MACC1 Overexpression Research Articles

Effects of curcumin on 5-fluorouracil resistance of colon cancer cells through the PI3K/AKT/mTOR pathway via MACC1

: Colon cancer (CC) represents a common malignancy of the digestive tract featured by high metastasis and recurrence rates. Despite the critical role of 5-fluorouracil (5-Fu) in tumor therapy, long-term and heavy use of 5-Fu leads to reduced treatment efficacy in CC patients. Curcumin can avert multidrug resistance as a sensitizing agent. This study sought to investigate the mechanism of curcumin in 5-Fu resistance of CC cells. : Firstly, the LoVo/5-Fu CC cell line was constructed using gradient 5-Fu, and subjected to treatment with curcumin or oe-MACC1 and pathway inhibitor BEZ235. Cell proliferation was assessed by MTT and clone formation assays, and IC50 values were calculated. Cell apoptosis was evaluated by means of Annexin V-FITC/PI double staining. Levels of drug-resistance factors and Vimentin and E-cadherin were measured using RT-qPCR. Levels of PI3K/AKT/mTOR pathway-associated proteins were determined with a Western blot assay. : LoVo/5-Fu cells presented with increased IC50 value, enhanced proliferation, weakened apoptosis, augmented levels of drug-resistance factors and Vimentin, and diminished E-cadherin levels. MACC1 was highly-expressed and the PI3K/AKT/mTOR-pathway-associated proteins phosphorylated in LoVo/5-Fu cells. Curcumin partly averted 5-Fu resistance and decreased the ratios of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR. MACC1 over-expression mitigated the action of curcumin on inhibiting 5-Fu resistance and elevating phosphorylation levels of PI3K/AKT/mTOR proteins. BEZ235 treatment abolished the action of over-expression of MACC1 on 5-Fu resistance of CC cells. : Our findings highlighted that curcumin suppressed the epithelial-mesenchymal transition of CC cells and thus reduced 5-Fu resistance of CC cells by regulating the PI3K/AKT/mTOR pathway via MACC1.

MACC1 Promotes the Progression and Is a Novel Biomarker for Predicting Immunotherapy Response in Colorectal Cancer.

Aims As one of the most prevalent malignant diseases in the world, the mechanisms of metastasis in colon cancer are poorly understood. The aim of this study was to investigate the role of the HGF/c-MET axis in the proliferation and metastasis in colon cancer. Methods The effect of MACC1 on cell proliferation and metastasis was analyzed through a series of in vitro experiments. The role of MACC1 in cancer cells was demonstrated by overexpression and silencing of MACC1 in gain or loss function experiments. To investigate the relationship between MACC1 and c-MET/HGF, we detected c-MET protein expression by disrupting with or overexpressing MACC1. The bioinformatics analysis was used to investigate the correlation between MACC1 and c-MET, and the c-MET expression after the interference of HGF with MACC1 was determined. Subsequently, the function of c-MET was verified in colon cancer cells by a series of experiments. The mouse tumor transplantation model experiment is most suitable in vivo. Results The results indicated that the overexpression of MACC1 could accelerate proliferation and facilitate metastasis in colon cancer cell lines. Furthermore, c-MET was determined to be the downstream regulator of MACC1. The addition of HGF could stimulate the expression of MACC1. With further exploration, we proved that c-MET is downstream of MACC1 in colon cancer and that overexpression of c-MET in colon cancer enhances cell proliferation and migration capability. At last, MACC1 expression level negatively correlates with the infiltration levels and several immune checkpoint biomarkers. High MACC1 expression has a lower response rate with ICIs in COAD. Conclusions We found that, under the regulation of the MACC1/HGF/c-MET axis, the proliferation and metastasis of colorectal cancer are increased by MACC1, which can be a novel biomarker for predicting ICIs response in colorectal cancer. Our findings provide a new idea for the targeted treatment of colorectal cancer.

Abstract 2879: MACC1 overexpression in colorectal cancer is driven by chromosomal instability and is associated with molecular subtype

