Proliferation Of Human Cancer Cells Research Articles

Ionizable lipid nanoparticles for RAS protease delivery to inhibit cancer cell proliferation

Mutations in RAS, a family of proteins found in all human cells, drive a third of cancers, including many pancreatic, colorectal, and lung cancers. However, there is a lack of clinical therapies that can effectively prevent RAS from causing tumor growth. Recently, a protease was engineered that specifically degrades active RAS, offering a promising new tool for treating these cancers. However, like many other intracellularly acting protein-based therapies, this protease requires a delivery vector to reach its site of action within the cell. In this study, we explored the incorporation of cationic lipids into ionizable lipid nanoparticles (LNPs) to develop a RAS protease delivery platform capable of inhibiting cancer cell proliferation in vitro and in vivo. A library of 13 LNPs encapsulating RAS protease was designed, and each formulation was evaluated for in vitro delivery efficiency and toxicity. A subset of four top-performing LNP formulations was identified and further evaluated for their impact on cancer cell proliferation in human colorectal cancer cells with mutated KRAS in vitro and in vivo, as well as their in vivo biodistribution and toxicity. In vivo, both the concentration of cationic lipid and type of cargo influenced LNP and cargo distribution. All lead candidate LNPs showed RAS protease functionality in vitro, and the top-performing formulation achieved effective intracellular RAS protease delivery in vivo, decreasing cancer cell proliferation in an in vivo xenograft model and significantly reducing tumor growth and size. Overall, this work demonstrates the use of LNPs as an effective delivery platform for RAS proteases, which could potentially be utilized for cancer therapies.

Structural Characteristic, Strong Antioxidant, and Anti-Gastric Cancer Investigations on an Oleoresin from Ginger (Zingiber officinale var. roscoe).

It is known that ginger oleoresin contains various active components and possesses bioactivities. In this study, ginger oleoresin from Chinese ginger (Zingiber officinale var. roscoe) was extracted using a CO2 supercritical fluid extraction method with a 0.52% yield (g/g), based on dry weights. Zingiberene with a content of 51.6 mg/g was the main volatile in the ginger oleoresin. In total, 17 phenolic compounds were identified, and their contents were calculated as 587.54 mg/g. Among them, a new gingertriol was detected in the Z. officinale. Antioxidant activity tests showed that the ginger oleoresin and six gingerols exhibited strong scavenging free radical activities, and the zingerone exhibited the strongest antioxidant activity, with IC50 values of 11.3 µg/mL for the 2, 2'-diphenyl-1-picrylhydrazyl radical and 19.0 µg/mL for the 2, 2'-amino-di (2-ethyl-benzothiazoline sulphonic acid-6) ammonium salt radical cation, comparable to vitamin C. Ginger oleoresin inhibits HGC-27 human gastric cancer cell proliferation at a rate of 4.05~41.69% and induces cell apoptosis at a rate of 10.4~20.9%. The Western blot result demonstrated that the AKT signaling pathway has the potential mechanism of ginger oleoresin acting on HGC-27 cells. The anticancer potential of the gingerol standards on HGC-27 cells followed the order of 8-gingerol > 6-gingerol > 10-gingerol > zingerone. The different antioxidant and anticancer potentials of the ginger phenolic compounds could be attributed to the presence of hydroxyl groups in the unbranched 1-alkyl chain and the length of carbon side chain. Consequently, ginger oleoresin shows substantial antioxidant and anticancer therapeutic potential and can be used for novel food-drug development.

Brexpiprazole Prevents the Malignant Progression of Human Colorectal Cancer Cells and Increases Its Sensitivity to EGFR Inhibition.

Colorectal cancer (CRC) is a major cause of mortality and morbidity. A study proved that brexpiprazole, as a novel dopamine receptor partial agonist, can also prevent CRC cell proliferation. Therefore, clarifying the molecular mechanism of brexpiprazole is vital to developing a novel therapeutic strategy for CRC. The effect of brexpiprazole on human colorectal cancer cell proliferation was measured with Cell Counting Kit-8 (CCK-8) kits. Cell migration capability was measured using wound healing and transwell. Cell apoptosis was evaluated with a flow cytometer. Western blots and immunohistochemical staining were used to evaluate protein expression. The effects observed in vitro were also confirmed in xenograft models. Brexpiprazole remarkably inhibited the proliferation, suppressed the migration ability, and induced apoptosis of colorectal cancer cells. Mechanism study showed that brexpiprazole exerted these effects by inhibiting the EGFR pathway. Brexpiprazole enhanced HCT116 cells' sensitivity to cetuximab, and a combination of brexpiprazole and cetuximab inhibited xenograft tumor growth in vivo. Our finding suggested that brexpiprazole inhibits proliferation, promotes apoptosis, and enhances CRC cells' sensitivity to cetuximab by regulating the EGFR pathway and it might be an efficacious treatment strategy for CRC.

