Metformin Promotes Anti-tumor Biomarkers in Human Endometrial Cancer Cells

AbstractMetformin (MET) is increasingly implicated in reducing the incidence of multiple cancer types in patients with diabetes. However, similar effects of MET in non-diabetic women with endometrial cancer (EC) remain unknown. In a pilot study, obese non-diabetic women diagnosed with type 1, grade 1/2 EC, and consenting to participate were randomly assigned to receive MET or no MET (control (CON)) during the pre-surgical window between diagnosis and hysterectomy. Endometrial tumors obtained at surgery (MET, n = 4; CON, n = 4) were analyzed for proliferation (Ki67), apoptosis (TUNEL), and nuclear expression of ERα, PGR, PTEN, and KLF9 proteins in tumor glandular epithelial (GE) and stromal (ST) cells. The percentages of immunopositive cells for PGR and for KLF9 in GE and for PTEN in ST were higher while those for ERα in GE but not ST were lower, in tumors of MET vs. CON patients. The numbers of Ki67- and TUNEL-positive cells in tumor GE and ST did not differ between groups. In human Ishikawa endometrial cancer cells, MET treatment (60 μM) decreased cell numbers and elicited distinct temporal changes in ESR1, KLF9, PGR, PGR-B, KLF4, DKK1, and other tumor biomarker mRNA levels. In the context of reduced KLF9 expression (by siRNA targeting), MET rapidly amplified PGR, PGR-B, and KLF4 transcript levels. Our findings suggest that MET acts directly in EC cells to modify steroid receptor expression and signaling network and may constitute a preventative strategy against EC in high-risk non-diabetic women.

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Nuclear upregulation of PI3K p110β correlates with increased rRNA transcription in endometrial cancer cells

10.1101/2019.12.20.884122 ◽

2019 ◽

Author(s):

Fatemeh Mazloumi Gavgani ◽

Thomas Karlsson ◽

Ingvild L Tangen ◽

Andrea Papdiné Morovicz ◽

Victoria Smith Arnesen ◽

...

Keyword(s):

Endometrial Cancer ◽

Cancer Cells ◽

Cell Lines ◽

Molecular Mechanisms ◽

Pi3k Pathway ◽

Clinical Samples ◽

Mrna Levels ◽

Dependent Manner ◽

Cytoplasmic Staining ◽

Ec Cells

AbstractGenes encoding for components of the phosphoinositide 3-kinase (PI3K) pathway are frequently mutated in cancer, including inactivating mutations of PTEN and activating mutations of PIK3CA, encoding the PI3K catalytic subunit p110α. PIK3CB, encoding p110β, is rarely mutated, but can contribute to tumourigenesis in some PTEN-deficient tumours. The underlying molecular mechanisms are however poorly understood. By analysing cell lines and annotated clinical samples, we have previously found that p110β is highly expressed in endometrial cancer (EC) cell lines and that PIK3CB mRNA levels increase early in primary tumours correlating with lower survival. Selective inhibition of p110α and p110β led to different effects on cell signalling and cell function, p110α activity being correlated to cell survival in PIK3CA mutant cells and p110β with cell proliferation in PTEN-deficient cells. To understand the mechanisms governing the differential roles of these isoforms, we assessed their sub-cellular localisation. p110α was cytoplasmic whereas p110β was both cytoplasmic and nuclear with increased levels in both compartments in cancer cells. Immunohistochemistry of p110β in clinically annotated patient tumour sections revealed high nuclear/cytoplasmic staining ratio, which correlated significantly with higher grades. Consistently, the presence of high levels of p110β in the nuclei of EC cells, correlated with high levels of its product phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) in the nucleus. Using immunofluorescence labelling, we observed both p110β and PtdIns(3,4,5)P3 in the nucleoli of EC cell lines. The production of nucleolar PtdIns(3,4,5)P3 was dependent upon p110β activity. EC cells with high levels of nuclear PtdIns(3,4,5)P3 and p110β showed elevated nucleolar activity as assessed by the increase in 47S pre-rRNA transcriptional levels in a p110β-dependent manner. Altogether, these results present a nucleolar role for the PI3K pathway that may contribute to tumour progression in endometrial cancer.