Abstract Metastasis-Associated in Colon Cancer 1 (MACC1) is a strong prognostic marker for human colorectal cancers (CRC). The gene is a key regulator of the HGF/c-Met pathway and its overexpression is associated with cancer cell proliferation, migration, invasiveness and metastasis. The contexture of MACC1 overexpression in cancers is still poorly understood. Here we used the data released by The Cancer Genome Atlas (TCGA) and in silico approaches for a deep and integrative analysis of MACC1 expression modalities in CRCs. First, we curated COAD-READ TCGA datasets and obtained full clinical, genomic, and transcriptomic annotation for a total of 259 CRC samples. In tumors, we showed high heterogeneity of MACC1 mRNA expression levels with data varying from 5.58 to 13.07 U (RNAseq log transformed data - RSEM normalized). By correlating MACC1 expression to clinical data, we showed its levels were associated with anatomic regions of the disease (p=6.35.10-5). The tumors from descending colon and rectum having higher levels than those of the ascending colon and caecum (p=7.44-06). High MACC1 was also associated with higher tumor stages (p=0.01) and nodal invasion (p=0.03). At genomic level, MACC1 mutation was not frequent in CRCs (<2%). The gene is however frequently subjected to gene copy number variation (CNV) due to chromosomal instability and polyploidy (MACC1 CNV values > normal in 155/259: 60%). Correlation analysis indicated strong positive association between MACC1 copy number variations and mRNA expression levels (p<10-16). In an independent dataset, we validate that gain of MACC1 gene copy number and associated higher expression levels correlated with metastasis dissemination and worse outcome of the patients. By analyzing MACC1 with respect of CRC subtypes, we confirmed higher MACC1 levels in MSS tumors with high chromosomal instability (CIN) compared to MSI or genomic stable tumors (GS) (p=1.16-14). Regarding association with CMS1-4 transcriptomic subtypes, the highest expression of MACC1 were observed in CMS2-(epithelial), and CMS4 (mesenchymal) as compared with CMS3 (epithelial with metabolic dysregulation) or CMS1 (MMR deficient). The subset of tumors with very low MACC1 expression were mainly those altered for MLH1 (MSI-H- MMR tumors), notably most of them have normal ploidy. Deconvolution analysis of immune contexture revealed negative correlation between MACC1 expression and immune activation. Stratification using MACC1 as prognostic marker within these tumor types will be evaluated and the gene will also be presented with respect of alterations of key cancer genes in these tumors. Overall, the present study gives insight into the contexture of MACC1 expression in CRC. We showed MACC1 overexpression is largely driven by chromosomal instability and DNA copy number gains. Its expression was dependent on the tumor molecular subtypes. Citation Format: Vincent Vuaroqueaux, Hoor Al-Hasani, Dennis Kobelt, Thomas Risch, Susen Burock, Marie-Laure Yaspo, Heinz-Herbert Fiebig, Ulrike Stein. MACC1 overexpression in colorectal cancer is driven by chromosomal instability and is associated with molecular subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2879.

Circ_0101802 functions as a sponge of miR-1236-3p to facilitate the proliferation, migration and invasion of colorectal cancer via regulating MACC1

Circular RNAs (circRNAs) have been shown to be involved in the progression of many diseases, including cancer. However, the role of circ_0101802 in the proliferation, migration and invasion of colorectal cancer (CRC) has not been studied. Our results showed that circ_0101802 was highly expressed in CRC tumor tissues and cells. Functional experiments suggested that circ_0101802 knockdown could inhibit the proliferation, migration and invasion of CRC cells in vitro and CRC tumorigenesis in vivo. In the terms of mechanism, we discovered that circ_0101802 could act as a sponge of miR-1236-3p, and miR-1236-3p could target MACC1. The rescue experiments revealed that miR-1236-3p inhibitor could reverse the inhibition effect of circ_0101802 silencing on CRC proliferation, migration and invasion, and MACC1 overexpression also could abolish the negative regulation of miR-1236-3p on CRC proliferation, migration and invasion. More important, our data confirmed that circ_0101802 sponged miR-1236-3p to positively regulate MACC1. In summary, our results revealed that circ_0101802 functioned as a tumor promoter in CRC, which could facilitate CRC proliferation, migration and invasion via regulating the miR-1236-3p/MACC1 axis.

Long Noncoding RNA FGD5-AS1 Knockdown Decrease Viability, Migration, and Invasion of Non-Small Cell Lung Cancer (NSCLC) Cells by Regulating the MicroRNA-944/MACC1 Axis.

Objective:Long noncoding RNA FGD5 antisense RNA 1 (FGD5-AS1) participates in the regulation of non-small cell lung cancer (NSCLC) progression, but the underlying mechanisms are not fully revealed. This study aimed to determine the regulatory mechanism of FGD5-AS1 on the viability, migration, and invasion of NSCLC cells.Methods:QRT-PCR was performed to measure the expression of FGD5-AS1, microRNA-944 (miR-944), and MACC1 in NSCLC. The correlation between FGD5-AS1 and clinicopathological features of NSCLC patients was analyzed. The viability of NSCLC cells were detected using MTT assay, and the migration and invasion were measured by transwell assay. Additionally, dual-luciferase reporter assay was used to demonstrate the interactions among FGD5-AS1, miR-944, and MACC1. Furthermore, exosomes were isolated from NSCLC cells and identified by transmission electron microscopy (TEM) and western blot. Then, the macrophages treated with exosomes were co-cultured with NSCLC cells to assess the effect of exosomes containing lower FGD5-AS1 level on NSCLC.Results:The expression of FGD5-AS1 and MACC1 was increased in NSCLC, but miR-944 expression was decreased. FGD5-AS1 expression had significantly correlation with TNM stage and metastasis in NSCLC patients. FGD5-AS1 knockdown decreased the viability, migration, and invasion of NSCLC cells. Additionally, FGD5-AS1 and MACC1 were both targeted by miR-944 with the complementary binding sites at 3’ UTR. In the feedback experiments, miR-944 inhibition or MACC1 overexpression reversed the reduction effect of FGD5-AS1 knockdown on the tumorigenesis of NSCLC. Moreover, silencing of FGD5-AS1 suppressed macrophages M2 polarization, and eliminated the promoting effects of exosomes mediated macrophages on NSCLC cell migration and invasion.Conclusions:FGD5-AS1 knockdown attenuated viability, migration, and invasion of NSCLC cells by regulating the miR-944/MACC1 axis, providing a new therapeutic target for NSCLC.