Retraction: Celastrol inhibit the proliferation, invasion and migration of human cervical HeLa cancer cells through down-regulation of MMP-2 and MMP-9.

Zhang J, Wang R, Cheng L, Xu H. Celastrol inhibit the proliferation, invasion and migration of human cervical HeLa cancer cells through down-regulation of MMP-2 and MMP-9. J Cell Mol Med. 2021;25:5335-5338. doi: 10.1111/jcmm.16488 The above article, published online on 4 May 2021 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor-in-Chief, Stefan N. Constantinescu, and the Foundation for Cellular and Molecular Medicine/John Wiley & Sons Ltd. Following publication, concerns were raised by third parties that portions of Figure1 are identical to Figure1 presented in another article (1) from a different author group and covering a different research topic. Further investigation confirmed these concerns and revealed additional duplication in Figures1 and 2, overlapping with Figures1 and 3 in (1). The authors of both articles were approached for an explanation but the authors of this article did not respond. The authors of (1) were unable to provide any underlying data. The retraction has been agreed because of concerns regarding the integrity of the research presented given that the figures were duplicated. (1) Shi X, Luo X, Chen T, etal. Naringenin inhibits migration, invasion, induces apoptosis in human lung cancer cells and arrests tumour progression invitro. J Cell Mol Med. 2021;25:2563-2571. doi: 10.1111/jcmm.16226.

Retraction: Naringenin inhibits migration, invasion, induces apoptosis in human lung cancer cells and arrests tumour progression invitro.

Shi X, Luo X, Chen T, Guo W, Liang C, Tang S, Mo J. Naringenin inhibits migration, invasion, induces apoptosis in human lung cancer cells and arrests tumour progression invitro. J Cell Mol Med. 2021; 25: 2563-2571. https://doi.org/10.1111/jcmm.16226 The above article, published online on 1 February 2021 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor-in-Chief, Stefan N. Constantinescu, and the Foundation for Cellular and Molecular Medicine/John Wiley & Sons Ltd. Following publication, concerns were raised by third parties that portions of Figure1 are identical to Figure1 in another article (1) from a different author group and covering a different research topic. Further investigation confirmed these concerns and revealed additional duplication in Figures1 and 3, overlapping with Figures1 and 2 in (1). The authors of both articles were approached for an explanation but were unable to address the concerns raised. The authors of (1) did not respond, and the authors of this article were unable to provide any underlying data. The retraction has been agreed because of concerns regarding the integrity of the research presented given that the figures were duplicated. (1) Zhang J, Wang R, Cheng L, Xu H. Celastrol inhibit the proliferation, invasion and migration of human cervical HeLa cancer cells through down-regulation of MMP-2 and MMP-9. J Cell Mol Med. 2021; 25: 5335-5338. https://doi.org/10.1111/jcmm.16488.

Inhibitory effects of medium-chain fatty acids on the proliferation of human breast cancer cells via suppression of Akt/mTOR pathway and modulating the Bcl-2 family protein.

Medium-chain fatty acids (MCFAs) have 6-12 carbon atoms and are instantly absorbed into the bloodstream before traveling to the portal vein and the liver, where they are immediately used for energy and may have antitumor effects. Its role in breast cancer is poorly understood. To investigate the apoptosis-inducing effect of MCFAs in breast cancer cells, cell viability assay, colony formation assay, cell migration assay, cell invasion assay, nuclear morphology, cell cycle assay, intracellular reactive oxygen species (ROS), matrix metalloproteinase (MMP), apoptosis, RT-qPCR analysis, and Western blot analysis were performed. In the present study, MCFA treatments reduced proliferative capability, increased ROS level, increased the depletion of MMP, induced G0/G1 and S phase cell cycle arrest, and late apoptosis of breast cancer cells in an effective concentration. Besides, MCFA treatment contributed to the upregulation of proapoptotic protein (BAK) and caspase-3, and the downregulation of antiapoptotic protein (Bcl-2). Mechanistically, phosphorylation levels of EGFR, Akt, and mTOR were significantly reduced in breast cancer cells treated with MCFAs. However, no significant changes in apoptosis and signaling-related proteins were observed in lauric acid-treated ER-positive cancer cells. Our findings suggested that MCFAs suppressed breast cancer cell proliferation by modulating the PI3K/Akt/mTOR signaling pathway. MCFAs may be a promising therapeutic drug for treating breast cancer.