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Landscape of Chimeric RNAs in Non-Cancerous Cells

Genes ◽

10.3390/genes12040466 ◽

2021 ◽

Vol 12 (4) ◽

pp. 466

Author(s):

Chen Chen ◽

Samuel Haddox ◽

Yue Tang ◽

Fujun Qin ◽

Hui Li

Keyword(s):

Cancer Cells ◽

Cell Lines ◽

Drug Targets ◽

Neoplastic Cell ◽

Multiple Cancer ◽

Chimeric Rnas ◽

Healthy Donors ◽

Cancer Types ◽

Chimeric Rna ◽

Cancerous Cells

Gene fusions and their products (RNA and protein) have been traditionally recognized as unique features of cancer cells and are used as ideal biomarkers and drug targets for multiple cancer types. However, recent studies have demonstrated that chimeric RNAs generated by intergenic alternative splicing can also be found in normal cells and tissues. In this study, we aim to identify chimeric RNAs in different non-neoplastic cell lines and investigate the landscape and expression of these novel candidate chimeric RNAs. To do so, we used HEK-293T, HUVEC, and LO2 cell lines as models, performed paired-end RNA sequencing, and conducted analyses for chimeric RNA profiles. Several filtering criteria were applied, and the landscape of chimeric RNAs was characterized at multiple levels and from various angles. Further, we experimentally validated 17 chimeric RNAs from different classifications. Finally, we examined a number of validated chimeric RNAs in different cancer and non-cancer cells, including blood from healthy donors, and demonstrated their ubiquitous expression pattern.

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Targeting RNF8 Effectively Reverse Cisplatin and Doxorubicin Resistance in Endometrial Cancer

10.21203/rs.3.rs-57263/v1 ◽

2020 ◽

Author(s):

Ben Yang ◽

Wang Ke ◽

Yingchun Wan ◽

Tao Li

Keyword(s):

Endometrial Cancer ◽

Cell Lines ◽

Quantitative Polymerase Chain Reaction ◽

Mrna Levels ◽

Mice Model ◽

Expression Levels ◽

Gynecological Malignancy ◽

Ec Cells

Abstract Background Endometrial cancer (EC) is one of the most frequent gynecological malignancy worldwide. However, resistance to chemotherapy remains one of the major difficulties in the treatment of EC. Thus, there is an urgent requirement to understand mechanisms of chemoresistance and identify novel regimens for patients with EC. Methods Cisplatin and doxorubicin resistant cell lines were acquired by continuous exposing parental EC cells to cisplatin or doxorubicin for 3 months. Cell viability was determined by using MTT assay. Protein Expression levels of protein were examined by western blotting assay. mRNA levels were measured by quantitative polymerase chain reaction (qPCR) assay. Ring finger protein 8 (RNF8) knockout cell lines were generated by clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 gene editing assay. Nonhomologous end joining (NHEJ) efficiency were quantified by plasmid based NHEJ assay. DNA double strand breaks (DSB) were generated using laser micro-irradiation. Protein recruitment to DSB was analyzed by immunofluorescent assay. Tumor growth was examined by AN3CA xenograft mice model. Results We found that protein and mRNA expression levels of RNF8 were significantly increased in both cisplatin and doxorubicin resistant EC cells. Cell survival assay showed that RNF deficiency significantly enhanced the sensitivity of resistant EC cells to cisplatin and doxorubicin (P < 0.01). In addition, chemoresistant EC cells exhibited increased NHEJ efficiency. Knockout of RNF8 in chemoresistant EC cells significantly reduced NHEJ efficiency and prolonged Ku80 retention on DSB. Moreover, cisplatin resistant AN3CA xenograft showed that RNF8 deficiency overcame cisplatin resistance. Conclusions Our in vitro and in vivo assays provide evidence for RNF8, which is a NHEJ factor, serving as a promising, novel target in EC chemotherapy.

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Noncanonical agonist PPARγ ligands modulate the response to DNA damage and sensitize cancer cells to cytotoxic chemotherapy

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1717776115 ◽

2018 ◽

Vol 115 (3) ◽

pp. 561-566 ◽

Cited By ~ 22

Author(s):

Melin J. Khandekar ◽

Alexander S. Banks ◽

Dina Laznik-Bogoslavski ◽

James P. White ◽

Jang Hyun Choi ◽

...

