Invasive Potential Of Cancer Cells Research Articles

Clematis erecta Extract Inhibits Migration, Invasion and Induce Apoptosis in Breast Cancer Cells

Migration and invasion of breast cancer cells to distant parts of the body is a hallmark of the disease. The disease becomes difficult to manage and treat with an increased invasion by cancer cells. Some drugs kill the cancer cells but causes general cytotoxicity but no drugs are known to inhibit the invasive potential of cancer cells. Previously, the natural components have been proven to exhibit anticancer and invasive potential. Clematis erecta L. (Ranunculaceae) Leaves infusion is traditionally used to treat syphilitic, cancerous and other foul ulcers. Moreover, methanolic extract and and ethyl acetate fraction exhibited significant analgesic and anti-inflammatory activity. The scientific literature still lacks support for the anticancer potential of C. erecta. Therefore, it was envisaged to investigate the anticancer activities of C. erecta aerial parts on breast cancer cells. The results obtained suggest that C. erecta has anti-invasive potential on triple-negative human breast cancer cells (MDA-MB-231). Three different extracts (chloroform, methanol and water) of aerial parts of C. erecta were evaluated for their effect on the growth and migration of human breast cancer cells MDA-MB-231. Interestingly, aqueous extract inhibits cell growth by more than 50% and also inhibits migration and invasion by 40 and 50%, respectively. DNA fragmentation of extract treated cells further suggested that C. erecta has the potential to kill cancer cells.

NALCN-mediated sodium influx confers metastatic prostate cancer cell invasiveness.

There is growing evidence that ion channels are critically involved in cancer cell invasiveness and metastasis. However, the molecular mechanisms of ion signaling promoting cancer behavior are poorly understood and the complexity of the underlying remodeling during metastasis remains to be explored. Here, using a variety of in vitro and in vivo techniques, we show that metastatic prostate cancer cells acquire a specific Na+ /Ca2+ signature required for persistent invasion. We identify the Na+ leak channel, NALCN, which is overexpressed in metastatic prostate cancer, as a major initiator and regulator of Ca2+ oscillations required for invadopodia formation. Indeed, NALCN-mediated Na+ influx into cancer cells maintains intracellular Ca2+ oscillations via a specific chain of ion transport proteins including plasmalemmal and mitochondrial Na+ /Ca2+ exchangers, SERCA and store-operated channels. This signaling cascade promotes activity of the NACLN-colocalized proto-oncogene Src kinase, actin remodeling and secretion of proteolytic enzymes, thus increasing cancer cell invasive potential and metastatic lesions in vivo. Overall, our findings provide new insights into an ion signaling pathway specific for metastatic cells where NALCN acts as persistent invasion controller.

STRUCTURAL ORGANIZATION OF COLLAGEN-CONTAINING CONNECTIVE TISSUE AS A PREDICTOR OF METASTATIC PROGRESSION OF PROSTATE CANCER

Summary. Aim: to determine at the optical level the role of the fibrillar organization of the collagen-containing connective tissue component of the primary tumor focus in prostate cancer (PCa) progression. Objects and methods: the morphological study is based on the analysis of the histological material of the primary tumor foci of 55 PCa patients without tumor progression in the postoperative period and bone metastases. The tumors were graded Gleason 6–9 at surgery. The features of the architecture of collagen-containing connective tissue in tumors were determined in histological slides stained by Van-Gieson. Results: the remodeling of the collagen-containing stromal component located both around the glandular structures (increase in the percentage of straight and aligned collagen fibers compared to the curved ones) and in the stroma itself has been found. Such remodeling is manifested by an increase in the total mass of fibrous structures, an increase in the width of collagen fibrils, their compaction in relation to the density of their location, peculiarities of desmoplasia, alignment and elongation. The specified features are consistent with the Gleason score and the postoperative disease course, namely, the occurrence of metastases. Conclusions: the desmoplastically changed collagen-containing connective tissue component of the PCa creates favorable conditions for the unimpeded migration and realization of the invasive potential of cancer cells, in particular those expressing bone tissue remodeling proteins.

The binding between NPM and H2B proteins signals for the diabetes-associated centrosome amplification.

