Inhibits Tumor Cell Invasion Research Articles

Coordination of AMPK and YAP by Spatholobi Caulis and Procyanidin B2 Provides Antioxidant Effects In Vitro and In Vivo.

The liver is vulnerable to oxidative attacks from heavy metals, such as iron, as well as some drugs, including acetaminophen. It has been shown that enhanced oxidative stress in the liver leads to excessive ROS production and mitochondrial dysfunction, resulting in organ injury. The beneficial effects of Spatholobi Caulis (SC), a natural herbal medicine, include treating ischemic stroke, inhibiting tumor cell invasion, pro-angiogenic activities, and anti-inflammatory properties. Scientific studies on its effects against hepatotoxic reagents (e.g., iron and acetaminophen), as well as their underlying mechanisms, are insufficient. This study examined the antioxidant effects and mechanisms of SC in vitro and in vivo. In cells, the proinflammatory mediator, arachidonic acid (AA), plus iron, significantly induced an increase in ROS generation, the damage in mitochondrial membrane potential, and the resulting apoptosis, which were markedly blocked by SC. More importantly, SC affected the activation of AMP-activated protein kinase (AMPK)-related proteins, which were vital to regulating oxidative stress in cells. In addition, SC mediated the expression of Yes-associated protein (YAP)-related proteins. Among the active compounds in SC, the procyanidin B2, but not liquiritigenin, daidzein, and genistein, significantly inhibited the cytotoxicity induced by AA + iron, and activated the LKB1-AMPK pathway. In mice, the oral administration of SC alleviated the elevations of ALT and histological changes by the acetaminophen-induced liver injury. These results reveal the potential of SC and a key bioactive component, procyanidin B2, as antioxidant candidates for hepatoprotection.

Engulfment and Cell Motility 1 (ELMO1) Regulates Tumor Cell Behavior and Predicts Prognosis in Colorectal Cancer.

Engulfment and cell motility 1 (ELMO1) plays a crucial role in the process of migration, chemotaxis, and metastasis of tumor cells. ELMO1 has been implicated in the pathogenesis of various cancers. However, the distinct function of ELMO1 in colorectal cancer (CRC) is unclear. We determined whether ELMO1 affects the oncogenic behavior of CRC cells and investigated its prognostic value in CRC patients. We investigated the impact of ELMO1 on tumor cell behavior using small interference RNA (siRNA) in CRC cell lines, including SW480 and DLD1. The expression of ELMO1 was investigated by reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, and enzyme-linked immunosorbent assay (ELISA) in cancer tissues and sera obtained from CRC patients. ELMO1 knockdown suppressed tumor cell proliferation in SW480 and DLD1 cells. ELMO1 knockdown-induced apoptosis through up-regulation of caspase-3, -7, and PARP activities and down-regulation of the anti-apoptotic Mcl-1 protein. ELMO1 knockdown-induced cell-cycle arrest by decreasing cyclin D1, cyclin-dependent kinase 2, 4 and 6, and the 25C cell division cycle (CDC25C). ELMO1 knockdown suppressed tumor cell invasion and migration. The expression of E-cadherin was increased, while that of Vimentin and Claudin 1 decreased following ELMO1 knockdown. The phosphorylation levels of PDK1, Akt, and GSK-3β and were down-regulated after ELMO1 knockdown. The expression of ELMO1 was found up-regulated in cancer tissues and sera taken from CRC patients. ELMO1 expression was significantly associated with tumor stage, lymph node metastasis, distant metastases, and poor survival. ELMO1 mediates tumor progression by increasing tumor cell motility and inhibiting apoptosis in human CRC.