Restoring Treatment Response in Colorectal Cancer Cells by Targeting MACC1-Dependent ABCB1 Expression in Combination Therapy.

Treatment failure of solid cancers, represented by the development of drug resistance in the primary tumor or later outgrowth of drug resistant metastases, is the major cause of death for cancer patients. It represents an urgent clinical need for predictive biomarkers which indicate the success or failure of standard treatment regimens. Besides treatment prediction, interfering with cellular processes associated with drug resistance might improve treatment response by applying combination therapies. Metastasis-associated in colon cancer (MACC) 1 was identified in our group as a prognostic biomarker in human colorectal cancer, and has been established as key player, prognostic, and predictive biomarker for tumor progression and metastasis in a variety of solid cancers. Besides increased cell proliferation and motility, subsequently contributing to growth and metastatic spread of the primary tumor, MACC1 has also been shown to dysregulate apoptosis and is contributing to treatment resistance. Here we report the MACC1 dependent treatment resistance of colorectal cancer (CRC) cells to standard therapeutics like doxorubicin by upregulating ATP-binding cassette subfamily B member 1 (ABCB1) protein. Overexpression of MACC1 in CRC cells increased both its presence on the ABCB1 promoter and its transcriptional activity, resulting in elevated ABCB1 expression and thus treatment resistance to standard therapeutics. In contrast, depleting MACC1 increased intracellular drug concentrations, leading to better treatment response. We already identified the first MACC1 transcriptional inhibitors, such as lovastatin, by high-throughput screening of clinically approved small molecule drugs. These compounds inhibited cell motility in vitro but also restricted metastasis development in xenograft mouse models by reducing MACC1 expression. Here we report, that treating high MACC1 expressing CRC cells with a combination of statins and standard therapeutics increased the rate of cytotoxicity and resulted in higher treatment response.

RETRACTED ARTICLE: MicroRNA-338-3p suppresses ovarian cancer cells growth and metastasis: implication of Wnt/catenin beta and MEK/ERK signaling pathways

BackgroundDownregulation of microRNA-338-3p (miR-338-3p) was detected in many malignant tumors, which indicated miR-338-3p might serve as a role of antioncogene in those cancers. The present study aimed to explore the roles of miR-338-3p in the growth and metastasis of ovarian cancer cells and elaborate the underlying possible molecular mechanism.MethodsMultiply biomedical databases query and KEGG pathway enrichment assay were used to infilter possible target genes and downstream pathways regulated by miR-338-3p. Overexpression miR-338-3p lentiviral vectors were transfected into ovarian cancer OVCAR-3 and OVCAR-8 cells, cell proliferation, migration and invasion were analyzed by MTT, colony formation, transwell, Matrigel assay and xenograft mouse model. One 3′-untranslated regions (UTRs) binding target gene of miR-338-3p, MACC1 (MET transcriptional regulator MACC1), and its regulated gene MET and downstream signaling pathway activities were examined by western blot.ResultsBiomedical databases query indicated that miR-338-3p could target MACC1 gene and regulate Met, downstream Wnt/Catenin beta and MEK/ERK pathways. Rescue of miR-338-3p could inhibit the proliferation, migration and invasion of ovarian cancer cells, and suppress the growth and metastasis of xenograft tumor. Restoration of miR-338-3p could attenuate MACC1 and Met overexpression induced growth, epithelial to mesenchymal transition (EMT) and activities of Wnt/Catenin beta and MEK/ERK signaling in vitro and in vivo.ConclusionsThe present data indicated that restoration of miR-338-3p could suppress the growth and metastasis of ovarian cancer cells, which might due to the inhibition of proliferation and EMT induced by MACC1, Met and its downstream Wnt/Catenin beta and MEK/ERK signaling pathways.