Diabetes not only increases the risk for cancer but also promotes cancer metastasis. Centrosome amplification (CA) is sufficient to initiate tumorigenesis and can enhance the invasion potential of cancer cells. We have reported that diabetes can induce CA, with diabetic pathophysiological factors as the triggers, which involves the signaling of nucleophosmin(NPM). Thus, CA can serve as a candidate biological link between diabetes and cancer. In the present study, we attempted to identify the NPMbinding partners and investigated whether the binding between NPMand its partner mediated the CA. We confirmed that high glucose, insulin, and palmitic acid cancer could elicit CA in the HCT16 colon cancer cells and found that the experimental treatment increased the binding between NPMand H2B, but not between p-NPMand H2B. The molecular docking analysis supported the fact that NPMand H2B could bind to each other through various amino acid residues. The treatment also increased the colocalization of NPMand H2B in the cytosol. Importantly, disruption of the NPM1-H2B complex by individualknockdown of the protein level of NPMor H2B led to the inhibition of the treatment-evoked CA. In conclusion, our results suggest that the binding between NPMand H2B proteins signals for the CA by high glucose, insulin, and palmitic acid.

Hexavalent chromium induces centrosome amplification through ROS-ATF6-PLK4 pathway in colon cancer cells.

Centrosome amplification (CA) refers to a numerical increase in centrosomes resulting in cells with more than two centrosomes. CA has been shown to initiate tumorigenesis and increase the invasive potential of cancer cells in genetically modified experimental models. Hexavalent chromium is a recognized carcinogen that causes CA and tumorigenesis as well as promotes cancer metastasis. Thus, CA appears to be a biological link between chromium and cancer. In the present study, we investigated how chromium triggers CA. Our results showed that a subtoxic concentration of chromium-induced CA in HCT116 colon cancer cells, resulted in the production of reactive oxygen species (ROS), activated ATF6 without causing endoplasmic reticulum stress, and upregulated the protein level of PLK4. Inhibition of ROS production, ATF6 activation, or PLK4 upregulation attenuated CA. Inhibition of ROS using N-acetyl-l-cysteine (NAC) inhibited chromium-induced activation of ATF6 and upregulation of PLK4. ATF6-specific siRNA knocked down the protein level and activation of ATF6, and upregulated PLK4, with no effect on ROS production. Knockdown of PLK4 protein had no effect on chromium-induced ROS production or activation of ATF6. In conclusion, our results suggest that hexavalent chromium induces CA via the ROS-ATF6-PLK4 pathway and provides molecular targets for inhibiting chromium-mediated CA, which may be useful for the assessment of CA in chromium-promoted tumorigenesis and cancer cell metastasis.

Royal Jelly and trans-10-Hydroxy-2-Decenoic Acid Inhibit Migration and Invasion of Colorectal Carcinoma Cells.

SUMMARYResearch backgroundAcquisition of migratory potential is pivotal for cancer cells, enabling invasion and metastasis of colorectal carcinoma. Royal jelly and its bioactive component trans-10-hydroxy-2-decenoic acid (10H2DA) showed remarkable antimetastatic potential, but the molecular mechanism underlying this activity is unclear.Experimental approachIdentification and quantification of 10H2DA in royal jelly originating from Serbia was done by HPLC method. Cytotoxicity of 10H2DA was measured by tetrazolium dye MTT test in concentration range 1-500 μg/mL after 24 and 72 h. Its effect on the collective and single-cell migration was measured by wound healing and transwell migration assays. Invasive potential of cancer cells was evaluated by a transwell method modified with collagen. Immunofluorescence was used for migratory and invasive protein expression, while the gene expression of these markers was evaluated by quantitative real time polymerase chain reaction (qRT-PCR). All assays were applied on human colorectal carcinoma HCT-116 and SW-480 cell lines and, except for MTT, evaluated after 24 h of treatment with two selected concentrations of royal jelly and 10H2DA.Results and conclusionsAccording to HPLC, the mass fraction of 10H2DA in royal jelly was 0.92% (m/m). Treatment with 10H2DA showed no cytotoxic effect; however, significant inhibitory potential of royal jelly and 10H2DA on the motility and invasiveness of colorectal cancer cells was observed. More pronounced effect was exerted by 10H2DA, which significantly suppressed collective cell migration and invasiveness of SW-480 cells, as well as single- and collective cell migration and invasive potential of HCT-116 cell line. Treatments increased epithelial markers E-cadherin and cytoplasmic β-catenin in HCT-116 cells, thus stabilizing intercellular connections. In SW-480 cells, 10H2DA increased E-cadherin on protein and gene level, and suppressed epithelial-mesenchymal transition (EMT) markers. In both cell lines, treatments induced significant suppression of promigratory/proinvasive markers: N-cadherin, vimentin and Snail on protein and gene level, which explains decreased migratory and invasive potential of HCT-116 and SW-480 cells.Novelty and scientific contributionOur study presents new findings and elucidation of royal jelly and 10H2DA molecular mechanism that underlies their antimigratory/antiinvasive activity on colorectal cancer cells. These findings are shown for the first time indicating that these natural products are a valuable source of anticancer potential and should be reconsidered for further antitumour therapy.