Chemotherapy treatment induces pro-invasive changes in liver ECM composition

Metastasis accounts for 90% of cancer-related deaths, yet the mechanisms by which cancer cells colonize secondary organs remain poorly understood. For breast cancer patients, metastasis to the liver is associated with poor prognosis and a median survival of 6 months. Standard of care is chemotherapy, but recurrence occurs in 30% of patients. Systemic chemotherapy has been shown to induce hepatotoxicity and fibrosis, but how chemotherapy impacts the composition of the liver extracellular matrix (ECM) remains unknown. Individual ECM proteins drive tumor cell proliferation and invasion, features that are essential for metastatic outgrowth in the liver. First, we find that the ECM of livers isolated from chemotherapy-treated MMTV-PyMT mice increases the invasion, but not proliferation, of metastatic breast cancer cells. Proteomic analysis of the liver ECM identified Collagen V to be more abundant in paclitaxel-treated livers. We show that Collagen V increases cancer cell invasion via α1β1 integrins and MAPK signaling, while also increasing the alignment of Collagen I, which has been associated with increased invasion. Treatment with obtustatin, an inhibitor specific to α1β1 integrins, inhibits tumor cell invasion in decellularized ECM from paclitaxel-treated livers. Overall, we show chemotherapy treatment alters the liver microenvironment, priming it as a pro-metastatic niche for cancer metastasis.

Bone Marrow Mesenchymal Stem Cells (BMSCs)-Derived miR-206 Promotes Breast Cancer Development by Activating Hedgehog Gene Signaling

Bone marrow-derived mesenchymal stem cells (BMSCs) are an integral part of the tumor microenvironment and involved in tumor evolution. Our aim is to further illuminate the relationship of exosomes of BMSC origin and breast cancer cells in breast cancer. Differential diagnosis was performed by identifying exosomal miR-206 secreted by BMSCs, and RT-PCR detected miR-206 expression in tumor tissues. Transwell assayed cell function and Target scan analyzed the regulatory relationship between Rab23 and miR-206. Rab23 expression was examined by western-blot after the addition of Rab23 and the effect of Rab23 on hedgehog was further verified. We demonstrated that exosomal miR-206 from BMSCs is expressed in tumor tissues and miR-206 mimics significantly inhibited tumor cell invasion and proliferation. miR-206 targets Rab23 and negatively regulates its expression. Further results showed that the addition of Rab23 could activate hedgehog signaling and promote the development of breast cancer. In conclusion, our study reveals that BMSC-derived miR-206 activates hedgehog gene signaling and promotes the breast carcinogenesis development by regulating Rab23 expression.

TRIM27-USP7 complex promotes tumour progression via STAT3 activation in human hepatocellular carcinoma.

TRIM27 is stabilized by binding to USP7 and mediates tumour progression in several cancers; however, the roles of TRIM27-USP7 complex on STAT3 activation in HCC are unknown. Regulations and functions of TRIM27 for activating STAT3 in HCC were assessed using 207 HCC samples or HCC cells. TRIM27 expression was increased in some cases of HCC. High TRIM27 expression was an independent predictor for poor prognosis in HCC after surgery. It was correlated with the expression of EpCAM, vimentin, MMP-9, and activation of STAT3 in HCC. TRIM27 expression was correlated with USP7 expression, and HCC with high TRIM27 expression together with high USP7 expression showed enhanced STAT3 activation, resulting in poorer prognosis. p-JAK1 expression was correlated with STAT3 activation in HCC with high TRIM27 expression. In vitro, USP7 knockdown decreased TRIM27 expression, suggesting that USP7 was essential for TRIM27 stabilization. Knocking down of TRIM27 or USP7 suppressed STAT3 activation and overexpression of TRIM27 accelerated STAT3 activation; therefore, the formation of TRIM27-USP7 complex was needed for STAT3 activation, which led to aggressive tumour proliferation and invasion by enhancing EMT and CSC-like property. Binding of JAK1 to TRIM27-USP7 complex was confirmed in vitro. Deletion of TRIM27-USP7 complex by USP7 inhibitor significantly inhibited tumour cell invasion by suppressing STAT3 activation. TRIM27 is stabilized by binding to USP7 and is related to aggressive tumour progression in HCC via STAT3 activation, resulting in poor prognosis after operation. Therefore, TRIM27-USP7 complex is a useful prognostic predictor and a promising therapeutic target for HCC.

Optical Cellular Micromotion: A New Paradigm to Measure Tumor Cells Invasion within Gels Mimicking the 3D Tumor Environments.