Keyword(s):

Dna Damage ◽

Cancer Cells ◽

Apoptotic Cell ◽

Peroxisome Proliferator ◽

Multiple Cancer ◽

P53 Signaling ◽

Response To Chemotherapy ◽

P53 Response ◽

Cancer Types ◽

Agonist Ligands

The peroxisome-proliferator receptor-γ (PPARγ) is expressed in multiple cancer types. Recently, our group has shown that PPARγ is phosphorylated on serine 273 (S273), which selectively modulates the transcriptional program controlled by this protein. PPARγ ligands, including thiazolidinediones (TZDs), block S273 phosphorylation. This activity is chemically separable from the canonical activation of the receptor by agonist ligands and, importantly, these noncanonical agonist ligands do not cause some of the known side effects of TZDs. Here, we show that phosphorylation of S273 of PPARγ occurs in cancer cells on exposure to DNA damaging agents. Blocking this phosphorylation genetically or pharmacologically increases accumulation of DNA damage, resulting in apoptotic cell death. A genetic signature of PPARγ phosphorylation is associated with worse outcomes in response to chemotherapy in human patients. Noncanonical agonist ligands sensitize lung cancer xenografts and genetically induced lung tumors to carboplatin therapy. Moreover, inhibition of this phosphorylation results in deregulation of p53 signaling, and biochemical studies show that PPARγ physically interacts with p53 in a manner dependent on S273 phosphorylation. These data implicate a role for PPARγ in modifying the p53 response to cytotoxic therapy, which can be modulated for therapeutic gain using these compounds.

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Tribbles Pseudokinase 3 Contributes to Cancer Stemness of Endometrial Cancer Cells by Regulating β-Catenin Expression

Cancers ◽

10.3390/cancers12123785 ◽

2020 ◽

Vol 12 (12) ◽

pp. 3785

Author(s):

Wen-Ling Wang ◽

Guan-Ci Hong ◽

Peng-Ju Chien ◽

Yu-Hao Huang ◽

Hsueh-Te Lee ◽

...

Keyword(s):

Endometrial Cancer ◽

Messenger Rna ◽

Tumor Development ◽

The Cancer Genome Atlas ◽

Scaffolding Protein ◽

Mrna Levels ◽

Cancer Stemness ◽

Gynecological Malignancy ◽

Ec Cells

Endometrial cancer (EC) is the second most common gynecological malignancy worldwide. Tribbles pseudokinase 3 (TRIB3) is a scaffolding protein that regulates intracellular signal transduction, and its role in tumor development is controversial. Here, we investigated the biological function of TRIB3 in EC. We found that the messenger RNA (mRNA) expression level of TRIB3 was significantly and positively correlated with shorter overall survival of EC patients in The Cancer Genome Atlas database. The protein expression of TRIB3 was found to be significantly increased in EC cancer stem cells (CSCs) enriched by tumorsphere cultivation. Knockdown of TRIB3 in EC cells suppressed tumorsphere formation, the expression of cancer stemness genes, and the in vivo tumorigenesis. The expression of β-catenin at both the protein and the mRNA levels was downregulated upon TRIB3 silencing. TRIB3 was found to interact with E74 Like ETS transcription factor 4 (ELF4) in the nucleus and bound to ELF4 consensus sites within the catenin beta 1 (CTNNB1) promoter in EC cell lines. These data indicated that TRIB3 may regulate CTNNB1 transcription by enhancing the recruitment of ELF4 to the CTNNB1 promoter. In conclusion, our results suggest that TRIB3 plays an oncogenic role in EC and positively regulates the self-renewal and tumorigenicity of EC-CSCs. Targeting TRIB3 is considered as a potential therapeutic strategy in future EC therapy.

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Impact of oestrogen and progesterone receptor expression in the cancer cells and myometrium on survival of patients with endometrial cancer

Journal of Obstetrics and Gynaecology ◽

10.1080/01443615.2017.1328591 ◽

2017 ◽

Vol 38 (1) ◽

pp. 96-102 ◽

Cited By ~ 3

Author(s):

Darko Tomica ◽

Snježana Ramić ◽

Damir Danolić ◽

Lucija Šušnjar ◽

Melita Perić-Balja ◽

...

Keyword(s):

Endometrial Cancer ◽

Progesterone Receptor ◽

Cancer Cells ◽

Receptor Expression ◽

Progesterone Receptor Expression

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Extracellular matrix regulates steady-state mRNA levels of the proliferation associated protein Ki-67 in endometrial cancer cells

Cancer Letters ◽

10.1016/s0304-3835(99)00066-x ◽

1999 ◽

Vol 140 (1-2) ◽

pp. 145-152 ◽

Cited By ~ 6

Author(s):

Marselina I Tan ◽

Elisabeth Strunck ◽

Thomas Scholzen ◽

Johannes Gerdes ◽

Günter Vollmer

Keyword(s):

