Hannah Schmucker1, Walker M Blanding1, Julia M Mook2, Jessica F Wade1, Jang Pyo Park3, Kerri Kwist3, Hiral Shah4, Brian W Booth5,6. 1. Department of Biological Sciences, Clemson University, Clemson, SC, USA. 2. Department of Microbiology, Clemson University, Clemson, SC, USA. 3. Institute for Biological Interfaces of Engineering, Clemson University, Clemson, SC, USA. 4. Department of Bioengineering, Institute for Biological Interfaces of Engineering, Clemson University, 401-1 Rhodes Engineering Research Center, Clemson, SC, 29634, USA. 5. Institute for Biological Interfaces of Engineering, Clemson University, Clemson, SC, USA. brbooth@clemson.edu. 6. Department of Bioengineering, Institute for Biological Interfaces of Engineering, Clemson University, 401-1 Rhodes Engineering Research Center, Clemson, SC, 29634, USA. brbooth@clemson.edu.
Abstract
PURPOSE: Tumor initiation and progression rely on cellular proliferation and migration. Many factors are involved in these processes, including growth factors. Amphiregulin (AREG) is involved in normal mammary development and the development of estrogen receptor (ER)-positive breast cancer. The aim of this project was to determine if AREG is involved in the proliferation and progression of HER2-positive breast cancer. METHODS: Mouse cell lines MMTV-neu, HC-11 and COMMA-D, as well as human cell lines MCF10A, SKBR3, HCC1954 and BT474 were used. Real-time PCR was used to quantify AREG expression and neutralizing antibodies were used to reduce the autocrine/paracrine effects of AREG. Transfections using siRNA and shRNA were used to knockdown AREG expression in the cancer cell lines. Free-floating sphere formation, colony forming, scratch wound and Transwell assays were used to assess the proliferation, tumor forming and migratory capacities of transfected cancer cells. RESULTS: We found AREG expression in both normal epithelial cell lines and tumor-derived cell lines. Knockdown of AREG protein expression resulted in reduced sphere sizes and reduced sphere numbers in both mouse and human cancer cells that overexpress erbB2/HER2. AREG was found to be involved in cancer cell migration and invasion. In addition, we found that AREG expression knockdown resulted in different migration capacities in normal and erbB2/HER2 overexpressing cancer cells. CONCLUSIONS: Based on our results we conclude that AREG is involved in regulating the proliferation and migration of erbB2/HER2-positive breast cancer cells.
PURPOSE:Tumor initiation and progression rely on cellular proliferation and migration. Many factors are involved in these processes, including growth factors. Amphiregulin (AREG) is involved in normal mammary development and the development of estrogen receptor (ER)-positive breast cancer. The aim of this project was to determine if AREG is involved in the proliferation and progression of HER2-positive breast cancer. METHODS:Mouse cell lines MMTV-neu, HC-11 and COMMA-D, as well as human cell lines MCF10A, SKBR3, HCC1954 and BT474 were used. Real-time PCR was used to quantify AREG expression and neutralizing antibodies were used to reduce the autocrine/paracrine effects of AREG. Transfections using siRNA and shRNA were used to knockdown AREG expression in the cancer cell lines. Free-floating sphere formation, colony forming, scratch wound and Transwell assays were used to assess the proliferation, tumor forming and migratory capacities of transfected cancer cells. RESULTS: We found AREG expression in both normal epithelial cell lines and tumor-derived cell lines. Knockdown of AREG protein expression resulted in reduced sphere sizes and reduced sphere numbers in both mouse and humancancer cells that overexpress erbB2/HER2. AREG was found to be involved in cancer cell migration and invasion. In addition, we found that AREG expression knockdown resulted in different migration capacities in normal and erbB2/HER2 overexpressing cancer cells. CONCLUSIONS: Based on our results we conclude that AREG is involved in regulating the proliferation and migration of erbB2/HER2-positive breast cancer cells.
Entities:
Keywords:
Amphiregulin; Breast cancer; HER2; Migration; Transfection; erbB2
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