Measuring tumor cell invasiveness through 3D tissues, particularly at the single-cell level, can provide important mechanistic understanding and assist in identifying therapeutic targets of tumor invasion. However, current experimental approaches, including standard in vitro invasion assays, have limited physiological relevance and offer insufficient insight into the vast heterogeneity in tumor cell migration through tissues. To address these issues, here the concept of optical cellular micromotion is reported on, where digital holographic microscopy is used to map the optical nano- to submicrometer thickness fluctuations within single-cells. These fluctuations are driven by the dynamic movement of subcellular structures including the cytoskeleton and inherently associated with the biological processes involved in cell invasion within tissues. It is experimentally demonstrated that the optical cellular micromotion correlates with tumor cells motility and invasiveness both at the population and single-cell levels. In addition, the optical cellular micromotion significantly reduced upon treatment with migrastatic drugs that inhibit tumor cell invasion. These results demonstrate that micromotion measurements can rapidly and non-invasively determine the invasive behavior of single tumor cells within tissues, yielding a new and powerful tool to assess the efficacy of approaches targeting tumor cell invasiveness.

MiR-3614-3p suppresses cell aggressiveness of human breast cancer by targeting AKT3 and HDAC1 expression.

BackgroundMicroRNAs (miRNAs) regulate various pathophysiological functions and pathobiological progression in various cancers. Our recent study reported that miR-3614-3p significantly suppressed the proliferation of breast cancer (BC) cells by downregulating its host gene TRIM25. However, other functional roles of miR-3614-3p migration and invasion in BC and its potential mechanisms are not clearly elucidated.MethodsIn this study, we investigated miR-3614-3p regulation of AKT3 and HDAC1 expression in BC. miR-3614-3p and AKT3/HDAC1 mRNA expression levels were determined using quantitative real-time PCR (qRT-PCR) in MCF-7 and MDA-MB-231 BC cells. The effects of miR-3614-3p on migration and invasion were measured using wound healing and transwell migration assays. In BC cells, miR-3614-3p levels were remarkably low, and AKT3 and HDAC1 mRNA and protein levels were high as assessed by qRT-PCR and western blot. Finally, we investigated the role of AKT3/HDAC1 using silent interfering RNA (siRNA) and confirmed the targeting of AKT3 and HDAC1 3' UTR through miR-3614-3p using a luciferase reporter assay.ResultsIn the present study, we found that overexpression of miR-3614-3p markedly suppressed tumor cell invasion and migration independent of TRIM25, whereas other target genes, AKT3 and HDAC1, were involved. Moreover, we found that the resulting silencing of AKT3 and HDAC1 suppressed cell migration.ConclusionsmiR-3614-3p is an anti-oncogene that can suppress BC cells by targeting AKT3 and HDAC1, revealing the potential role of miR-3614-3p in suppressing BC metastasis. Therefore, miR-3614 may act as a potential biomarker for BC prognosis.

Oncolytic Adenovirus with SPAG9 shRNA Driven by DD3 Promoter Improved the Efficacy of Docetaxil for Prostate Cancer.

Prostate cancer (PCa) is a common malignant tumor of the male urinary system and ranks the second in the causes of tumor-related deaths. Differential display code 3 (DD3) is a noncoding gene that is specifically expressed in PCa. High expression of sperm-associated antigen 9 (SPAG9) is closely related to tumorigenesis of PCa, and SPAG9 is a therapeutic target for PCa. In this study, a new oncolytic adenovirus DD3-ZD55-SPAG9 was constructed by using DD3 promoter to enhance the efficacy and safety of adenovirus. The combined use of DD3-ZD55-SPAG9 and docetaxel showed that DD3-ZD55-SPAG9 significantly improved the anti-tumor efficacy of docetaxel in PCa both in vitro and in vivo. The mechanism was related to the induction of tumor cell apoptosis and the inhibition of tumor cell invasion. In conclusion, DD3-ZD55-SPAG9 combined with docetaxel is an effective strategy for PCa therapy.