Abstract 1858: Saffron restricts MACC1-dependent cell proliferation and motility of colorectal cancer cells, and alters the microbiome structure

Abstract Saffron has proven a beneficial effect as a supplement in the treatment of many conditions through its perceived anti-oxidative and anti-inflammatory properties. We previously identified the gene MACC1 and demonstrated its importance as metastasis inducer, prognostic and predictive biomarker for colorectal cancer (CRC). It is confirmed as decisive driver for tumorigenesis and metastasis for a broad range of solid cancers. Therefore, we explored the inhibitory impact of saffron extracts on MACC1-induced cancer cell growth and motility and its effects on the gut microbiome. First, we assessed cell proliferation of MACC1-high and MACC1-low expressing cells using MTT and the IncuCyte real-time cell imaging system. We employed the endogenously MACC1-low expressing colon cancer cell line SW480 and clones with ectopic MACC1 overexpression. In addition, SW620 cells with endogenous high MACC1 expression and CRISPR-Cas9 mediated MACC1 knock-out were used. Cells were treated with increasing amounts of extracts from 4 different saffron species and concentration-dependent proliferation was analyzed by MTT assay. Changes in cell cycle were assessed by FACS. In vitro migration was measured by Boyden chamber assays. Activation of caspases was analyzed by Western blot. In addition, rats treated with saffron had their gut microbiome analyzed through 16S rDNA sequencing of fecal DNA samples. Upon addition of saffron, cells with high MACC1 expression showed a growth delay compared to cells with reduced MACC1 expression. Further, MACC1-dependent migration was reduced when cells were treated with saffron extracts. We did not find activation of caspases, but we showed a cell cycle arrest of MACC1 positive cells treated with saffron extracts. Using real-time measurement of cell proliferation we identified crocin as most active compound, in our system. Furthermore, rats fed with saffron showed major changes at the phylum level of their gut microbiome. A dramatic reduction/depletion of Cyanobacteria and Proteobacteria and a decrease of Bacteroidetes/Firmicutes ratio was found within the saffron treated rats. These reductions were accompanied by enrichment in Spirochaetes, Tenericutes and Candidatus saccharribacteria phyla in these rats. This is the first identification of saffron-based compounds restricting cancer cells proliferation and motility progression via the novel target MACC1. In addition, change of microbiome composition by saffron favors short chain fatty acid synthesis. The use of saffron or some of its extracts might be of therapeutic value to CRC patients. Citation Format: Nazli Güllü, Dennis Kobelt, Hassan Brim, Lena Timm, Janice Smith, Hamid Shoraka, Akbar Soleimani, Silvia Bisti, Marjorie Gondre-Lewis, Hassan Ashktorab, Ulrike S. Stein. Saffron restricts MACC1-dependent cell proliferation and motility of colorectal cancer cells, and alters the microbiome structure [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1858.

Abstract 1607: Saffron restricts MACC1-dependent cell proliferation and motility of colorectal cancer cells, and alters the microbiome structure

Abstract Saffron has proven a beneficial effect as a supplement in the treatment of many conditions through its perceived anti-oxidative and anti-inflammatory properties. We previously identified the gene MACC1 and demonstrated its importance as metastasis inducer, prognostic and predictive biomarker for colorectal cancer (CRC). It is confirmed as decisive driver for tumorigenesis and metastasis for a broad range of solid cancers. Therefore, we explored the inhibitory impact of saffron extracts on MACC1-induced cancer cell growth and motility and its effects on the gut microbiome. First, we assessed cell proliferation of MACC1-high and MACC1-low expressing cells using MTT and the IncuCyte real-time cell imaging system. We employed the endogenously MACC1-low expressing colon cancer cell line SW480 and clones with ectopic MACC1 overexpression. In addition, SW620 cells with endogenous high MACC1 expression and CRISPR-Cas9 mediated MACC1 knock-out were used. Cells were treated with increasing amounts of extracts from 4 different saffron species and concentration-dependent proliferation was analyzed by MTT assay. Changes in cell cycle were assessed by FACS. In vitro migration was measured by Boyden chamber assays. Activation of caspases was analyzed by Western blot. In addition, rats treated with saffron had their gut microbiome analyzed through 16S rDNA sequencing of fecal DNA samples. Upon addition of saffron, cells with high MACC1 expression showed a growth delay compared to cells with reduced MACC1 expression. Further, MACC1-dependent migration was reduced when cells were treated with saffron extracts. We did not find activation of caspases, but we showed a cell cycle arrest of MACC1 positive cells treated with saffron extracts. Using real-time measurement of cell proliferation we identified crocin as most active compound, in our system. Furthermore, rats fed with saffron showed major changes at the phylum level of their gut microbiome. A dramatic reduction/depletion of Cyanobacteria and Proteobacteria and a decrease of Bacteroidetes/Firmicutes ratio was found within the saffron treated rats. These reductions were accompanied by enrichment in Spirochaetes, Tenericutes and Candidatus saccharribacteria phyla in these rats. This is the first identification of saffron-based compounds restricting cancer cells proliferation and motility progression via the novel target MACC1. In addition, change of microbiome composition by saffron favors short chain fatty acid synthesis. The use of saffron or some of its extracts might be of therapeutic value to CRC patients. Citation Format: Nazli Güllü, Hassan Brim, Marjorie Gondre-Lewis, Silvia Bisti, Hamid Shoraka, Dennis Kobelt, Akbar Soleimani, Lena Timm, Janice Smith, Hassan Ashktorab, Ulrike Stein. Saffron restricts MACC1-dependent cell proliferation and motility of colorectal cancer cells, and alters the microbiome structure [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1607.

