Amphiregulin Expression Levels Research Articles

MiR-149 Suppresses Breast Cancer Metastasis by Blocking Paracrine Interactions with Macrophages.

Paracrine activation of cells contained in the tumor microenvironment promotes tumor progression and metastasis. In breast cancer, malignant cells recruit and educate macrophages into a M2 tumor-promoting phenotype that supports the metastatic spread of cancer cells. Here, we show that miR-149 functions as a metastasis-suppressing microRNA in breast cancer cells by limiting colony-stimulating factor-1 (CSF1)-dependent recruitment and M2 polarization of macrophages. In lymph node-positive, triple-negative breast cancer (TNBC) tissues, low miR-149 expression correlated with macrophage infiltration and reduced patient survival. By directly targeting CSF1, miR-149 expression in TNBC cell lines (MDA-MB-231 and BT-549) inhibited the recruitment of human monocytic THP-1 cells and primary human macrophages. Furthermore, in macrophages cocultured with MDA-MB-231 cells expressing miR-149, epidermal growth factor (EGF) and amphiregulin expression levels were strongly reduced, resulting in reduced EGF receptor activation in the cancer cells. In vivo, lung metastases developing from orthotopic MDA-MB-231 tumors were reduced by 75% by miR-149 expression, and this was associated with impaired M2 macrophage infiltration of the primary tumors. These data suggest that miR-149 downregulation functionally contributes to breast tumor progression by recruiting macrophages to the tumor and facilitating CSF1 and EGF receptor cross-talk between cancer cells and macrophages. SIGNIFICANCE: These findings contribute to the understanding of tumor-stroma interactions by showing that miR-149 downregulation in TNBC enhances reciprocal growth factor signaling between macrophages and cancer cells, which promotes tumor progression and metastasis. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/6/1330/F1.large.jpg.

Prognostic role of Amphiregulin and the correlation with androgen receptor in invasive breast cancer

BackgroundIn androgen-sensitive prostate cancer, androgenic stimulation induces the synthesis of amphiregulin (AREG). Research is lacking on the role of AREG in invasive breast cancer and the co-expression with androgen receptor (AR) status. Materials and methodsThe present study investigated the prognostic role of AREG in invasive breast cancer cases (N = 298) and the co-expression with the AR status as analysed by immunohistochemistry (IHC). ResultsThe samples were divided into groups according to AREG expression levels: low/no expression (AREGlow/no) and high expression (AREGhigh). As shown by cytoplasmic immunostaining, 46.0% (137/298) of invasive breast cancers were AREGhigh, and 54.0% (161/298) of cases were AREGlow/no. Co-expression of the AR and AREG accounted for 62.4% (186/298) of cases. A Kaplan–Meier analysis revealed that AREGhigh and AR+/AREGhigh decreased patients’ overall survival (OS) (P = 0.002 and P = 0.006, respectively) and disease-free survival (DFS) (P < 0.001 and P < 0.001, respectively). In Cox models, AR+/AREGhigh remained an independent prognostic indicator of OS and DFS in invasive breast cancer (hazard ratio [HR], 0.591, 95% confidence interval [CI], 0.407-0.859, P = 0.006; HR, 0.449, 95% CI, 0.236-0.853, P = 0.014, respectively). AREGhigh remained an independent prognostic indicator of OS and DFS in estrogen receptor (ER)-negative tumours (P < 0.05). ConclusionsThis study suggested that AREG and the AR were co-expressed in invasive breast cancer. Thus, AREG and the AR may be valuable prognostic biomarkers in invasive breast cancer and promising therapeutic targets, especially in ER-negative breast cancer.

Abstract 2578: Expression of epiregulin/amphiregulin associates with prognosis in recurrent/metastatic head and neck cancer treated with cetuximab