Toxoplasma gondii suppresses proliferation and migration of breast cancer cells by regulating their transcriptome

BackgroundBreast cancer is the most common cancer in women worldwide. Toxoplasma gondii (T. gondii) has shown anticancer activity in breast cancer mouse models, and exerted beneficial effect on the survival of breast cancer patients, but the mechanism was unclear.MethodsThe effect of tachyzoites of T. gondii (RH and ME49 strains) on human breast cancer cells (MCF-7 and MDA-MB-231 cells) proliferation and migration was assessed using cell growth curve and wound healing assays. Dual RNA-seq was performed for T. gondii-infected and non-infected cells to determine the differentially expressed genes (DEGs). Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Protein–Protein Interaction Networks analysis (PPI) were performed to explore the related signaling pathway and hub genes. Hub genes were validated using the Kaplan–Meier plotter database, and Pathogen Host Interaction (PHI-base) database. The results were verified by qRT-PCR.ResultsThe tachyzoites of T. gondii decreased the expression of Ki67 and increased the expression of E-cadherin, resulting in suppressing the proliferation and migration of infected human breast cancer cells. The inhibitory effect of T. gondii on breast cancer cells showed a significant dose–response relationship. Compared with the control group, 2321 genes were transcriptionally regulated in MCF-7 cells infected with T. gondii, while 169 genes were transcriptionally regulated in infected MDA-MB-231 cells. Among these genes, 698 genes in infected MCF-7 cells and 67 genes in infected MDA-MB-231 cells were validated by the publicly available database. GO and KEGG analyses suggested that several pathways were involved in anticancer function of T. gondii, such as ribosome, interleukin-17 signaling, coronavirus disease pathway, and breast cancer pathway. BRCA1, MYC and IL-6 were identified as the top three hub genes in infected-breast cancer cells based on the connectivity of PPI analysis. In addition, after interacting with breast cancer cells, the expression of ROP16 and ROP18 in T. gondii increased, while the expression of crt, TgIST, GRA15, GRA24 and MIC13 decreased.ConclusionsT. gondii transcriptionally regulates several signaling pathways by altering the hub genes such as BRCA1, MYC and IL-6, which can inhibit the breast tumor growth and migration, hinting at a potential therapeutic strategy.

ELK3 destabilization by speckle-type POZ protein suppresses prostate cancer progression and docetaxel resistance.

Accumulating evidence demonstrates that the activity regulation of ELK3, a member of the E26 transformation-specific oncogene family, is critical to regulating cell proliferation, migration, and survival in human cancers. However, the molecular mechanisms of how ELK3 induces chemoresistance in prostate cancer (PCa) have not been elucidated. In this study, we found that SPOP and ELK3 are an interacting partner. The interaction between SPOP and ELK3 resulted in increased ELK3 ubiquitination and destruction, assisted by checkpoint kinase-mediated ELK3 phosphorylation. Notably, the modulation of SPOP-mediated ELK3 protein stability affected the c-Fos-induced cell proliferation and invasion of PCa cells. The clinical involvement of the SPOP-ELK3 axis in PCa development was confirmed by an immunohistochemical assay on 123 PCa tissues, with an inverse correlation between increased ELK3 and decreased SPOP being present in ~80% of the specimens. This observation was supported by immunohistochemistry analysis using a SPOP-mutant PCa specimen. Finally, docetaxel treatment induced cell death by activating checkpoint kinase- and SPOP-mediated ELK3 degradation, while SPOP-depleted or SPOP-mutated PCa cells showed cell death resistance. Notably, this observation was correlated with the protein levels of ELK3. Taken together, our study reveals the precise mechanism of SPOP-mediated degradation of ELK3 and provides evidence that SPOP mutations contribute to docetaxel resistance in PCa.

Bergapten attenuates human papillary thyroid cancer cell proliferation by triggering apoptosis and the GSK-3β, P13K and AKT pathways.