MACC1 overexpression in carcinoma‑associated fibroblasts induces the invasion of lung adenocarcinoma cells via paracrine signaling.

Carcinoma‑associated fibroblasts (CAFs) are essential for initiating lung cancer cell invasion and metastasis. An elevated MACC1 expression has been implicated in the progression of lung adenocarcinoma. Hitherto, the role of MACC1 in lung adenocarcinoma‑derived CAFs remains unclear. In this study, CAFs isolated from the tissues of patients with lung adenocarcinoma expressed typical CAF markers (shown by immunohistochemical and immunofluorescence analysis) and exhibited enhanced migration and invasion abilities when co‑cultured with A549 cells in a microfluidic model. MACC1‑overexpressing CAFs not only demonstrated an increased invasion, but also exerted a promoting effect on the invasion of tumor cells. The reduced expression of MACC1 impaired the invasive ability of the CAFs. Western blot analysis and RT‑qPCR analysis demonstrated that multiple paracrine pathways were activated in the MACC1‑overexpressing CAFs. Overall, this study presents a novel role of MACC1 in CAF‑induced lung adenocarcinoma cell invasion, which possibly occurs via paracrine signaling. Furthermore, MACC1 was indicated to be a potential therapeutic target for lung adenocarcinoma.

Evaluation of the correlation of MACC1, CD44, Twist1, and KiSS-1 in the metastasis and prognosis for colon carcinoma

BackgroundMetastasis-associated in colon cancer 1 (MACC1) has been reported to promote tumor cell invasion and metastasis. Cancer stem cells and epithelial-mesenchymal transition (EMT) have also been reported to promote tumor cell proliferation, invasion, and metastasis. KiSS-1, a known suppressor of metastasis, has been reported to be down-regulated in various tumors. However, the associations of MACC1, CD44, Twist1, and KiSS-1 in colonic adenocarcinoma (CAC) invasion and metastasis remain unclear. The purpose of this study is to investigate the roles of MACC1, CD44, Twist1, and KiSS-1 in CAC invasion and metastasis and their associations with each other and with the clinicopathological characteristics of CAC patients.MethodsImmunohistochemistry and multivariate analysis were carried out to explore the expression of MACC1, CD44, Twist1, and KiSS-1 in 212 whole-CAC-tissue specimens and the corresponding normal colon mucosa tissues. Demographic, clinicopathological, and follow-up data were also collected.ResultsThe results of this study showed MACC1, CD44, and Twist1 expression to be up-regulated, and KiSS-1 expression was down-regulated in CAC tissues. Positive expression of MACC1, CD44, and Twist1 was found to be positively correlated with invasion, tumor grades, and lymph- node-metastasis (LNM) stages and tumor-node-metastasis (TNM) stages for patients with CAC. Positive expression of KiSS-1 was inversely associated with invasion, tumor size, LNM stage, and TNM stage. The KiSS-1-positive expression group had significantly more favorable OS than did the KiSS-1-negative group. Univariate analysis indicated that overexpression of MACC1, CD44, and Twists1 was negatively associated with longer overall survival (OS) time, and there was a positive relationship between KiSS-1-positive expression and OS time for patients with CAC. Multivariate Cox analysis demonstrated that overexpression of MACC1, CD44, Twist1, and low expression of KiSS-1 and LNM and TNM stages were independent predictors of prognosis in patients with CAC.ConclusionsThe results in this study indicated that levels of expression of MACC1, CD44, Twist1, and KiSS-1 are related to the duration of OS in patients with CAC. MACC1, CD44, Twist1, and KiSS-1 may be suitable for use as biomarkers and therapeutic targets in CAC.

Elevated MACC1 expression predicts poor prognosis in small invasive lung adenocarcinoma.