Tumor Targeting by Conditioned Medium Derived From Human Amniotic Membrane: New Insight in Breast Cancer Therapy.

Objectives:Traditional breast cancer treatments have challenges including inefficiency, multidrug resistance, severe side effects, and targeting non-specifically. The development of alternative treatment strategies has attracted a great deal of interest. Using the amniotic membrane has become a promising and convenient new approach for cancer therapy. This study aimed to evaluate the anti-cancer ability of conditioned medium extracted from the human amniotic membrane (hAM-CM) on breast cancer cells.Methods:Conditioned medium was collected after 48 h incubation of hAM in epithelial up manner. MTT, cell cycle, apoptosis, colony formation, and sphere assays were used to determine the impact of hAM-CM on breast cancer cell lines. The effects of hAM-CM on the migration and invasion of breast cancer cells were determined using scratch wound healing and transwell assays, respectively.Results:Based on the results, cell viability was significantly decreased by hAM-CM in a dose-dependent manner. The hAM-CM remarkably induced apoptosis and necrosis of cancer cells. Moreover, cell migration and invasion potential of cancer cells decreased after the hAM-CM treatment. Further, both the number of colonies and their morphologies were affected by the treatment. In the treated group, a significant decrease in the number of colonies along with an obvious change in their morphologies from holoclone shape to a dominant paracolone structure was observed.Conclusion:Our results indicate that the conditioned medium derived from the human amniotic membrane able to inhibit proliferation and metastasis of tumor cells and can be considered a natural and valuable candidate for breast cancer therapy.

Sustained IKKβ phosphorylation and NF-κB activation by superoxide-induced peroxynitrite-mediated nitrotyrosine modification of B56γ3 and PP2A inactivation

Apart from its physiological role in inflammation and immunity, the nuclear factor-kappa B (NF-κB) protein complex has been implicated in tumorigenesis and its progression. Here, we provide evidence that a pro-oxidant milieu is an upstream effector of oncogenic NF-κB signaling. Through pharmacological or genetic inhibition of SOD1, we show that elevated intracellular superoxide (O2-) mediates sustained IKK phosphorylation, and induces downstream degradation of IκBα, leading to the nuclear localization and transcriptional activation of NF-κB. Mechanistically, we show that such sustained NF-κB signaling is a function of protein phosphatase 2A (PP2A) inactivation brought about by the nitrative modification of its substrate-binding sub-unit B56γ. Importantly, the pro-oxidant driven NF-κB activation enhances the migratory and invasive potential of cancer cells. In summary, our work highlights the critical involvement of O2--dependent peroxynitrite production in inhibiting PP2A-mediated dephosphorylation of IKK, thereby facilitating cancers to acquire an invasive phenotype. Given that NF-κB is a key player of chronic inflammation and carcinogenesis, our work unravels a novel synergistic node involving O2--driven redox milieu and deregulated PP2A as a potential therapeutic target.

Genistein inhibits radiation-induced invasion and migration of glioblastoma cells by blocking the DNA-PKcs/Akt2/Rac1 signaling pathway