Abstract Objectives: The anti-EGFR antibody cetuximab, combined with chemotherapy, is the standard therapy for recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). However, the response rate and survival benefit are limited. The purpose of this study was to investigate the expression of the epidermal growth factor receptor ligands epiregulin (EREG) and amphiregulin (AREG), as predictive prognostic markers for the efficacy of cetuximab combined with chemotherapy in R/M HNSCC. Materials and methods: We conducted a retrospective analysis of 37 patients diagnosed with R/M HNSCC. Tumor samples from biopsy specimens or surgical specimens were analyzed for EREG and AREG mRNA gene expression, and clinical outcomes evaluated. Survival rates were estimated by Kaplan-Meier analysis. Cox proportional hazard regression was used to test the potential association between ligand expression, overall survival (OS), and progression free survival (PFS). Results: Values of EREG ≤ 15.87 and AREG ≤ 11.80, determined by receiver operating characteristic (ROC) curve analysis, were associated with significantly longer OS (p=0.03 and p=0.01, respectively), as well as PFS (p=0.04 and p=0.04, respectively). EREG and AREG expression levels were significantly correlated (r=0.73, p&amp;lt;0.01). Multivariate analysis revealed that AREG expression was significantly associated with OS and PFS. Conclusion: R/M HNSCC patients with higher EREG and AREG gene expression appear to benefit more from cetuximab combined with chemotherapy than those with lower expression. The potential for these ligands as prognostic biomarkers for cetuximab-containing regimens requires further evaluation. Citation Format: Yasunao Kogashiwa, Hitoshi Inoue, Kiyomi Kuba, Masanori Yasuda, Mitsuhiko Nakahira, Masashi Sugasawa. Expression of epiregulin/amphiregulin associates with prognosis in recurrent/metastatic head and neck cancer treated with cetuximab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2578.

Amphiregulin (AREG) expression and response to first-line panitumumab (pmab) plus FOLFIRI in metastatic colorectal cancer (mCRC).

731 Background: Biomarker analyses have shown that patients (pts) with RAS wild-type (WT) mCRC can achieve overall survival (OS) benefits with first-line pmab plus chemotherapy. Other biomarkers may exist that could optimize pt selection. Epidermal growth factor receptor ligand (eg AREG) levels have been correlated with OS during anti-EGFR therapy. Here we investigate the relationship between AREG expression and treatment outcome in a single-arm first-line mCRC study of pmab + FOLFIRI. Methods: Qualified reverse transcription quantitative polymerase chain reaction (RT-qPCR) assays were used to measure AREG RNA expression in archival formalin-fixed, paraffin embedded tumor samples from mCRC pts in two pmab trials (STEPP and 314). The STEPP analysis was used to establish a cut-off point in AREG expression that identified the best responders. This cut-off was applied prospectively to samples previously analyzed for KRAS in the 314 trial. Using the KRASMT subgroup as a non-responding comparator, Cox proportional hazards (PH) models were used to evaluate AREG expression levels as a continuous covariate. Decision curves were used to estimate the progression-free survival (PFS) hazard ratio (HR) with increasing levels of baseline AREG expression. Results: In the 314 trial 100 pts had evaluable AREG levels. Among 50 KRAS WT pts, high AREG expression was associated with objective response (OR) (Table). The high AREG group had better PFS HRs (KRAS WT/KRAS MT: 0.30 [95% CI, 0.12–0.75]) compared with the low AREG group (PFS HR 0.49 [95% CI 0.21–1.1]). There was a significant biomarker-by-AREG expression interaction in the Cox PH model (p=0.03). Conclusions: Treatment decision curves based on the PH model suggest that most KRAS WT patients express AREG at levels where treatment benefit is predicted. Future analysis of samples from a RASWT population may provide further insights. Clinical trial information: NCT00508404. [Table: see text]

Abstract 3297: PTEN loss enhances amphiregulin-specific signaling and gene expression in triple-negative breast cancer

Abstract The epidermal growth factor receptor (EGFR) is frequently overexpressed in triple-negative breast cancer, and approximately half of all basal breast cancers show autocrine activation of EGFR by amphiregulin (AREG). When AREG is the activating ligand, the dynamics of EGFR signaling are different than those of EGF-bound EGFR. AREG-ligated EGFR accumulates at the cell surface, resulting in altered receptor trafficking and downstream signaling. To further explore the role of amphiregulin in cell signaling, we performed reverse-phase protein array (RPPA) analysis on SUM149 cells, a highly tumorigenic and metastatic cell line that expresses high levels of AREG and EGFR, and MCF10A cells, a non-transformed mammary epithelial cell line cultured in the presence of either EGF or AREG. This analysis identified a number of signaling components mediated specifically by amphiregulin, including decreased caspase expression, increased Gab2 expression, elevated fibronectin levels and marked changes in integrin expression. In addition, AREG-mediated EGFR signaling resulted in increased expression and activation of c-src. Thus, a major effect of AREG signaling in both cell lines was enhanced activation of the canonical focal adhesion signaling pathway. In addition to increased expression of EGFR and AREG, SUM149 cells are known to have a genomic alteration at the PTEN locus resulting in complete abrogation of PTEN message and protein expression. RPPA analysis of signaling in SUM-149 cells confirmed this PTEN loss and, not surprisingly, revealed elevated AKT activity. In addition, several proteins in the WNT signaling pathway were altered in SUM-149 cells, including reduced expression of GSK3β, increased phosphorylation of GSK3β, and increased expression of Dvl. These results suggest enhanced Wnt-beta catenin activity in SUM-149 cells that is likely the result of enhanced AKT activity resulting from PTEN loss. Although we previously had difficulty measuring Wnt signaling in SUM-149 cells in monolayer culture, mammosphere assays performed with SUM149 cells expressing a Wnt reporter construct confirmed that Wnt signaling was highly active in mammospheres in 3D culture. We also analyzed the genes that are regulated in expression by EGFR in MCF-10A cells in the presence of EGF or AREG, and in SUM-149 cells. These results identified AREG-specific gene expression profiles in the isogenic system. In addition, SUM-149 cell-specific genes regulated by EGFR were also identified, some of which are likely to be the result of PTEN loss and enhanced AKT signaling in these cells. Finally, we found that AREG knockdown in SUM-149 cells significantly reduced the tumorigenicity of SUM149 cells in NOD-SCID mice. Analysis of signaling and gene expression in these AREG knock-down cells are consistent with the importance of AREG expression levels and of the modulation of the focal adhesion canonical pathway in tumor potential of the cells in vivo. Citation Format: Christiana Kappler, Robert Wilson, Bridget Varughese, Stephen P. Ethier. PTEN loss enhances amphiregulin-specific signaling and gene expression in triple-negative breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3297. doi:10.1158/1538-7445.AM2014-3297