Patients with small (⩽ 2 cm) invasive lung adenocarcinoma are at high risk of poor prognosis and disease recurrence after complete surgical resection. Therefore, identification of high-risk individuals from these patients emerges as an urgent problem. Elevated MACC1 expression predicts a poor prognosis in multiple types of cancer that are independent of TNM staging. This study investigated the prognostic value of MACC1 expression in patients with small invasive lung adenocarcinoma. The current study aimed to evaluate the relationship between MACC1 expression in patients' tumor tissue and prognosis in small invasive lung adenocarcinoma. The records of 131 patients with small invasive lung adenocarcinoma who underwent complete surgical resection were reviewed. The MACC1 expression was detected by immunohistochemical staining in all specimens. Meanwhile, western blot and real-time quantitative PCR were used to examine the expression level of MACC1 in human lung adenocarcinoma cell lines. The effect of clinicopathological risk factors on patients' survival was analyzed using the Kaplan-Meier approach and multivariable Cox models. Elevated MACC1 expression was observed in 53 (40.5%) specimens, and in A549, H358, H460 and H322 lung adenocarcinoma cell lines. MACC1 overexpression was associated with differentiation (P= 0.005) and blood vessel invasion (P= 0.001). Compared with low MACC1 expression, elevated MACC1 expression was associated with significantly shorter overall survival (odds ratio = 6.515; 95% confidence interval: 1.382-30.721; P= 0.018) and disease-free survival (odds ratio = 3.270; 95% confidence interval: 1.117-9.569; P= 0.031). Multivariate analyses demonstrated high MACC1 expression is an independent risk factor of worse overall survival (odds ratio = 5.684; 95% confidence interval: 1.145-28.210; P= 0.034) and disease-free survival (odds ratio = 4.667; 95% confidence interval: 1.372-15.877; P= 0.014). MACC1 is an independent prognostic marker in patients with small invasive lung adenocarcinoma after complete surgical resection. Differential outcomes are associated with MACC1 expression level.

MACC1 regulates Fas mediated apoptosis through STAT1/3 – Mcl-1 signaling in solid cancers

MACC1 was identified as a novel player in cancer progression and metastasis, but its role in death receptor-mediated apoptosis is still unexplored. We show that MACC1 knockdown sensitizes cancer cells to death receptor-mediated apoptosis. For the first time, we provide evidence for STAT signaling as a MACC1 target. MACC1 knockdown drastically reduced STAT1/3 activating phosphorylation, thereby regulating the expression of its apoptosis targets Mcl-1 and Fas. STAT signaling inhibition by the JAK1/2 inhibitor ruxolitinib mimicked MACC1 knockdown-mediated molecular signatures and apoptosis sensitization to Fas activation. Despite the increased Fas expression, the reduced Mcl-1 expression was instrumental in apoptosis sensitization. This reduced Mcl-1-mediated apoptosis sensitization was Bax and Bak dependent. MACC1 knockdown also increased TRAIL-induced apoptosis. MACC1 overexpression enhanced STAT1/3 phosphorylation and increased Mcl-1 expression, which was abrogated by ruxolitinib. The central role of Mcl-1 was strengthened by the resistance of Mcl-1 overexpressing cells to apoptosis induction. The clinical relevance of Mcl-1 regulation by MACC1 was supported by their positive expression correlation in patient-derived tumors. Altogether, we reveal a novel death receptor-mediated apoptosis regulatory mechanism by MACC1 in solid cancers through modulation of the STAT1/3-Mcl-1 axis.

Retraction Note to: Evaluation of gene expression level of CDC5L and MACC1 in poor prognosis and progression of osteosarcoma.

Current evidences have indicated that osteosarcoma is strongly associated with abnormal genetic and epigenetic changes that lead to the abnormal expression of oncogenes or methylation of tumor suppressor genes. In the present study, MACC1 and CDC5L mRNA levels in the patients with osteosarcoma were evaluated using quantitative real-time PCR. Our results demonstrated that CDC5L mRNA levels were higher in tumor tissues than in adjacent normal tissues (2.713 ± 0.738 vs. 1.071 ± 0.629; P < 0.05). Moreover, MACC1 was upregulated in tumor bone tissues than in adjacent normal tissues (3.221 ± 0. 624 vs. 1.427 ± 0.456; P < 0.05). Our result demonstrated that high expression of CDC5L was significantly related to advanced TNM stage (P = 0.032). No significant difference was determined between CDC5L mRNA expression and other clinicopathological parameters including age, gender, tumor diameter, location, tumor grade, and histological type. In addition, overexpression of MACC1 was strongly correlated with advanced TNM stage (P = 0.027) and high tumor grade (P = 0.035). Our findings indicated that mRNA level of CDC5L is correlated with advanced TNM stage, and MACC1 may be involved in progression of osteosarcoma.

MACC1 Induces Tumor Progression in Transgenic Mice and Colorectal Cancer Patients via Increased Pluripotency Markers Nanog and Oct4.