CircEPS15 Overexpression in Hepatocellular Carcinoma Modulates Tumor Invasion and Migration.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. Recent evidence has shown that circular RNAs (circRNAs) play important roles in tissue development, gene transcription, signal regulation and tumorigenesis. However, whether circRNAs are involved in HCC progression and encode functional proteins remains largely unknown. In the present study, we aimed to explore the function and molecular mechanism of circRNAs in HCC. First, many circRNAs were found to be differentially expressed in HCC samples and paired adjacent normal liver tissues. The validation of dysregulated circRNAs by qRT-PCR revealed that circEPS15 expression was downregulated in HCC tissues, and the survival curves showed that low circEPS15 levels were associated with poor overall survival in HCC patients. Then, the overexpression of circEPS15 suppressed tumor cell invasion and migration by inhibiting the TJP1/CDH2/VIM signaling pathway and retarded cell cycle progression, which was confirmed by the Transwell culture system, wound healing assays, flow cytometry and western blot assays. After that, the spanning junction open reading frame in circEPS15 driven by IRES was shown to encode a novel protein, which was verified by western blotting with full-length, mutated, and truncated sequences of circEPS15 with a FLAG tag. Moreover, ceRNA analysis and qRT-PCR results suggest a possible circRNA (circEPS15)-miRNA-mRNA network in HCC. Collectively, our study reveals that endogenous circEPS15 plays a novel role in repressing HCC through the ceRNA network and encodes a functional protein.

BHMPS Inhibits Breast Cancer Migration and Invasion by Disrupting Rab27a-Mediated EGFR and Fibronectin Secretion

Simple SummaryNumerous studies targeting Rab GTPases and its multiple effectors have been attempted since exocytosis has been shown to alter tumor malignancy by modulating cancer cell behavior and tumor microenvironment. Here, we demonstrated that BHMPS inhibits migration and invasion of breast cancer cells by blocking the interaction between Rab27a and Slp4. BHMPS interfered with vesicle trafficking and secretion by decreasing FAK and JNK activation. In addition, BHMPS suppressed tumor growth in Rab27a-overexpressing MDA-MB-231 xenograft mice. This study highlighted the importance of understanding the mechanisms of Rab27a-mediated metastasis in improving the therapeutic options for metastatic cancers.Our previous work demonstrated that (E)-N-benzyl-6-(2-(3, 4-dihydroxybenzylidene) hydrazinyl)-N-methylpyridine-3-sulfonamide (BHMPS), a novel synthetic inhibitor of Rab27aSlp(s) interaction, suppresses tumor cell invasion and metastasis. Here, we aimed to further investigate the mechanisms of action and biological significance of BHMPS. BHMPS decreased the expression of epithelial-mesenchymal transition transcription factors through inhibition of focal adhesion kinase and c-Jun N-terminal kinase activation, thereby reducing the migration and invasion of breast cancer. Additionally, knockdown of Rab27a inhibited tumor migration, with changes in related signaling molecules, whereas overexpression of Rab27a reversed this phenomenon. BHMPS effectively prevented the interaction of Rab27a and its effector Slp4, which was verified by co-localization, immunoprecipitation, and in situ proximity ligation assays. BHMPS decreased the secretion of epidermal growth factor receptor and fibronectin by interfering with vesicle trafficking, as indicated by increased perinuclear accumulation of CD63-positive vesicles. Moreover, administration of BHMPS suppressed tumor growth in Rab27a-overexpressing MDA-MB-231 xenograft mice. These findings suggest that BHMPS may be a promising candidate for attenuating tumor migration and invasion by blocking Rab27a-mediated exocytosis.

Research Progress on the Antitumor Mechanism of Compound Kushen Injection

Compound Kushen Injection (CKI), as a clinical traditional Chinese medicine preparation, has prominent antitumor effect but with several side effects. A large number of studies have shown that CKI plays an antitumor role by regulating tumor cell proliferation, inducing tumor cell differentiation and apoptosis, inhibiting tumor cell invasion and metastasis, reducing tumor angiogenesis, regulating the immunity, and so on. Clinically, CKI is widely used to treat various tumors, where it is often combined with surgery, chemotherapy, radiotherapy, targeted therapy, and other antitumor treatments. This article reviews the antitumor mechanism of CKI and the progress of its clinical application in order to provide a theoretical basis for further clinical application.