Expression of Amphiregulin and EGFRvIII Affect Outcome of Patients with Squamous Cell Carcinoma of the Head and Neck Receiving Cetuximab–Docetaxel Treatment

Constitutive activation of epidermal growth factor receptor (EGFR) as a result of gene amplification, mutation, or overexpression of its ligands has been associated with response to EGFR targeting strategies. The role of these molecular mechanisms for the responsiveness of squamous cell carcinoma of the head and neck (SCCHN) to cetuximab-containing regimens remains unknown. Tumor biopsies from 47 patients, enrolled in a single-arm phase II multicenter study for second-line treatment of recurrent or metastatic SCCHN with cetuximab and docetaxel, were analyzed by immunohistochemistry for expression of EGFR, its deletion variant III (EGFRvIII) and its ligand amphiregulin (AREG). The relation between expression levels and disease control rate (DCR) was evaluated by logistic regression. Association between expression levels, progression-free survival (PFS), and overall survival (OS) was determined by Kaplan-Meier analysis, log-rank test, and uni- and multivariate Cox regression analysis. High expression of EGFR, EGFRvIII, and AREG was detected in 73%, 17%, and 45% of SCCHN cases, respectively. Expression levels of EGFR had no impact on PFS or OS. High expression levels of EGFRvIII were significantly associated with reduced DCR and shortened PFS (HR: 3.3, P = 0.005) but not with OS. Patients with high AREG expression in tumor cells had significantly shortened OS (HR: 2.2, P = 0.002) and PFS (HR 2.2, P = 0.019) compared with patients with low expression score. Multivariate Cox analysis revealed an independent association of AREG and EGFRvIII with PFS but only AREG was an independent prognosticator of OS. High EGFRvIII and AREG expression levels identify SCCHN patients who are less likely to benefit from combination treatment with cetuximab and docetaxel.

Amphiregulin Is a Promising Prognostic Marker for Liver Metastases of Colorectal Cancer

Aberrant activation of epidermal growth factor receptors (EGFR/HER1) by ligand stimulation or heterodimerization with human epidermal growth factor 2 (HER2) is considered to play an important role in the development of colorectal carcinoma. Amphiregulin (AR) is a ligand of EGFR that might be related to the development and progression of gastrointestinal tumors. The aim of this study was to determine the AR, EGFR, and HER2 protein expression levels and to evaluate their prognostic relevance to the clinical course of colorectal cancer. The AR, EGFR, and HER2 protein levels in primary tumors of colorectal cancer (n = 106) were examined using immunohistochemistry. Metastatic sites in liver specimens (n = 16) were also analyzed in the same manner. Thirteen (81.6%) metastatic lesions of the liver stained positive for AR. Among the primary lesions of colorectal cancer, 58 (54.7%) stained positive for AR, 13 (12.3%) stained positive for EGFR, and 5 (4.7%) stained positive for HER2. When the relationships between each protein expression level and the clinicopathologic factors were examined, only the AR expression level was significantly related to liver metastasis (P = 0.0296). A multivariate analysis of liver metastasis proved that AR expression was an independent prognostic factor of liver metastasis from colorectal cancer (P = 0.0217). AR expression in primary lesions of colorectal cancer is an important predictive marker of liver metastasis.