Background and purposeRadiotherapy is the most important therapeutic measure against glioblastoma multiforme (GBM), which is regarded as the most common and highly lethal type of brain cancer. Nevertheless, most relapses originate in the close vicinity of the irradiated target volume. Genistein is a natural product that can suppress the invasive potential of cancer cells. In this study, DNA-dependent protein kinase catalytic subunit (DNA-PKcs)-proficient and -deficient GBM cells were selected for in vitro and in vivo studies to investigate the inhibiting effects of genistein on radiation-induced invasion and migration and the corresponding mechanism. Materials and methodsGBM cell lines with or without genistein pre-treatment were irradiated with X-rays. Cell survival was determined using colony formation assay and the rate of cellular proliferation was analyzed with a real-time cell electronic sensing system. For in vitro study, invasion and migration abilities were evaluated via wound-healing and transwell assays, while protein expression was determined with western blotting. Genistein interaction with DNA-PKcs was estimated with pull-down, recombinant and binding assays. For in vivo study, cells were stereotactically injected into NOD-SCID mice to establish tumors. Hematoxylin and eosin and immunohistochemistry were used to assess the invasive potential of GBM. ResultsX-ray irradiation enhanced the migration and invasion of DNA-PKcs-positive but not DNA-PKcs-negative GBM cells. It also activated the DNA-PKcs/Akt2/Rac1 signaling pathway, which contributed to GBM malignant progression by aggravating GBM cell invasive potential. The study successfully demonstrated that genistein can specifically bind to DNA-PKcs and block the DNA-PKcs/Akt2/Rac1 pathway, thereby effectively inhibiting radiation-induced invasion and migration of GBM cells in vitro and in vivo. The present study emphasized that radiation-induced invasive potential is initiated by DNA-PKcs, which is a well-known double strand breaks (DSB) repair protein, and determined the exact site for genistein binding to DNA-PKcs. ConclusionDNA-PKcs is not only a potential target for cancer therapy, but also a reliable biomarker for predicting radiation-induced invasion and migration of GBM cells. Thus, genistein might serve as a novel therapeutic strategy for treating GBM.

ITLN1 modulates invasive potential and metabolic reprogramming of ovarian cancer cells in omental microenvironment

Advanced ovarian cancer usually spreads to the omentum. However, the omental cell-derived molecular determinants modulating its progression have not been thoroughly characterized. Here, we show that circulating ITLN1 has prognostic significance in patients with advanced ovarian cancer. Further studies demonstrate that ITLN1 suppresses lactotransferrin’s effect on ovarian cancer cell invasion potential and proliferation by decreasing MMP1 expression and inducing a metabolic shift in metastatic ovarian cancer cells. Additionally, ovarian cancer-bearing mice treated with ITLN1 demonstrate marked decrease in tumor growth rates. These data suggest that downregulation of mesothelial cell-derived ITLN1 in the omental tumor microenvironment facilitates ovarian cancer progression.

A 3D biophysical model for cancer spheroid cell-enhanced invasion in collagen-oriented fiber microenvironment**Project supported by the National Natural Science Foundation of China (Grant Nos. 11974066 and 11674043), the Fundamental Research Funds for the Central Universities, China (Grant No. 2019CDYGYB007), and the Natural Science Foundation of Chongqing, China (Grant No. cstc2019jcyj-msxmX0477).

The process of in situ tumors developing into malignant tumors and exhibiting invasive behavior is extremely complicated . From a biophysical point of view, it is a phase change process affected by many factors, including cell-to-cell, cell-to-chemical material, cell-to-environment interaction, etc. In this study, we constructed spheroids based on green fluorescence metastatic breast cancer cells MDA-MB-231 to simulate malignant tumors in vitro, while constructed a three-dimensional (3D) biochip to simulate a micro-environment for the growth and invasion of spheroids. In the experiment, the 3D spheroid was implanted into the chip, and the oriented collagen fibers controlled by collagen concentration and injection rate could guide the MDA-MB-231 cells in the spheroid to undergo directional invasion. The experiment showed that the oriented fibers greatly accelerated the invasion speed of MDA-MB-231 cells compared with the traditional uniform tumor micro-environment, namely obvious invasive branches appeared on the spheroids within 24 hours. In order to analyze this interesting phenomenon, we have developed a quantitative analyzing approach to explore strong angle correlation between the orientation of collagen fibers and invasive direction of cancer cell. The results showed that the oriented collagen fibers produced by the chip can greatly stimulate the invasion potential of cancer cells. This biochip is not only conducive to modeling cancer cell metastasis and studying cell invasion mechanisms, but also has the potential to build a quantitative evaluation platform that can be used in future chemical drug treatments.

Telomerase Reverse Transcriptase (TERT) Expression, Telomerase Activity, and Expression of Matrix Metalloproteinases (MMP)-1/-2/-9 in Feline Oral Squamous Cell Carcinoma Cell Lines Associated With Felis catus Papillomavirus Type-2 Infection.