High Expression of the DNMT3A Embedded Pri- Mir -1301 Associates with Improved Outcome in Patients with Acute Myeloid Leukemia

Introduction: MicroRNA miR-1301 is embedded in intron 1 in the DNMT3A gene, which encodes the DNA (cytosine-5)-methyltransferase 3A, is frequently mutated in older acute myeloid leukemia (AML) patients (pts) & has been associated with clonal hematopoiesis. In glioma cells miR-1301 suppresses tumor cell invasion by regulating the activity & function of the p53 pathway & inhibits cell proliferation by targeting NRAS . The role of miR-1301 in AML remains to be elucidated.Here, we investigated the clinical & prognostic impact of pri- miR-1301 in a homogenously treated set of mainly older AML pts who received allogeneic stem cell transplantation (HSCT) as consolidation.Patients & Methods: We analyzed 126 AML pts (median age at HSCT 64 years [y]; range 38-75 y) with available pretreatment bone marrow (BM). All received HSCT after non-myeloablative (NMA) conditioning (Fludarabine 30mg/m2 at day -4 to -2 & 2Gy total body irradiation at day 0) at our institution between January 2000 & June 2012 in complete remission (CR, 83%, n=104) or CR with incomplete peripheral recovery (Cri, 17%, n=22). Cytogenetics were determined at diagnosis using standard techniques for banding & fluorescence in-situ hybridization. Using flow cytometry, BM mononuclear cells were assessed for presence of CD2, CD7, CD11b, CD13, CD14, CD15, CD33, CD34, CD38, CD45, CD56, CD61, CD64, CD65, CD117 & Glycophorin A. For pts with material available, the presence of FLT3 -ITD, FLT3 -TKD & expression status of EVI1, as well as the mutation status of the CEBPA, DNMT3A, IDH1, IDH2 &NPM1 genes were determined. Pri- miR-1301 expression was measured using a SYBR-Green assay based quantitative reverse transcription polymerase chain reaction & normalized to 18S expression as internal control. The third quartile of normalized gene expression was used to define high & low pri- miR-1301 expressers. Median follow-up after HSCT was 6.0 y for pts alive.Results: At diagnosis, high pri- miR-1301 expressers had lower white blood cell counts (P=.02). High pri- miR-1301 expression associated with higher percentage of CD7 (P=.04) & by trend of CD2 (P=.10) positive cells (both T-cell associated antigens) & lower percentage of CD15 (P=.01), CD33 (P=.002) & CD64 (P=.05) positive cells (myeloid antigens). Furthermore, high pri- miR-1301 expressers were less likely to be NPM1 mutated (P=.06) by trend. Since miR-1301 is embedded in the DNMT3A gene, we tested whether pri- miR-1301 expression associated with the presence of DNMT3A mutations, but did not observe any association (pts with DNMT3A mutations: high pri- miR-1301 expression: 18% vs . low pri- miR-1301 expression: 14%, P=.73). Pts with high pri- miR-1301 expression had longer leukemia-free survival (LFS, P=.07; Figure 1A) by trend & significantly longer overall survival (OS, P=.03; Figure 1B). In multivariate analysis, high pri- miR-1301 expression retained its prognostic impact on LFS (Hazard Ratio [HR] 0.51, Confidence Interval [CI] 0.28-0.93, P=.03) after adjustment for age at HSCT, hemoglobin level at diagnosis &EVI1 expression status at diagnosis & was the only significant factor for OS (HR 0.52, CI 0.29-0.95, P=.03).Conclusion: High pri- miR-1301 expression associated with distinct clinical & prognostic features, including higher expression of the T-cell associated antigens CD2 & CD7, lower expression of myeloid antigens CD15, CD33 & CD64 & a lower incidence of NPM1 mutations by trend. Whereas DNMT3A embedded miR-1301, no association of pri- miR-1301 expression & occurrence of DNMT3A mutation was found. Importantly, high pri- miR-1301 expressers had by trend a longer LFS & significantly longer OS independent of other prognostic markers. Further studies are warranted to elucidate the AML biology associated with high expression of the DNMT3A embedded miR-1301 in AML. [Display omitted] DisclosuresFranke:Novartis: Honoraria, Research Funding; Takeda: Consultancy; Pfizer: Honoraria. Schwind:Novartis: Consultancy.

