,3,6­tetrahydropyridine hydrochloride (MPTP­HCl; 30 mg/kg/day for 5 days), and a PD cellular model was established by treating SH­SY5Y cells with different concentrations of 1­methyl­4­phenylpyridinium (MPP+) for 24 h. The expression levels of circular RNA sterile α motif domain containing 4A (circSAMD4A) and microRNA (miR)­29c­3p in both midbrain tissues and SH­SY5Y cells were detected via reverse transcription­quantitative PCR. The interaction between circSAMD4A and miR­29c­3p was verified using a dual­luciferase reporter experiment. Apoptosis­, autophagy­ and 5'AMP­activated protein kinase (AMPK)/mTOR cascade­associated proteins in midbrain tissues and SH­SY5Y cells were detected using western blotting. Furthermore, TUNEL staining and flow cytometry were used to analyze cell apoptosis. It was found that circSAMD4A was upregulated, while miR­29c­3p was downregulated in both PD animal and cellular models. Moreover, circSAMD4A directly targeted and negatively regulated miR­29c­3p. Further studies identified that circSAMD4A knockdown inhibited MPTP­ or MPP+­induced apoptosis and autophagy; however, these effects were abolished by an miR­29c­3p inhibitor. In addition, circSAMD4A knockdown repressed phosphorylated­AMPK expression and increased mTOR expression in MPTP­ or MPP+­induced PD models, the effects of which were reversed by a miR­29c­3p inhibitor. Collectively, these results suggested that circSAMD4A participated in the apoptosis and autophagy of dopaminergic neurons by modulating the AMPK/mTOR cascade via miR­29c­3p in PD.

Prohibitin, STAT3 and SH2D4A physically and functionally interact in tumor cell mitochondria.

Chromosome 8p is frequently deleted in various cancer entities and has been shown to correlate with poor patient survival. SH2D4A is located on chromosome 8p and prevents the nuclear translocation of the pro-tumorigenic transcription factor STAT3. Here, we investigated the interaction of SH2D4A and STAT3 to shed light on the non-canonical functions of STAT3 in cooperation with the tumor suppressor SH2D4A. Using an immunoprecipitation-mass spectrometry (IP-MS) approach, we identified the mitochondrial scaffold proteins prohibitin 1 (PHB1) and prohibitin 2 (PHB2) among other proteins to potentially bind to SH2D4A. Co-immunoprecipitation and proximity ligation assays confirmed direct interactions of STAT3, PHB1, and SH2D4A in situ and in vitro. In addition, cell fractionation and immunofluorescence staining revealed co-localization of these proteins with mitochondria. These interactions were selectively interrupted by the small molecule and PHB ligand FL3. Furthermore, FL3 led to a reduction of STAT3 protein levels, STAT3 transcriptional activity, and HIF1α protein stabilization upon dimethyloxalylglycine (DMOG) treatment. Besides, mitochondrial fusion and fission markers, L-OPA1, Mfn1, and FIS1, were dysregulated upon FL3 treatment. This dysregulated morphology was accompanied by significant reduction of mitochondrial respiration, thus, FL3 significantly diminished mitochondrial respirational capacity. In contrast, SH2D4A knockout increased mitochondrial respiration, whereas FL3 reversed the effect of SH2D4A knockout. The here described results indicate that the interaction of SH2D4A and PHB1 is involved in the mitochondrial function and integrity. The demonstrated interaction with STAT3, accompanied by its reduction of transcriptional activity, further suggests that SH2D4A is linking STAT3 to its mitochondrial functions, and inhibition of PHB-interaction may have therapeutic effects in tumor cells with STAT3 activation.

The tumor suppressor Zinc finger protein 471 suppresses breast cancer growth and metastasis through inhibiting AKT and Wnt/ß-catenin signaling.

BACKGROUND: Zinc-finger protein 471 (ZNF471) is a member of the Krüppel-associated box domain zinc finger protein (KRAB-ZFP) family. ZNF471 is methylated in squamous cell carcinomas of tongue, stomach and esophageal. However, its role in breast carcinogenesis remains elusive. Here, we studied its expression, functions, and molecular mechanisms in breast cancer. METHODS: We examined ZNF471 expression by RT-PCR and qPCR. Methylation-specific PCR determined its promoter methylation. Its biological functions and related molecular mechanisms were assessed by CCK-8, clonogenicity, wound healing, Transwell, nude mice tumorigenicity, flow cytometry, BrdU-ELISA, immunohistochemistry and Western blot assays. RESULTS: ZNF471 was significantly downregulated in breast cell lines and tissues due to its promoter CpG methylation, compared with normal mammary epithelial cells and paired surgical-margin tissues. Ectopic expression of ZNF471 substantially inhibited breast tumor cell growth in vitro and in vivo, arrested cell cycle at S phase, and promoted cell apoptosis, as well as suppressed metastasis. Further knockdown of ZNF471 verified its tumor-suppressive effects. We also found that ZNF471 exerted its tumor-suppressive functions through suppressing epithelial-mesenchymal transition, tumor cell stemness and AKT and Wnt/ß-catenin signaling. CONCLUSIONS: ZNF471 functions as a tumor suppressor that was epigenetically inactivated in breast cancer. Its inhibition of AKT and Wnt/ß-catenin signaling pathways is one of the mechanisms underlying its anti-cancer effects.

BCL11B suppresses tumor progression and stem cell traits in hepatocellular carcinoma by restoring p53 signaling activity.