Telomerase activity contributes to cell immortalization by avoiding telomere shortening at each cell division; indeed, its catalytic subunit telomerase reverse transcriptase (TERT) is overexpressed in many tumors, including human oral squamous cell carcinoma (hOSCC). In these tumors, matrix metalloproteinases (MMPs), a group of zinc-dependent endopeptidases involved in cell migration, contribute to invasive potential of cancer cells. A proportion of hOSCC is associated with infection by high-risk human papillomavirus (HR-HPVs), whose E6 oncogene enhances TERT and MMPs expression, thus promoting cancer progression. Feline oral squamous cell carcinoma (FOSCC) is a malignant tumor with highly invasive phenotype; however, studies on telomerase activity, TERT, and MMPs expression are scarce. In this study, we demonstrate telomerase activity, expression of TERT, and its transcriptional activator cMyc along with expression of MMP-1, -2, and -9 in FOSCC-derived cell lines SCCF2 and SCCF3, suggesting a contribution by these pathways in cell immortalization and invasion in these tumors. Recent studies suggest that a sub-group of FOSCC as well as SCCF2 and SCCF3 are associated with Felis catus PV type-2 (FcaPV-2) infection. However, in this work, FcaPV-2 E6 gene knock-down caused no shift in either TERT, cMyc, or MMPs levels, suggesting that, unlike its human counterpart, the viral oncogene plays no role in their regulation.

Epithelial Plasticity, Autophagy and Metastasis: Potential Modifiers of the Crosstalk to Overcome Therapeutic Resistance.

Epithelial-to-mesenchymal transition (EMT) initiates malignant transformation of cancer cells and is responsible for the generation of heterogenic subsets of cancer stem cells (CSCs). Signals in the form of environmental cues and paracrine factors within tumor microenvironment (TME) niche, support the possibility of generation of pool of CSCs with two distinct functional transition states. Cyclic CSCs with predominant epithelial phenotype, self-renew and differentiate into mature cancer cells. Subsets of autophagic/ non-cyclic CSCs with predominant mesenchymal phenotype have capacity to invade, metastasize, resist to apoptosis, escape immunosurveillance, survive chemotherapies and are majorly responsible for cancer mortality. Differences in phenotypic plasticity may form the basis of differential impact of therapeutic outcomes on heterogeneous subpopulations of CSCs. Activation of autophagy is responsible for the recycling of damaged organelles and protein aggregates, regulates EMT, confers the survival advantage to neoplastic cells to anti-cancer therapies, significantly affects the invasive potential of cancer cells and supports their metastatic dissemination in a tissue and tumor stage dependent manner. Therapy resistance is the primary obstacle in the complete ablation of tumor cells. Combinational treatments based on targeting autophagic CSCs and inhibiting EMT regulators may represent potential anticancer strategies for the prevention of cancer invasion, metastatic spread and disease relapse.

Characterization of the Effects of Migrastatic Inhibitors on 3D Tumor Spheroid Invasion by High-resolution Confocal Microscopy.

Drug discovery and development in cancer research is increasingly being based on drug screens in a 3D format. Novel inhibitors targeting the migratory and invasive potential of cancer cells, and consequently the metastatic spread of disease, are being discovered and considered as complementary treatments in highly invasive cancers such as gliomas. Thus, generating data enabling the detailed analyses of cells in a 3D environment following the addition of a drug is required. The methodology described here, combining spheroid invasion assays with high-resolution image capture and data analysis by confocal laser scanning microscopy (CLSM), enabled detailed characterization of the effects of the potential anti-migratory inhibitor MI-192 on glioma cells. Spheroids were generated from cell lines for invasion assays in low adherent 96-well plates and then prepared for CLSM analysis. The described workflow was preferred over other commonly used spheroid-generating techniques due to both ease and reproducibility. This, combined with the enhanced image resolution attained by confocal microscopy compared to conventional wide-field approaches, allowed the identification and analysis of distinct morphological changes in migratory cells in a 3D environment following treatment with the migrastatic drug MI-192.

Stilbenoid-Mediated Epigenetic Activation of Semaphorin 3A in Breast Cancer Cells Involves Changes in Dynamic Interactions of DNA with DNMT3A and NF1C Transcription Factor.