Over the past few decades, thyroid cancer (TC) incidence has steadily increased globally. The most common TC is human papillary thyroid carcinoma (PTC), which is poorly responsive to the current treatments. Hence, finding a successful therapeutic is urgently required. Bergapten (BG) is a furanocoumarin, a natural psoralen derivative isolated from numerous species of citrus and bergamot oil that has demonstrated anti-tumor activity. However, there are no reports available on the efficacy of BG on PTC cells. The current research investigated the anti-cancer activity of BG on human BCPAP cells, with cytotoxicity and apoptosis evaluated using MTT assay, AO/EB, DAPI, PI, ELISA, mRNA, and western blot. Bergapten (control group, 10 μM/mL and 15 μM/mL) inhibited PTC cell proliferation and stimulated apoptosis by enhancing Bax and caspase and reducing Bcl-2, cyclin-D1, c-myc, and survivin in a dose-dependent manner. Furthermore, BG expressively attenuated PI3K/AKT/GSK-3β signaling, creating an uneven Bax/Bcl-2 ratio that triggered Cyt-c, caspase cascade and apoptosis in human PTC cells. Our findings emphasize that BG has the potential to be used as a protective natural remedy for human PTC cells.

Compound 225# inhibits the proliferation of human colorectal cancer cells by promoting cell cycle arrest and apoptosis induction.

Colorectal cancer (CRC) ranks as the second leading cause of cancer‑related death worldwide due to its aggressive nature. After surgical resection, >50% of patients with CRC require adjuvant therapy. As a result, eradicating cancer cells with medications is a promising method to treat patients with CRC. In the present study, a novel compound was synthesized, which was termed compound 225#. The inhibitory activity of compound 225# against CRC was determined by MTT assay, EdU fluorescence labeling and colony formation assay; the effects of compound 225# on the cell cycle progression and apoptosis of CRC cells were detected by flow cytometry and western blotting; and the changes in autophagic flux after the administration of compound 225# were detected using the double fluorescence fusion protein mCherry‑GFP‑LC3B and western blotting. The results demonstrated that compound 225# exhibited antiproliferative properties, inhibiting the proliferation and expansion of CRC cell lines in a time‑ and dose‑dependent manner. Furthermore, compound 225# triggered G2/M cell cycle arrest by influencing the expression of cell cycle regulators, such as CDK1, cyclin A1 and cyclin B1, which is also closely related to the activation of DNA damage pathways. The cleavage of PARP and increased protein expression levels of PUMA suggested that apoptosis was triggered after treatment with compound 225#. Moreover, the increase in LC3‑II expression and stimulation of autophagic flux indicated the activation of an autophagy pathway. Notably, compound 225# induced autophagy, which was associated with endoplasmic reticulum (ER) stress. In accordance with the in vitro findings, the in vivo results demonstrated that compound 225# effectively inhibited the growth of HCT116 tumors in mice without causing any changes in their body weight. Collectively, the present results demonstrated that compound 225# not only inhibited proliferation and promoted G2/M‑phase cell cycle arrest and apoptosis, but also initiated cytoprotective autophagy in CRC cells by activating ER stress pathways. Taken together, these findings provide an experimental basis for the evaluation of compound 225# as a novel potential medication for CRC treatment.

Ginsenoside Re Suppress the Proliferation and Migration of MCF-7 Human Breast Cancer Cells

Ginsenoside Re Suppress the Proliferation and Migration of MCF-7 Human Breast Cancer Cells

Abstract 4377: AT rich interaction domain 5a promotes gastrointestinal tumorigenesis by regulating YAP and STAT3 pathways