Extracellular Matrix ◽

Endometrial Cancer ◽

Steady State ◽

Cancer Cells ◽

Mrna Levels ◽

Ki 67

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Regulation of PD-L1 expression in the tumor microenvironment

Journal of Hematology & Oncology ◽

10.1186/s13045-020-01027-5 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Ming Yi ◽

Mengke Niu ◽

Linping Xu ◽

Suxia Luo ◽

Kongming Wu

Keyword(s):

Cancer Cells ◽

Immune Escape ◽

Multiple Cancer ◽

Laboratory Findings ◽

Limiting Step ◽

Programmed Death ◽

Programmed Cell Death 1 ◽

Anti Cancer ◽

Potential Applications ◽

Cancer Types

AbstractProgrammed death-ligand 1 (PD-L1) on cancer cells engages with programmed cell death-1 (PD-1) on immune cells, contributing to cancer immune escape. For multiple cancer types, the PD-1/PD-L1 axis is the major speed-limiting step of the anti-cancer immune response. In this context, blocking PD-1/PD-L1 could restore T cells from exhausted status and eradicate cancer cells. However, only a subset of PD-L1 positive patients benefits from α-PD-1/PD-L1 therapies. Actually, PD-L1 expression is regulated by various factors, leading to the diverse significances of PD-L1 positivity. Understanding the mechanisms of PD-L1 regulation is helpful to select patients and enhance the treatment effect. In this review, we focused on PD-L1 regulators at the levels of transcription, post-transcription, post-translation. Besides, we discussed the potential applications of these laboratory findings in the clinic.

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ApoStream, a novel device for antibody-independent capture of circulating cancercells (CCCs) from blood of patients with various types of cancer.

Journal of Clinical Oncology ◽

10.1200/jco.2012.30.15_suppl.e13500 ◽

2012 ◽

Vol 30 (15_suppl) ◽

pp. e13500-e13500

Author(s):

Vishal Gupta ◽

Insiya Jafferji ◽

Miguel Garza ◽

Sujita Sukumaran ◽

Jacky Woo ◽

...

Keyword(s):

Cancer Patient ◽

Cancer Cells ◽

Laser Scanning ◽

Mononuclear Cells ◽

Metastatic Cancer ◽

Multiple Cancer ◽

Tumor Biopsy ◽

Peripheral Blood Mononuclear ◽

Skov3 Cells ◽

Cancer Types

e13500 Background: The detection of circulating tumor cells (CTCs) using immunomagnetic EpCAM-based capture methods has been conceptually accepted as a “liquid tumor biopsy”. However, these methods have limited the recovery of CTCs for molecular profiling applications. We developed a novel continuous flow dielectrophoresis field-flow fractionation (DEP-FFF) device, ApoStream for antibody-independent capture of circulating cancer cells (CCCs), with improved recovery across multiple cancer types and preserved viability of CCCs for downstream characterization. Methods: The performance of ApoStream was demonstrated using a low EpCAM expressing cell line, SKOV3. ApoStream was further used to enrich CCCs from various cancer patient blood. Prostate, breast and NSCLC CCCs were stained for cytokeratin (CK), CD45, and DAPI; melanoma CCCs were stained with S100, CD45 and DAPI. CCC enumeration was performed using laser scanning cytometry. Results: In system precision performance studies, average inter-day recovery on the ApoStream was 80.3 ± 3.5%, CV = 4.3% when cancer cells were spiked into buffer, and 78.5 ± 3.0%, CV = 3.3% when cancer cells spiked into ~10 million healthy peripheral blood mononuclear cells (PBMCs). Linearity performance was demonstrated with spiking 5-2600 SKOV3 cells into 10 million PBMCs (R2=1). Cell viability was not affected by processing through ApoStream device. High CCC recovery from metastatic cancer patient blood samples was obtained with counts ranging from 0 - 2630 (NSCLC, n=66), 0 - 3490 (prostate, n=29), 10 - 968 (breast, n=11), and 3 - 3120 (melanoma, n=13) CCCs per 7.5 mL blood. Positive CCC counts were obtained in 90% of NSCLC samples, 93% of prostate cancer samples, 100% breast cancer and melanoma specimens. There were no CK+ cells detected in healthy donor blood controls. Conclusions: Improved CCC recovery from various cancer types was demonstrated with the ApoStream device. ApoStream provides an antibody-independent method for capture of viable CCCs that enables further downstream molecular characterization of rare cells for use in clinical applications. Acknowledgements: Funded by NCI Contract No. HHSN261200800001E.

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