No abstract

Amygdalin, D-mandelonitrile-β-D-glucoside-6-β-glucoside, belongs to aromatic cyanogenic glycoside group derived from rosaceous plant seed. Mounting evidence has supported the anti-cancer effects of amygdalin. However, whether amygdalin indeed acts as an anti-tumor agent against breast cancer cells is not clear. The present study aimed to investigate the effect of amygdalin on the proliferation of human breast cancer cells. Here, we show that amygdalin exerted cytotoxic activities on estrogen receptors (ER)-positive MCF7 cells, and MDA-MB-231 and Hs578T triple-negative breast cancer (TNBC) cells. Amygdalin induced apoptosis of Hs578T TNBC cells. Amygdalin downregulated B-cell lymphoma 2 (Bcl-2), upregulated Bcl-2-associated X protein (Bax), activated of caspase-3 and cleaved poly ADP-ribose polymerase (PARP). Amygdalin activated a pro-apoptotic signaling molecule p38 mitogen-activated protein kinases (p38 MAPK) in Hs578T cells. Treatment of amygdalin significantly inhibited the adhesion of Hs578T cells, in which integrin α5 may be involved. Taken together, this study demonstrates that amygdalin induces apoptosis and inhibits adhesion of breast cancer cells. The results suggest a potential application of amygdalin as a chemopreventive agent to prevent or alleviate progression of breast cancer, especially TNBC.

Cadmium (Cd(2+)) is a major environmental pollutant that induces cytotoxicity by heavy-metal accumulation. Prolonged Cd(2+) exposure leads to cell damage by oxidative stress mainly in the kidneys, a critical organ for detoxification. To identify reliable on invasive protein biomarkers for Cd(2+)-induced nephrotoxicity, we performed 2-dimensional gel electrophoresis/matrix-assisted laser desorption/ionization time of flight mass spectra and stable isotope labeling by amino acids in cell culture/liquid chromatography-mass spectrometry analyses using conditioned media (CM) of HK-2 human kidney epithelial cells treated with CdCl2. Here, we identified heat shock cognate 71 kDa protein isoform1 (HSPA8) and α-enolase (ENO1) as potential biomarker candidates for the evaluation of Cd(2+)-induced nephrotoxicity. Treatment with CdCl2 increased the protein level of HSPA8 in CM and lysates of HK-2 cells. The mRNA level of HSPA8 was also increased by CdCl2 treatment, indicating transcriptional regulation. The level of ENO1 was increased in CM, but not in lysates of CdCl2-treated HK-2 cells. CdCl2 did not affect the mRNA level of ENO1. We provide evidence that the increases of HSPA8 and ENO1 in CM were due to Cd(2+)-induced cell death through oxidative stress. The increases of HSPA8 and ENO1 levels were also detected in CM of HK-2 cells treated with other nephrotoxic agents, such as HgCl2, NaAsO2, cisplatin, amphotericin B, and cyclosporine A. Urine and kidney tissues of CdCl2-treated rats showed increased levels of HSPA8. Taken together, this study identified HSPA8 and ENO1 as noninvasive biomarker candidates by 2 comparative proteomic analyses. These new biomarker candidates may have potential as alternatives to traditional biomarkers for the efficient and sensitive assessment of nephrotoxicity.

Tumor‐associated macrophages (TAMs) are major components of tumor microenvironment that promote invasion and metastasis of cancer cells. In this study, we investigated the effect of TAMs on phenotypic conversion of non‐neoplastic MCF10A human breast epithelial cells using an indirect co‐culture system. Co‐culture with TAMs induced epithelial‐to‐mesenchymal transition, invasive phenotype, and MMP‐9 upregulation in MCF10A cells. Comparative proteomic analysis revealed that endoplasmic reticulum oxidoreductase (ERO)1‐α was increased in MCF10A cells co‐cultured with TAMs compared to that in mono‐cultured cells. ERO1‐α was crucial for TAMs‐induced invasive phenotype and MMP‐9 upregulation involving transcription factors c‐fos and c‐Jun. Cytokine array analysis showed that levels of interleukin (IL)‐6, C‐X‐C motif ligand (CXCL)1, C‐C motif ligand (CCL)2, growth‐regulated oncogene (GRO), IL‐8, and granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) were increased in conditioned media of co‐cultured cells. Among these cytokines increased in conditioned media of co‐cultured cells, CCL2 was secreted from TAMs, leading to induction of ERO1‐α, MMP‐9 upregulation, and invasiveness in MCF10A cells. Our findings elucidated a molecular mechanism underlying the aggressive phenotypic change of non‐neoplastic breast cells by co‐culture with TAMs, providing useful information for prevention or treatment of recurrent breast cancer. Support or Funding Information The present study was supported by the National Research Foundation of Korea (No. 2013R1A2A2A04013379, No.2015M3A9B6074045, and No. 2016R1A6A1A03007648).

Tumor-associated macrophages (TAMs) are major components of tumor microenvironment that promote invasion and metastasis of cancer cells. In this study, we investigated the effect of TAMs on phenotypic conversion of non-neoplastic MCF10A human breast epithelial cells using an indirect co-culture system. Co-culture with TAMs induced epithelial-to-mesenchymal transition, invasive phenotype, and MMP-9 upregulation in MCF10A cells. Comparative proteomic analysis revealed that endoplasmic reticulum oxidoreductase (ERO)1-alpha was increased in MCF10A cells co-cultured with TAMs compared to that in mono-cultured cells. ERO1-alpha was crucial for TAMs-induced invasive phenotype and MMP-9 upregulation involving transcription factors c-fos and c-Jun. Cytokine array analysis showed that levels of interleukin (IL)-6, C-X-C motif ligand (CXCL)1, C-C motif ligand (CCL)2, growth-regulated protein (GRO), IL-8, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were increased in conditioned media of co-cultured cells. Among these cytokines increased in conditioned media of co-cultured cells, CCL2 was secreted from TAMs, leading to induction of ERO1-alpha, MMP-9 upregulation, and invasiveness in MCF10A cells. Our findings elucidated a molecular mechanism underlying the aggressive phenotypic change of non-neoplastic breast cells by co-culture with TAMs, providing useful information for prevention or treatment of recurrent breast cancer Note: This abstract was not presented at the meeting. Citation Format: Seungeun Lee, Eunhye Lee, EunYi Ko, Mina Ham, Hye Min Lee, Eun-Sook Kim, Minsoo Koh, Hyun Kyung Lim, Joohee Jung, So Yeon Park, Aree Moon. Tumor-associated macrophages secrete CCL2 and induce the invasive phenotype of human breast epithelial cells through upregulation of ERO1-alpha and MMP-9 [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 129.