Abstract Background: Chronic inflammation and an inflammatory cytokine, interleukin(IL)-6, play important roles in gastrointestinal tumorigenesis and metastasis. Recently, autocrine IL-6 has been reported to promote cancer development and progression in gastrointestinal cancer. However, it is still unknown how IL-6 level is regulated in gastrointestinal cancer cells. RNA-binding proteins (RBPs) are known to be one of the regulatory mechanisms of cytokine production, including IL-6, especially in immune cells. AT rich interaction domain 5a (Arid5a), one of the RBPs, has been reported to bind to IL6 3’UTR and regulates its stability and expression. Therefore, we aimed to elucidate the role of Arid5a in gastrointestinal cancer, especially gastric and colon cancer. Methods: Human gastric and colon cancer cell lines were used in this study. Arid5a knocked-down gastric and colon cancer cells were generated with lentiviral transduction of shRNAs targeting Arid5a and their phenotypes were investigated by several assays including MTT, apoptosis and sphere formation assays. Bulk RNA-seq analysis was performed using Arid5a knocked-down gastric and colon cancer cells. Results: In both human gastric and colon cancer cell lines, Arid5a knockdown suppressed cancer cell proliferation and spheroid formation, and increased apoptosis. Unexpectedly, IL6 mRNA expression was not correlated with Arid5a expression level. We found that Arid5a knockdown suppresses the activation of YAP and STAT3 by western blot analysis. We identified several new Arid5a target genes by bulk RNA-seq analysis. Conclusion: Arid5a plays an important role in the regulation of cancer cell proliferation, apoptosis and stemness in human gastric and colon cancer cells, perhaps independently of IL-6. Arid5a regulates YAP and STAT3 pathways by unknown mechanisms. We are now investigating how Arid5a regulates YAP and STAT3 pathways by focusing new Arid5a target genes. Citation Format: Koji Taniguchi, Kazuya Hamada, Shugo Tanaka, Hiroyuki Kurosu, Issei Kawakita, Kentaro Kumagai, Kentaro Nakazono, Sari Iwasaki, Satoshi Tanaka. AT rich interaction domain 5a promotes gastrointestinal tumorigenesis by regulating YAP and STAT3 pathways [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4377.

Effects of Viable and Heat-Inactivated Bifidobacterium longum D42 on Proliferation and Apoptosis of HT-29 Human Colon Cancer Cells.

Bifidobacterium longum is a common probiotic; both viable and heat-inactivated Bifidobacterium longum have many probiotic effects, such as anticancer effects. But some mechanisms of anticancer effects are still unclear, especially for heat-inactivated probiotics. In this study, we analyzed the effects of viable and heat-inactivated Bifidobacterium longum D42 on human colon cancer cells (HT-29). Cell proliferation, membrane permeability and apoptosis were detected by using the CCK-8 method, LDH method and Annexin V-FITC/PI kits. The ROS level and mitochondrial membrane potential were examined using the fluorescent probes DCFH-DA and JC-1. Real-time fluorescence quantitative PCR (RT-qPCR) and Western blot were used to detect the expression of mitochondrial apoptosis pathway genes and proteins. The results showed that viable and heat-inactivated Bifidobacterium longum D42 at concentrations of 1 × 106 CFU/mL significantly inhibited the proliferation of and increased the level of LDH release of HT-29 colon cancer cells. We found that they could increase the apoptosis rate of HT-29 cells. Moreover, they could also induce apoptosis by inducing cells to produce ROS and destroying the mitochondrial membrane potential of cells. Further studies found that they could increase the mRNA transcription and protein expression levels of the Caspase-3, Caspase-9 and Bax genes in cells, and reduce the mRNA transcription and protein expression levels of the Bcl-2 gene. In summary, our findings revealed that viable and heat-inactivated Bifidobacterium longum D42 have inhibitory effects on proliferation and promote the apoptosis of human colon cancer cells, and also have certain adjuvant drug therapeutic effects and have potential application value in the adjuvant treatment of colon cancer.

Targeted inhibition of colorectal cancer proliferation: Thedual-modulatory role of 2,4-DTBP on anti-apoptotic Bcl-2 and Survivin proteins.

The anti-apoptotic proteins, Bcl-2 and Survivin, are consistently overexpressed in numerous human malignancies, notably in colorectal cancer. 2,4-Di-tert-butylphenol (2,4-DTBP) is a naturally occurring phenolic compound known for its diverse biological activities, including anti-cancer properties. The mechanism behind 2,4-DTBP-induced inhibition of cell proliferation and apoptosis in human colorectal cancer cells, specifically regarding Bcl-2 and Survivin, remains to be elucidated. In this study, we employed both in silico and invitro methodologies to underpin this interaction at the molecular level. Molecular docking demonstrated a substantial binding affinity of 2,4-DTBP towards Bcl-2 (ΔG = -9.8 kcal/mol) and Survivin (ΔG = -5.6 kcal/mol), suggesting a potential inhibitory effect. Further, molecular dynamic simulations complemented by MM-GBSA calculations confirmed the significant binding of 2,4-DTBP with Bcl-2 (dGbind = -54.85 ± 6.79 kcal/mol) and Survivin (dGbind = -32.36 ± 1.29 kcal/mol). Invitro assays using HCT116 colorectal cancer cells revealed that 2,4-DTBP inhibited proliferation and promoted apoptosis in both a dose- and time-dependent manner. Fluorescence imaging and scanning electron microscopy illustrated the classical features associated with apoptosis upon 2,4-DTBP exposure. Cell cycle analysis through flow cytometry highlighted a G1 phase arrest and apoptosis assay demonstrated increased apoptotic cell population. Notably, western blotting results indicated a decreased expression of Bcl-2 and Survivin post-treatment. Considering the cytoprotective roles of Bcl-2 and Survivin through the inhibition of mitochondrial dysfunction, our findings of disrupted mitochondrial bioenergetics, characterized by reduced ATP production and oxygen consumption, further accentuate the functional impairment of these proteins. Overall, the integration of in silico and invitro data suggests that 2,4-DTBP holds promise as a therapeutic agent targeting Bcl-2 and Survivin in colorectal cancer.