Targeting the tetraspanin CD81 reduces cancer invasion and metastasis

Tetraspanins are an evolutionary conserved family of proteins involved in multiple aspects of cell physiology, including proliferation, migration and invasion, protein trafficking, and signal transduction; yet their detailed mechanism of action is unknown. Tetraspanins have no known natural ligands, but their engagement by antibodies has begun to reveal their role in cell biology. Studies of tetraspanin knockout mice and of germline mutations in humans have highlighted their role under normal and pathological conditions. Previously, we have shown that mice deficient in the tetraspanin CD81 developed fewer breast cancer metastases compared to their wild-type (WT) counterparts. Here, we show that a unique anti-human CD81 antibody (5A6) effectively halts invasion of triple-negative breast cancer (TNBC) cell lines. We demonstrate that 5A6 induces CD81 clustering at the cell membrane and we implicate JAM-A protein in the ability of this antibody to inhibit tumor cell invasion and migration. Furthermore, in a series of in vivo studies we demonstrate that this antibody inhibits metastases in xenograft models, as well as in syngeneic mice bearing a mouse tumor into which we knocked in the human CD81 epitope recognized by the 5A6 antibody.

Next‑generation sequencing analysis reveals that MTH‑3, a novel curcuminoid derivative, suppresses the invasion of MDA‑MB‑231 triple‑negative breast adenocarcinoma cells.

Triple-negative breast cancer (TNBC) behaves aggressively in the invasive and metastatic states. Our research group recently developed a novel curcumin derivative, (1E,3Z,6E)-3-hydroxy-5-oxohepta-1,3,6-triene-1,7-diyl)bis(2-methoxy-4,1-phenylene)bis(3-hydroxy-2-hydroxymethyl)-2-methyl propanoate (MTH-3), and previous studies showed that MTH-3 inhibits TNBC proliferation and induces apoptosis in vitro and in vivo with a superior bioavailability and absorption than curcumin. In the present study, the effects of MTH-3 on TNBC cell invasion were examined using various assays and gelatin zymography, and western blot analysis. Treatment with MTH-3 inhibited MDA-MB-231 cell invasion and migration, as shown by Transwell assay, 3D spheroid invasion assay, and wound healing assay. The results of the gelatin zymography experiments revealed that MTH-3 decreased matrix metalloproteinase-9 activity. The potential signaling pathways were revealed by next-generation sequencing analysis, antibody microarray analysis and western blot analysis. In conclusion, the results of the present study show that, MTH-3 inhibited tumor cell invasion through the MAPK/ERK/AKT signaling pathway and cell cycle regulatory cascade, providing significant information about the potential molecular mechanisms of the effects of MTH-3 on TNBC.

Cancer-associated mutations reveal a novel role for EpCAM as an inhibitor of cathepsin-L and tumor cell invasion

BackgroundEpCAM (Epithelial cell adhesion molecule) is often dysregulated in epithelial cancers. Prior studies implicate EpCAM in the regulation of oncogenic signaling pathways and epithelial-to-mesenchymal transition. It was recently demonstrated that EpCAM contains a thyroglobulin type-1 (TY-1) domain. Multiple proteins with TY-1 domains are known to inhibit cathepsin-L (CTSL), a cysteine protease that promotes tumor cell invasion and metastasis. Analysis of human cancer sequencing studies reveals that somatic EpCAM mutations are present in up to 5.1% of tested tumors.MethodsThe Catalogue of Somatic Mutations in Cancer (COSMIC) database was queried to tabulate the position and amino acid changes of cancer associated EpCAM mutations. To determine how EpCAM mutations affect cancer biology we studied C66Y, a damaging TY-1 domain mutation identified in liver cancer, as well as 13 other cancer-associated EpCAM mutations. In vitro and in vivo models were used to determine the effect of wild type (WT) and mutant EpCAM on CTSL activity and invasion. Immunoprecipitation and localization studies tested EpCAM and CTSL protein binding and determined compartmental expression patterns of EpCAM mutants.ResultsWe demonstrate that WT EpCAM, but not C66Y EpCAM, inhibits CTSL activity in vitro, and the TY-1 domain of EpCAM is responsible for this inhibition. WT EpCAM, but not C66Y EpCAM, inhibits tumor cell invasion in vitro and lung metastases in vivo. In an extended panel of human cancer cell lines, EpCAM expression is inversely correlated with CTSL activity. Previous studies have demonstrated that EpCAM germline mutations can prevent EpCAM from being expressed at the cell surface. We demonstrate that C66Y and multiple other EpCAM cancer-associated mutations prevent surface expression of EpCAM. Cancer-associated mutations that prevent EpCAM cell surface expression abrogate the ability of EpCAM to inhibit CTSL activity and tumor cell invasion.ConclusionsThese studies reveal a novel role for EpCAM as a CTSL inhibitor, confirm the functional relevance of multiple cancer-associated EpCAM mutations, and suggest a therapeutic vulnerability in cancers harboring EpCAM mutations.