Loci-specific increase in DNA methylation occurs in cancer and may underlie gene silencing. It is investigated whether dietary stilbenoids, resveratrol, and pterostilbene exert time-dependent effects on DNA methylation patterns and specifically methylation-silenced tumor suppressor genes in breast cancer cells. Following genome-wide DNA methylation analysis with Illumina-450K, changes characteristic of early and late response to stilbenoids are identified. Interestingly, often the same genes but at different CpG loci, the same gene families, or the same functional gene categories are affected. CpG loci that lose methylation in exposed cells correspond to genes functionally associated with cancer suppression. There is a group of genes, including SEMA3A, at which the magnitude of hypomethylation in response to stilbenoids rises with increasing invasive potential of cancer cells. Decreased DNA methylation at SEMA3A promoter and concomitant gene upregulation coincide with increased occupancy of active histone marks. Open chromatin upon exposure to stilbenoids may be linked to decreased DNMT3A binding followed by increased NF1C transcription factor occupancy. Sequestration of DNMT3A is possibly a result of stilbenoid-mediated increase in SALL3 expression, which was previously shown to bind and inhibit DNMT3A activity. The findings define mechanistic players in stilbenoid-mediated epigenetic reactivation of genes suppressing cancer.

Scutellarin inhibits the invasive potential of malignant melanoma cells through the suppression epithelial-mesenchymal transition and angiogenesis via the PI3K/Akt/mTOR signaling pathway

Malignant melanoma is the leading cause of death from skin disease, due in large part to its propensity to metastasize. Scutellarin is an active flavone extracted from traditional Chinese herb Erigeron breviscapus (Vant.) Hand. Mazz. Recent studies have reported that scutellarin can be utilized to treat various types of tumors. In this study, we investigated the effects of scutellarin on melanoma cancer cell invasive potential and the molecular mechanisms underlying these effects using A375 melanoma cells lines. The in-vitro antitumor activity of scutellarin was evaluated by CCK-8 assay, wound-healing assay, transwell assays, adhesion assays, and tube formation assays to assess the cell viability, migration, invasion, adhesion, and angiogenesis, respectively. Also, western blotting assay was used to assess the level of PI3K/Akt/mTOR signaling pathway proteins in A375 cells. We found that scutellarin significantly inhibited melanoma cell lines and HUVECs viability in a time- and concentration-dependent manners. Additionally, scutellarin effectively suppressed tumor cell migration, invasion, adhesion through the suppression of EMT and angiogenesis by inhibiting the PI3K/Akt/mTOR signaling pathway. These results indicated that scutellarin could markedly inhibit the invasive potential of melanoma cell lines by suppressing the EMT and angiogenesis through the PI3K/Akt/mTOR signaling pathway. It suggests that scutellarin might be a potential compound in malignant melanoma treatment.

Senescence-related deterioration of intercellular junctions in the peritoneal mesothelium promotes the transmesothelial invasion of ovarian cancer cells

Mechanisms of transmesothelial invasion of ovarian cancer are still poorly understood. Here we examined whether this phenomenon may be determined by an expression of intercellular junctions in peritoneal mesothelial cells (PMCs). Analysis of ovarian tumors showed that cancer cells are localized below an intact layer of PMCs. The PMCs located near the invaded cancer cells displayed low expression of connexin 43, E-cadherin, occludin, and desmoglein, as well as expressed SA-β-Gal, a marker of senescence. Experiments in vitro showed that senescent PMCs exhibited decreased levels of the four tested intercellular junctions, and that the invasion of ovarian cancer cells through the PMCs increased proportionally to the admixture of senescent cells. Intervention studies showed that the expression of connexin 43, E-cadherin, occludin, and desmoglein in senescent PMCs could be restored upon the blockade of p38 MAPK, NF-κB, AKT, JNK, HGF, and TGF-β1. When these molecules were neutralized, the efficiency of the transmesothelial cancer cell invasion was diminished. Collectively, our findings show that the integrity of the peritoneal mesothelium, which is determined by the expression of junctional proteins, is critical for the invasion of ovarian cancer. They also indicate a mechanism by which senescent PMCs may promote the invasive potential of cancer cells.

OR34-2 SREBP1 Drives KRT80-Dependent Cytoskeletal Changes and Invasive Behaviour in Endocrine-Resistant ERα Breast Cancer