Isoalantolactone inhibits the proliferation of human liver cancer cells by inducing intrinsic apoptosis

Purpose: To investigate isoalantolactone's potential as an anticancer agent targeting liver cancer cells and to elucidate the underlying mechanism. 
 Methods: Cell counting kit-8 (CCK-8) and colony formation assays were employed to analyze the anti-growth effects of isoalantolactone against liver cancer cells. Cell apoptosis was studied using Acridine orange/ethidium bromide (AO/EB) and Annexin V-FITC/Propidium iodide (PI) staining methods. The intracellular levels of reactive oxygen species (ROS) were estimated using the 2’-7’-Dichlorodihydrofluorescein diacetate (DCF-DA) method. Liver cancer cell invasion was assessed through Transwell assays. 
 Results: Isoalantolactone inhibited the proliferation and colony formation of HuH7 cells by inducing apoptosis. Isoalantolactone showed IC50 of 9 μM against HuH7 liver cancer cells. The MRC-5 normal cells treated with isoalantolactone also showed loss of viability and the IC50 was estimated to be 40 μM. HuH7 cancer cells administered with isoalantolactone exhibited modulation of apoptotic marker protein expression levels. Apoptosis was shown to result from ROS elevation. Isoalantolactone also restricted liver cancer cell invasion. 
 Conclusion: Isoalantolactone shows anti-proliferative and anti-metastatic effects against liver cancer cells via ROS-mediated apoptosis induction thereby making it a potential source of potent therapeutic agents against human cancer.

CLCN3 in mediating the proliferation of human ovarian cancer cells.

Chloride channel-3 (CLCN3), a crucial component of the voltage-gated chloride channel family, is implicated in numerous physiological and pathophysiological processes. This study aimed to investigate the characteristics of CLCN3 in pancancer and its influence on the immune response through the use of a range of databases. Concurrently, we assessed the impact of CLCN3 on the proliferation of ovarian cancer (OC) cells and explored its potential mechanisms. We employed the Tumor Immune Estimation Resource (TIMER) 2.0 and Clinical Proteomic Tumor Analysis Consortium (CPTAC) databases to examine the messenger RNA (mRNA) and the protein expression of CLCN3 across various cancers. The prognostic significance of CLCN3 was evaluated using the Gene Expression Profiling Interactive Analysis 2.0 (GEPIA 2.0) database. The University of Alabama at Birmingham Cancer Data Analysis Portal (UALCAN) facilitated the analysis of CLCN3 promoter methylation levels. The association between CLCN3 expression and tumor-infiltrating immune cells was investigated using various algorithms. The cBioportal database facilitated the analysis of CLCN3 mutations and mutation sites across various cancers. The Tumor-Immune System Interactions Database (TISIDB) database was employed to explore the correlation between CLCN3 expression and immune or molecular subtypes across a variety of cancer types. We collected ovarian tissue samples, encompassing both normal ovarian and OC tissues. The human OC cell lines, SKOV3 cells and OVCAR433 cells, were cultured. CLCN3 expression was determined via reverse-transcription quantitative polymerase chain reaction (RT-qPCR), while phosphatidylinositol 3-kinase/Akt kinase (PI3K/AKT) expression was detected using Western blot. We utilized small interfering RNA (siRNA) technology to suppress CLCN3 expression. The proliferative capacity of SKOV3 and OVCAR433 cells was assessed using the Cell Counting Kit 8 (CCK-8) assay. CLCN3 demonstrated an aberrant expression in a number of cancer types and was markedly reduced in OC tissues. Poor prognosis in cervical squamous cell cancer and myeloid leukemia was linked to excessive expression of CLCN3. The examination of immune cell infiltration, which included CD8+ T cells, B cells, T regulatory cells, and cancer-associated fibroblast cells, showed a strong association with aberrant CLCN3 expression. Following the use of siRNA technology, the ability of the ovarian carcinoma cell line SKOV3 and OVCAR433 to proliferate as well as the expression of PI3K/AKT both increased. CLCN3 is a possible biomarker for immune-related processes and the prognosis of cancer, and the PI3K/AKT signaling pathway may affect OC cells' ability to proliferate.