We have previously identified the gene MACC1 as a strong prognostic biomarker for colorectal cancer metastasis and patient survival. Here, we report for the first time the generation of transgenic mouse models for MACC1. We generated mice with transgenic overexpression of MACC1 in the intestine driven by the villin promoter (vil-MACC1) and crossed them with Apc(Min) mice (vil-MACC1/Apc(Min)). vil-MACC1/Apc(Min) mice significantly increased the total number of tumors (P = 0.0056). This was particularly apparent in large tumors (≥3-mm diameter; P = 0.0024). A detailed histopathologic analysis of these lesions demonstrated that the tumors from the vil-MACC1/Apc(Min) mice had a more invasive phenotype and, consequently, showed a significantly reduced survival time than Apc(Min) mice (P = 0.03). Molecular analysis revealed an increased Wnt and pluripotency signaling in the tumors of vil-MACC1/Apc(Min) mice. Specifically, we observed a prominent upregulation of the pluripotency markers Oct4 and Nanog in these tumors compared with Apc(Min) controls. Finally, we could also validate that Oct4 and Nanog are regulated by MACC1 in vitro and strongly correlate with MACC1 levels in a cohort of 60 tumors of colorectal cancer patients (r = 0.7005 and r = 0.6808, respectively; P > 0.0001 and P > 0.0002, respectively). We provide proof of principle that MACC1-induced tumor progression in colorectal cancer acts, at least in part, via the newly discovered MACC1/Nanog/Oct4 axis. These findings might have important implications for the design of novel therapeutic intervention strategies to restrict tumor progression. Clin Cancer Res; 22(11); 2812-24. ©2016 AACR.

Evaluation of gene expression level of CDC5L and MACC1 in poor prognosis and progression of osteosarcoma.

Current evidences have indicated that osteosarcoma is strongly associated with abnormal genetic and epigenetic changes that lead to the abnormal expression of oncogenes or methylation of tumor suppressor genes. In the present study, MACC1 and CDC5L mRNA levels in the patients with osteosarcoma were evaluated using quantitative real-time PCR. Our results demonstrated that CDC5L mRNA levels were higher in tumor tissues than in adjacent normal tissues (2.713 ± 0.738 vs. 1.071 ± 0.629; P < 0.05). Moreover, MACC1 was upregulated in tumor bone tissues than in adjacent normal tissues (3.221 ± 0. 624 vs. 1.427 ± 0.456; P < 0.05). Our result demonstrated that high expression of CDC5L was significantly related to advanced TNM stage (P = 0.032). No significant difference was determined between CDC5L mRNA expression and other clinicopathological parameters including age, gender, tumor diameter, location, tumor grade, and histological type. In addition, overexpression of MACC1 was strongly correlated with advanced TNM stage (P = 0.027) and high tumor grade (P = 0.035). Our findings indicated that mRNA level of CDC5L is correlated with advanced TNM stage, and MACC1 may be involved in progression of osteosarcoma.

Influence of the MACC1 gene on sensitivity to chemotherapy in human U251 glioblastoma cells.

This study was conducted to determine the influence of MACC1 expression on chemotherapy sensitivity in human U251 glioblastoma cells. Expression of the MACC1 gene in 49 cases of human brain glioma was determined by quantitative real-time PCR. Silencing effects of RNA interference on MACC1 was detected by Western-blotting. Flow cytometry methods and methyl thiazolyl tetrazolium assay (MTT) were used to determine the apoptosis and growth inhibitory rates of the U251 cells with MACC1 silencing. before and after treatment with cisplatin (DDP). MACC1 mRNA in gliomas was up-regulated remarkably, to 158.8% of that in peri-cancerous tissues (P<0.05). The siRNA-MACC1 could inhibit the expression of MACC1 protein significantly (p<0.05), associated with an increase in apoptosis rate from 2.57% to 5.39% in U251 cells and elevation of the growth inhibitory rate from 1.5% to 17.8% (p<0.05 for both). After treatment with DDP at various concentrations (1, 3, 5μg/ml), compared with control U251 cells, the apoptosis rate of MACC1-silenced U251 cells rose from 8.41%, 13.2% and 19.5% to 12.8%, 17.8% and 25.8%; the growth inhibitory rate increased from 16.2%, 19.3% and 24.5% to 23.7%, 28.4% and 36.3%. There is a notable relationship between over-expression of MACC1 and the characteristics of glioma cells. Silencing of MACC1 was found to enhance the apoptosis and growth inhibitory rates of U251 glioma cells, and thereby increase their sensitivity to DDP chemotherapy.