MiR-3614-3p suppresses cell aggressiveness of human breast cancer by targeting AKT3 and HDAC1 expression

Purpose: MicroRNAs (miRNA) have been reported in the regulation of various pathobiological progression in cancer. Our recent study has reported that miR-3614-3p significantly suppressed the proliferation of Breast Cancer (BC) cells through the downregulation its host gene TRIM25. However, the other functional role of miR3614-3p migration and invasion in BC and its mechanism have not been investigated thoroughly. Materials and methods: The MDA-MB-231 and MCF-7 BC cell lines were purchased. The cell line expression levels of miR-3614- 3p and AKT3/HDAC1 were determined by quantitative real-time PCR (qPCR). The wound healing assay and transwell migration assay were determined. We next measured protein levels of AKT3/HDAC1 by Western blot. Finally, we investigated the role of AKT3/HDAC1 using siRNA; and confirmed the targeting of 3’UTR of AKT3 and HDAC1 through miR-3614-3p using a luciferase reporter assay. Results: In the present research, we studied that overexpression of miR-3614-3p markedly suppressed tumor cell invasion and migration independent TRIM25, whereas through regulated another targets AKT3 and HDAC1 expression. Notably, TRIM25 is also a target gene of miR-3614 which bind to pri-miR-3614 caused TRIM25 silence. Conclusion: miR-3614-3p is an anti-oncogene that can suppress breast cancer cell aggressiveness by targeting AKT3 and HDAC1, which reveals the potential values of miR-3614-3p for suppression of metastasis of BC. Keywords: Breast cancer; miR-3614; AKT3; HDAC1.

The tumor suppressive role of TIMP3 in the human osteosarcoma cells

BackgroundTissue inhibitor of metalloproteinase 3 (TIMP3) regulates a variety of cellular activities such as proliferation, viability, apoptosis, and motility. Functional loss of TIMP3 is reported in several human cancers. However, its role in osteosarcoma (OS) remains largely unclear. MethodsIn this study, we explored the mechanism underlying the modulation of TIMP3 in the growth and aggressiveness of U2OS and 143B human OS cells at both cellular and molecular levels. ResultsOur results show that overexpression of TIMP3 inhibits endogenous MMP activity and represses a series of oncogenic phenotypes of tumor cells independent of MMP inhibition, including reduced proliferation and survival, induced apoptosis, as well as improved sensitivity of tumor cells in response to cisplatin chemotherapy. TIMP3 overexpression also suppresses tumor cell invasion via its MMP inhibitory capacity. Importantly, TIMP3 modulates tumor cell oncogenesis via its induction of PTEN and subsequent inactivation of the PI3K/AKT pathway. ConclusionOur results suggest that TIMP3 is an oncosuppressor in human OS cells. Reactivation of TIMP3 function may be considered as a potential therapy for the treatment of this bone cancer.

Anti-Tumor Drug Discovery Based on Natural Product β-Elemene: Anti-Tumor Mechanisms and Structural Modification.

Natural products are important sources for drug discovery, especially anti-tumor drugs. β-Elemene, the prominent active ingredient extract from the rhizome of Curcuma wenyujin, is a representative natural product with broad anti-tumor activities. The main molecular mechanism of β-elemene is to inhibit tumor growth and proliferation, induce apoptosis, inhibit tumor cell invasion and metastasis, enhance the sensitivity of chemoradiotherapy, regulate the immune system, and reverse multidrug resistance (MDR). Elemene oral emulsion and elemene injection were approved by the China Food and Drug Administration (CFDA) for the treatment of various cancers and bone metastasis in 1994. However, the lipophilicity and low bioavailability limit its application. To discover better β-elemene-derived anti-tumor drugs with satisfying drug-like properties, researchers have modified its structure under the premise of not damaging the basic scaffold structure. In this review, we comprehensively discuss and summarize the potential anti-tumor mechanisms and the progress of structural modifications of β-elemene.