About one third of oestrogen receptor alpha breast cancer (ERα BC) patients progress to invasive metastatic disease despite targeted endocrine therapies1. Our previous study showed how breast cancer cells developing resistance to aromatase inhibitors (AI) endogenously trigger cholesterol biosynthesis potentially through activation of sterol regulatory element binding protein 1 (SREBP1) to promote sustained oestrogen independent, ERα activation2. Tumour cells are phenotypically characterized by an aberrant architecture and, although this feature is associated with cell migration and invasion, the mechanisms underlying the cytoskeletal reorganization are still poorly understood. In this study, we used breast cancer models to investigate the relationship between acquisition of drug resistance and invasive potential. We showed that cells acquiring resistance to AI undergo active cytoskeleton re-organisation via Keratin 80 (KRT80) and F-Actin remodelling. This process is driven by epigenetic reprogramming at the type II keratin locus leading to KRT80 upregulation. Chromatin Immunoprecipitation coupled with Next Generation Sequencing (ChIP-seq) revealed that reprogramming is dependent on de novo SREBP1 binding to a single enhancer that is activated upon chronic AI treatment. AI-treated patients’ samples are characterized by KRT80 cytoskeletal re-organization and an increased number of KRT80 positive cells at relapse (seventy-five human breast specimens and ten metastatic lymph nodes were selected for immunostaining with the Trust Tissue Bank approval). Using confocal microscopy imaging we found that KRT80 activation and redeployment led to increased F-actin deposition and increased focal adhesion. Additionally, results from 3D organoids invasion assay showed KRT80 manipulation directly contributed to changes in cellular stiffness and invasive potential. Intriguingly, radiological exam using shear wave elasticity imaging performed on twenty prospectively recruited cancer patients confirmed that KRT80 levels correlate with stiffer tumours in vivo. Our data strongly suggested that therapy plays a direct role in shaping the biophysical properties and invasive potential of cancer cells, by inducing epigenetic rearrangements leading to KRT80 upregulation and concomitant cytoskeletal reorganization. We also described an unexpected role for intermediate filaments in promoting cancer cell invasion by showing for the first time that KRT80 promotes actin fibre formation as well as focal adhesion maturation and lamellipodia formation. The link between epigenetic and cytoskeleton reprogramming offers an intriguing axis for drug development and biomarker discovery, especially within the goal of preventing metastatic invasion in BC patients.

MCP-1/CCL2 Mediated by Autocrine Loop of PDGF-BB Promotes Invasion of Lung Cancer Cell by Recruitment of Macrophages Via CCL2–CCR2 Axis

Platelet-derived growth factor-BB (PDGF-BB) is recognized as a potential player in a paracrine manner in tumor stroma development. PDGF-BB has an autocrine growth function in lung cancer cells; however, the mechanism in nonsmall-cell lung cancer is not fully understood. In this study, we report that PDGF-BB increased monocyte chemoattractant protein-1 (MCP-1)-dependent macrophage recruitment and that expression of metastatic genes increased in A549 cells cocultured with RAW 264.7 macrophages. Similar to exogenous PDGF-BB, PDGF-BB might have a self-stimulation in invasion of cancer cells during reciprocal activation of cancer cells and tumor-associated macrophages through secretion of soluble proteins. Also, we found that PDGF-BB upregulates both mRNA and protein level of MCP-1 expression in human A549 cells through mitogen-activated protein kinases and phosphatidylinositol 3-kinase/Akt cell signaling pathways, binding NFκB to MCP-1 promoter site and PDGF-Rβ as critical receptor. These results suggested that MCP-1/chemokine (C-C motif) ligand 2 (CCL2) expression mediated by the autocrine loop of PDGF-BB enhanced recruitment of macrophages through CCL2-CCR2 axis, which could ultimately increase expression of metastatic genes in lung cancer cells, finally promoting invasive potential of cancer cells.

Histone Deacetylase Inhibitors and Phenotypical Transformation of Cancer Cells.

Histone deacetylase inhibitors (HDIs) are a group of potent epigenetic drugs which have been investigated for their therapeutic potential in various clinical disorders, including hematological malignancies and solid tumors. Currently, several HDIs are already in clinical use and many more are on clinical trials. HDIs have shown efficacy to inhibit initiation and progression of cancer cells. Nevertheless, both pro-invasive and anti-invasive activities of HDIs have been reported, questioning their impact in carcinogenesis. The aim of this review is to compile and discuss the most recent findings on the effect of HDIs on the epithelial-mesenchymal transition (EMT) process in human cancers. We have summarized the impact of HDIs on epithelial (E-cadherin, β-catenin) and mesenchymal (N-cadherin, vimentin) markers, EMT activators (TWIST, SNAIL, SLUG, SMAD, ZEB), as well as morphology, migration and invasion potential of cancer cells. We further discuss the use of HDIs as monotherapy or in combination with existing or novel anti-neoplastic drugs in relation to changes in EMT.