MACC1 Suppresses Cell Apoptosis in Hepatocellular Carcinoma by Targeting the HGF/c-MET/AKT Pathway

Background & Aims: To investigate the expression and prognostic value of MACC1 in patients with HCC and identify the mechanism by which MACC1 inhibits HCC cell apoptosis. Methods: MACC1 and p-AKT expression was studied using immunohistochemistry of both HCC tissues and adjacent liver tissues. qRT-PCR and western immunoblotting were used to examine the expression of target genes at the mRNA and protein levels, respectively. The MTT assay was used to assess cell viability, and cell apoptosis was determined by DAPI staining, Annexin V/PI staining and Caspase 3/7 assay. Nude mice were used to perform in vivo experiments. Results: The overexpression of MACC1 was found in HCC tissues and was correlated with poor postsurgical prognosis. There was a positive relationship between MACC1 and p-AKT expression in HCC tissues. In vitro experiments showed that MACC1 repressed HCC cell apoptosis and promoted cell growth. Knockdown of c-MET abolished the anti-apoptotic function of MACC1. Next, MACC1 was verified to activate PI3K/AKT signaling by sensitizing HGF/c-MET signaling in HCC. MACC1 overexpression enhanced the HGF-driven phosphorylation of BAD, Caspase 9 and FKHRL1 and inhibited their pro-apoptotic functions in HCC cells. Finally, MACC1 was shown to inhibit cell apoptosis and promote HCC growth in vivo. Conclusions: This investigation revealed that MACC1 overexpression predicted worse prognosis after liver resection, which was attributed to the repression of HCC cell apoptosis via a molecular mechanism in which MACC1 accelerated the activation of the HGF/c-MET/PI3K/AKT pathway and phosphorylated BAD, Caspase 9 and FKHRL1, ultimately preventing their nuclear translocation and their pro-apoptotic function.

Low SOX17 expression is a prognostic factor and drives transcriptional dysregulation and esophageal cancer progression

The transcriptional network of the SRY (sex determining region Y)-box 17 (SOX17) and the prognostic impact of SOX17 protein expression in human cancers remain largely unclear. In this study, we evaluated the prognostic effect of low SOX17 protein expression and its dysregulation of transcriptional network in esophageal squamous cell carcinoma (ESCC). Low SOX17 protein expression was found in 47.4% (73 of 154) of ESCC patients with predicted poor prognosis. Re-expression of SOX17 in ESCC cells caused reduced foci formation, cell motility, decreased ESCC xenograft growth and metastasis in animals. Knockdown of SOX17 increased foci formation in ESCC and normal esophageal cells. Notably, 489 significantly differential genes involved in cell growth and motility controls were identified by expression array upon SOX17 overexpression and 47 genes contained putative SRY element in their promoters. Using quantitative chromatin immunoprecipitation-PCR and promoter activity assays, we confirmed that MACC1, MALAT1, NBN, NFAT5, CSNK1A1, FN1 and SERBP1 genes were suppressed by SOX17 via the SRY binding-mediated transcriptional regulation. Overexpression of FN1 and MACC1 abolished SOX17-mediated migration and invasion suppression. The inverse correlation between SOX17 and FN1 protein expression in ESCC clinical samples further strengthened our conclusion that FN1 is a transcriptional repression target gene of SOX17. This study provides compelling clinical evidence that low SOX17 protein expression is a prognostic biomarker and novel cell and animal data of SOX17-mediated suppression of ESCC metastasis. We establish the first transcriptional network and identify new suppressive downstream genes of SOX17 which can be potential therapeutic targets for ESCC.

Promoter identification and transcriptional regulation of the metastasis gene MACC1 in colorectal cancer

MACC1, Metastasis associated in colon cancer 1, is a newly identified prognostic biomarker for colorectal cancer metastasis and patient survival, when determined in the primary tumor or patient blood. MACC1 induces cell motility and proliferation in cell culture and metastasis in mouse models. MACC1 acts as a transcriptional regulator of the receptor tyrosine kinase gene Met via binding to its promoter. However, no information about the promoter of the MACC1 gene and its transcriptional regulation has been reported so far. Here we report the identification of the MACC1 promoter using a promoter luciferase construct that directs transcription of MACC1. To gain insights into the essential domains within this promoter region, we constructed 5′ truncated deletion constructs. Our results show that the region from −426 to −18 constitutes the core promoter and harbors functional motifs for the binding of AP-1, Sp1, and C/EBP transcription factors as validated by site directed mutagenesis study. Using electrophoretic mobility shift assay and chromatin immunoprecipitation assay, we demonstrated the physical interaction of these transcription factors to a minimal essential MACC1 core promoter sequence. Knock down of these transcription factors using RNAi strategy reduced MACC1 expression (P < 0.001), and resulted in decrease of cell migration (P < 0.01) which could be specifically rescued by ectopic overexpression of MACC1. In human colorectal tumors, expression levels of c-Jun and Sp1 correlated significantly to MACC1 (P = 0.0007 and P = 0.02, respectively). Importantly, levels of c-Jun and Sp1 also showed significant correlation to development of metachronous metastases (P = 0.01 and P = 0.001, respectively). This is the first study identifying the MACC1 promoter and its transcriptional regulation by AP-1 and Sp1. Knowledge of the transcriptional regulation of the MACC1 gene will implicate in enhanced understanding of its role in cancer progression and metastasis.