MiR-128-3p inhibits the proliferation, migration and invasion of human nasopharyngeal carcinoma cells via EMT

Purpose: To investigate the effect of overexpressing miR-128-3p on the migration, invasion, and proliferation of human nasopharyngeal cancer cells, and associated pathways.Methods: Cell counting kit-8 (CCK-8) and clone creation tests were used to investigate the effect of miR-128-3p inhibitor on the growth of C666-1 cells. The inhibitor control and miR-128-3p inhibitor were transfected into human nasopharyngeal cancer cells (C666-1) and allowed to incubate for 48 h. Cell migration and invasion assays were conducted using Transwell and Scratch assays. Cell cycle was assessed using propidium iodide (PI) single-staining method, while expression of miR-128-3p was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Expression levels of vimentin, E-cadherin, and N-cadherin were determined by Western blot.Results: MiR-128-3p inhibitor transfection in C666-1 cells significantly increased cell proliferation, cell viability, and clonal proliferation, and also significantly reduced miR-128-3p expression levels, and enhanced cell migration and invasion. In C666-1 cells, inhibition of miR-128-3p induced epithelial mesenchymal transition (EMT).Conclusion: MiR-128-3p inhibits nasopharyngeal cancer cells from proliferating, migrating, and invading by regulating epithelial mesenchymal transition. MiR-128-3p may therefore be a viable therapeutic target for nasopharyngeal cancer. Future studies are required to yield pertinent data in support of this hypothesis.

MiRNA-449c-5p regulates the JAK-STAT pathway in inhibiting cell proliferation and invasion in human breast cancer cells by targeting ERBB2.

Breast cancer is a highly prevalent disease worldwide, and early diagnosis and treatment could reduce the mortality rate of breast cancer patients. microRNAs (miRNA) have been shown to regulate the occurrences and progression of many types of cancers. Thus, it is crucial to find novel biomarkers in breast cancer. miR-449c-5p acted as a biomarker in non-small cell lung cancer, gastric carcinoma, and so forth. ERBB2 is an ideal target for breast cancer therapy. However, the molecular mechanisms between miR-449c-5p and ERBB2 in breast cancer remain poorly understood. Our study focused on the regulatory role of miR-449c-5p in breast cancer and its targeting relationship with ERBB2. The miR-449c-5p expression in breast cancer tissue and normal tissue was searched from the online database (Starbase). The clinical prognosis of miR-449c-5p and ERBB2 was predicted by using the Kaplan-Meier analysis method. The expression of miR-449c-5p mimics and inhibitors was measured by qRT-PCR. T47D cells were transfected with miR-449c-5p mimics and miR-449c-5p inhibitors. After that, CCK-8, colony formation assays and Transwell assays were used to evaluate the cell proliferation ability, migration and invasion. Whether ERBB2 was the target gene of the miR-449c-5p was predicted by Starbase and verified by dual-luciferase activity assay. In addition, protein levels and the relationship between signalling pathways were measured and validated using western blotting analysis. We confirmed that miR-449c-5p was highly expressed in breast cancer tissue, and its downregulation was linked with poor prognosis. Overexpression of miR-449c-5p inhibited the proliferation, migration and invasion of breast cancer cells. ERBB2 was a target of miR-449c-5p. The invasion, migration, and proliferation of breast cancer cells were inhibited by miR-449c-5p/ERBB2 through JAK-STAT. This study demonstrated that miR-449c-5p inhibits breast cancer cell proliferation, migration and invasion by targeting ERBB2 via JAK/STAT, which means miR-449c-5p, is a potential biomarker for breast cancer and provides a novel insight for diagnosis.