Abstract PO051: Fibroblast lineage tracing in metastatic and pre-metastatic microenvironments

Abstract Metastatic breast cancer is one of the leading causes of cancer-related mortality among women. However, the complex process of tumor cell dissemination and colonization of distant tissues is incompletely understood and largely incurable using existing therapies. Metastatic disease is not solely driven by tumor cell-intrinsic properties but is instead a consequence of dynamic interactions between cancer cells and other cell types in the tumor microenvironment (TME) including cancer-associated fibroblasts (CAFs). However, CAFs are known to display paradoxical functions across different cancer types, suggesting that heterogeneous CAF subpopulations play diverse or opposing roles in the TME. It was recently shown that periostin, a TGFβ-induced matricellular protein important for collagen cross-linking and extracellular matrix stiffening, can distinguish CAF subtypes in breast cancer. This specific CAF subpopulation is not well characterized, and their functional properties and contributions to disease progression have not been well-studied in vivo. Here, we have used in vivo genetic labelling of periostin+ subpopulations (Postn-Cre:ZSGreenl/s/l) and their progeny to assess their expansion and function during mammary tumor growth and metastasis. We find that ZSGreen-labelled periostin-expressing CAFs arise from mesenchymal-lineage cells and are more abundant in primary tumors, metastatic, and pre-metastatic sites of high-metastatic tumors versus their low-metastatic counterparts. We also find differential collagen organization in high- versus low-metastatic tumors: high-metastatic tumors display longer, aligned collagen fibers whereas low-metastatic tumors display shorter, disorganized fibers. Knocking down periostin in primary human breast CAFs inhibits tumor cell invasion through collagen in a CAF/tumor cell co-culture spheroid assay, indicating that periostin-expressing CAFs may remodel the extracellular matrix via collagen cross-linking to facilitate cell invasion. Finally, we show that periostin/ZSGreen+ cells in highly metastatic tumors are larger and more spindle-shaped, and that periostin knockdown in primary human breast CAFs reduces their size and ability to migrate, suggesting that periostin expression associates with CAF biomechanical properties such as spreading and motility. Our results indicate that highly-metastatic cancer cells mobilize periostin-expressing CAFs in the TME, which may promote collagen remodeling and collective migration of CAFs and cancer cells during tissue invasion. Citation Format: Jamie L. Null, Dae Joong Kim, Yu Zhang, James V. McCann, Thomas H. Barker, Andrew C. Dudley. Fibroblast lineage tracing in metastatic and pre-metastatic microenvironments [abstract]. In: Proceedings of the AACR Virtual Special Conference on the Evolving Tumor Microenvironment in Cancer Progression: Mechanisms and Emerging Therapeutic Opportunities; in association with the Tumor Microenvironment (TME) Working Group; 2021 Jan 11-12. Philadelphia (PA): AACR; Cancer Res 2021;81(5 Suppl):Abstract nr PO051.

Targeting galectin-3 with a high-affinity antibody for inhibition of high-grade serous ovarian cancer and other MUC16/CA-125-expressing malignancies

The lectin, galectin-3 (Gal3), has been implicated in a variety of inflammatory and oncogenic processes, including tumor growth, invasion, and metastasis. The interactions of Gal3 and MUC16 represent a potential targetable pathway for the treatment of MUC16-expressing malignancies. We found that the silencing of Gal3 in MUC16-expressing breast and ovarian cancer cells in vitro inhibited tumor cell invasion and led to attenuated tumor growth in murine models. We therefore developed an inhibitory murine monoclonal anti–Gal3 carbohydrate-binding domain antibody, 14D11, which bound human and mouse Gal3 but did not bind human Galectins-1, -7, -8 or -9. Competition studies and a docking model suggest that the 14D11 antibody competes with lactose for the carbohydrate binding pocket of Gal3. In MUC16-expressing cancer cells, 14D11 treatment blocked AKT and ERK1/2 phosphorylation, and led to inhibition of cancer cell Matrigel invasion. Finally, in experimental animal tumor models, 14D11 treatment led to prolongation of overall survival in animals bearing flank tumors, and retarded lung specific metastatic growth by MUC16 expressing breast cancer cells. Our results provide evidence that antibody based Gal3 blockade may be a viable therapeutic strategy in patients with MUC16-expressing tumors, supporting further development of human blocking antibodies against Gal3 as potential cancer therapeutics.