Accumulating evidence indicates that hepatocellular carcinoma (HCC) tumorigenesis, recurrence, metastasis, and therapeutic resistance are strongly associated with liver cancer stem cells (CSCs), a rare subpopulation of highly tumorigenic cells with self-renewal capacity and differentiation potential. Previous studies identified B cell leukemia/lymphoma-11b (BCL11B) as a novel tumor suppressor with impressive capacity to restrain CSC traits. However, the implications of BCL11B in HCC remain unclear. In this study, we found that low BCL11B expression was an independent indicator for shorter overall survival (OS) and time to recurrence (TTR) for HCC patients with surgical resection. In vitro and in vivo experiments confirmed BCL11B as a tumor suppressor in HCC with inhibitory effects on proliferation, cell cycle progression, apoptosis, and mobility. Furthermore, BCL11B could suppress CSC traits, as evidenced by dramatically decreased tumor spheroid formation, self-renewal potential and drug resistance. A Cignal Finder Array and dual-luciferase activity reporter assays revealed that BCL11B could activate the transcription of P73 via an E2F1-dependent manner. Thus, we concluded that BCL11B is a strong suppressor of retaining CSC traits in HCC. Ectopic expression of BCL11B might be a promising strategy for anti-HCC treatment with the potential to cure HBV-related HCC regardless of P53 mutation status.

Targeting prohibitin with small molecules to promote melanogenesis and apoptosis in melanoma cells.

Prohibitins 1 and 2 (PHB1/2) are scaffold proteins that are involved in both melanogenesis and oncogenic pathways. We hypothesized that a PHB1 ligand, melanogenin, may display anti-cancer effects in addition to its known melanogenic activity in melanocytes. Here, we disclose a convenient synthesis of melanogenin, and its analogs. We found that, among 57 new melanogenin analogs, two (Mel9 and Mel41) significantly promoted both melanogenesis in melanocytes by activating one of the PHB2-interacting proteins, microtubule-associated protein light chain 3 (LC3), and upregulating the expression of microphthalmia associated transcription factor (MITF). These analogs also activate ERK. Besides, in addition to their promelanogenic activities, we uncovered that melanogenin and its active analogs induce apoptosis in several cancer cell lines, including melanoma cells, and that this effect is caused by an inhibition of AKT survival pathway. Our findings present a new putative function for PHBs as regulators of LC3/ERK/MITF melanogenic signaling, and suggest that Mel9 and Mel41 may provide the basis for the development of new drugs candidates to treat melanoma and other types of cancers.

Human papilloma virus-specific T cells can be generated from naïve T cells for use as an immunotherapeutic strategy for immunocompromised patients.

Human papilloma virus (HPV) is a known cause of cervical cancer, squamous cell carcinoma and laryngeal cancer. Although treatments exist for HPV-associated malignancies, patients unresponsive to these therapies have a poor prognosis. Recent findings from vaccine studies suggest that T-cell immunity is essential for disease control. Because Epstein-Barr Virus (EBV)-specific T cells have been highly successful in treating or preventing EBV-associated tumors, we hypothesized that the development of a manufacturing platform for HPV-specific T cells from healthy donors could be used in a third-party setting to treat patients with high-risk/relapsed HPV-associated cancers. Most protocols for generating virus-specific T cells require prior exposure of the donor to the targeted virus and, because the seroprevalence of high-risk HPV types varies greatly by age and ethnicity, manufacturing of donor-derived HPV-specific T cells has proven challenging. We, therefore, made systematic changes to our current Good Manufacturing Practice (GMP)-compliant protocols to improve antigen presentation, priming and expansion for the manufacture of high-efficacy HPV-specific T cells. Like others, we found that current methodologies fail to expand HPV-specific T cells from most healthy donors. By optimizing dendritic cell maturation and function with lipopolysaccharide (LPS) and interferon (IFN)γ, adding interleukin (IL)-21 during priming and depleting memory T cells, we achieved reliable expansion of T cells specific for oncoproteins E6 and E7 to clinically relevant amounts (mean, 578-fold expansion; n = 10), which were polyfunctional based on cytokine multiplex analysis. In the third-party setting, such HPV-specific T-cell products might serve as a potent salvage therapy for patients with HPV-associated diseases.

HPV16 E6 Promotes Breast Cancer Proliferation via Upregulation of COX-2 Expression.

Background. Breast cancer remains the leading cause of cancer-related mortality worldwide. It has been indicated that human papillomaviruses 16 (HPV16) might participate in the pathogenesis and development of breast cancer. However, the detected rate of HPV16 varies with region. We will investigate HPV16 E6 expression in North China and explore the effects and mechanism of HPV16 E6 on breast cancer proliferation in this study. Methods. The expressions of HPV16 E6 and COX-2 in paraffin-embedded tissues of the invasive ductal breast cancer were detected by qPCR and IHC. The effects of HPV16 E6 on breast cancer proliferation were determined by function studies. The mechanism of HPV16 E6 in promoting breast cancer proliferation was explored by Western blot and Dual-Luciferase Reporter Assay. Results. HPV16 E6 was positive in 28% invasive ductal breast carcinoma in North China; HPV16 E6 promoted breast cancer proliferation. Inhibition of COX-2 by siCOX-2 or Celecoxib attenuated the proliferation of breast cancer cells with HPV16 E6 expression; and the upregulation of COX-2 could be suppressed by the inhibition of NF-κB activity. Conclusion. HPV16 E6 promotes breast cancer proliferation by activation of NF-κB signaling pathway and increase of COX-2 expression. COX-2 will be a potential target for HPV16 E6-associated breast cancer.