Abstract A076: The role of PARP-1-mediated FoxA1 ADP-ribosylation in breast cancer biology

Abstract Breast cancer is the most common cancer diagnosed in women. At the time of diagnosis, approximately 70% of breast cancers express Estrogen Receptor alpha (ERα), a key signature of luminal subtype tumors. Equally important in these cases is the expression of FoxA1, a subtype-specific transcription factor (TF) essential for ERα-mediated cell proliferation. FoxA1 is a key pioneer TF that assists ERα with the assembly of enhancers and the transcription of estrogen (E2)-regulated target genes. Together, the FoxA1-ERα axis is a major driver of cancer cell growth. Our recently published work implicates poly(ADP-ribose) polymerase 1 (PARP-1) and ADP-ribosylation (ADPRylation) in the regulation of E2-dependent transcriptional responses in ERα+ breast cancer cells. To determine the underlying mechanisms, we identified FoxA1 as a substrate of PARP-1-mediated ADPRylation. We hypothesize that (1) PARP-1 elicits its effects on E2 signaling and ERα enhancer function, in part, by ADPRylating FoxA1 at specific residues and (2) Loss of these regulatory ADPRylation events alters FoxA1 activity, which in turn can alter the assembly of ERα enhancers and transform the breast cancer cell transcriptome. Indeed, the sites of FoxA1 ADPRylation are recurrently mutated in breast cancers and are highly enriched in metastatic cases. Using an integrated set of biochemical, molecular, cell based, genomic and proteomic approaches, we have validated the ADPRylation of FoxA1 by PARP-1 in vitro and in vivo. We selected two FoxA1 ADPRylation sites, D226 and E255, for further analysis due to their positions in a mutation ‘hot spot’ region corresponding to the C-terminal forkhead DNA-binding domain. Mutation of these sites blocks FoxA1 ADPRylation and enhances cell proliferation in MCF-7 human breast cancer cells. Additionally, preliminary ChIP-seq analysis indicates that the binding of both ERα and FoxA1 are globally reduced at ERα enhancers in MCF-7 cells expressing the FoxA1 ADPRylation site mutants compared to wild-type but may also increase both at selected enhancers. While there is a global reduction in mutant FoxA1 binding near the canonical estrogen-regulated genes, interestingly, we observe an overall enhanced binding of FoxA1 D226N compared to wild-type outside of this canonical program. Collectively, these studies will (1) reveal new molecular endpoints modulated by PARP-1, providing new insights about the use of PARPi to treat ERα+ breast cancers, and (2) suggest new therapeutic strategies to disrupt the aberrant signaling pathways associated with mutant FoxA1 luminal breast cancers, and more importantly metastatic cases, which is of utmost clinical significance. Citation Format: Cristel V. Camacho, Aishwarya Sundaresan, Tulip Nandu, W. Lee Kraus. The role of PARP-1-mediated FoxA1 ADP-ribosylation in breast cancer biology [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A076.

ZNF692 promotes cell proliferation, invasion and migration of human prostate cancer cells by targeting the EMT signaling pathway

BackgroundProstate cancer poses a considerable threat to human health. At present, the mechanism of tumor progression remains unclear. ZNF692 is overexpressed in many tumors, and the high expression of ZNF692 is correlated with tumor aggressiveness and tumor phenotype of prostate cancer, suggesting that ZNF692 may play an important role in tumor biology of prostate cancer. This paper aims to elucidate the relationship between them.MethodsThe expression level of ZNF692 was verified in normal prostate cells (RWPE-1) and prostate cancer cells (LNCaP, PC3, DU145). PC3 cells were selected to construct the ZNF692 knockout prostate cancer cell line. The changes of cell proliferation, apoptosis, invasion and metastasis were detected by CCK8, Edu staining, Transwell assay and scratch assay. The expression levels of related proteins were detected by Western blot.ResultsAt the cellular level, ZNF692 was overexpressed to varying degrees in prostate cancer cell lines, with the highest expression in PC3 cell lines. CCK8 and Edu results showed that the proliferation of prostate cancer PC3 cells that knocked down ZNF692 was slowed. Transwell assay and scratch assay showed reduced invasion and migration of prostate cancer PC3 cells that knocked out ZNF692. Flow cytometry showed that the apoptosis rate of prostate cancer PC3 cells after ZNF692 knockout was increased. In addition, after ZNF692 silencing, the expression level of epithelial phenotype E-cadherin increased in PC3 cells, while the expression level of interstitial phenotype N-cadherin, Vimentin, c-Myc, and CyclinA1 decreased. The state of prostate cancer PC3 cells that overexpressed ZNF692 was reversed from the state after ZNF692 was knocked down.ConclusionZNF692 can be used as a new prognostic marker and a potential biologic therapeutic target for PCa. By inhibiting the expression of c-myc and cyclinA1, the EMT signaling pathway is regulated to provide evidence for its potential molecular mechanism.