Comparative analysis of gene regulatory networks of highly metastatic breast cancer cells established by orthotopic transplantation and intra-circulation injection

Abstract

Metastasis signature genes in breast cancer have been studied comparing transcriptomic profiles of highly metastatic cancer cell lines established by intra-circulation injection with that of their parental cell line. However, this method is not suitable to analyze the initial steps of metastasis including invasion into local tissues and the circulatory system. To characterize the molecular mechanisms of early metastasis, we established highly metastatic MDA-MB-231 cell lines that metastasized to lung by the two animal transplantation models: the orthotopic transplantation method, which mimics all steps of metastasis, or intra-circulation injection method. We then performed data-mining and network analysis of gene expression profiles of metastatic cell lines established by each transplantation method. Transcriptome analysis of seven metastatic cell lines revealed novel lung metastasis signature genes, including known metastasis promoting genes and signature genes. In the OXconc (orthotopic xenograft concentration) signature, 'chemotaxis' and 'cell adhesion' terms were enriched. In the TVIconc (tail vein injection concentration) signature, 'antigen recognition' and 'cell adhesion' were enriched. Furthermore, network analysis of the metastasis signature genes highlighted hub genes in the gene regulatory network. Our findings show that expression profiles of highly metastatic cell lines were different between the orthotopic transplantation and intra-circulation injection method. It also indicates that some metastatic signature genes have been missed in previous studies. Characterization of metastasis genes using the orthotopic transplantation method will be helpful in understanding the multi-step mechanisms of metastasis. Signature genes in OXconc may have the potential to become prognostic markers.

Establishment and characterization of highly osteolytic luminal breast cancer cell lines by intracaudal arterial injection

“…Human breast cancer cell lines MCF7‐ mSlc7a1 ‐ luc2 (IDAC, Tohoku University, Miyagi, Japan) and MDA‐MB‐231‐ mslc7a1‐luc2 (MDA‐MB‐231 parent) (Nakayama, Ito, Fujimoto, Watanabe, & Semba, ) were cultured in RPMI (Fujifilm Wako) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin G and 100 µg/ml streptomycin. CMG14‐12 cells were cultured in Dulbecco's modified Eagle medium (DMEM, Life Technology) supplemented with 10% FBS, 100 U/ml penicillin G (Meiji Seika Pharma Co., Ltd.) and 100 µg/ml streptomycin (Meiji Seika Pharma Co., Ltd.).…”

“…Human breast cancer cell lines MCF7-mSlc7a1-luc2 (IDAC, Tohoku University, Miyagi, Japan) and MDA-MB-231-mslc7a1-luc2 (MDA-MB-231 parent) (Nakayama, Ito, Fujimoto, Watanabe, & Semba, 2017) were cultured in RPMI (Fujifilm Wako) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin G and 100 µg/ml streptomycin.…”

“…Human cancer cell lines expressing luciferase were established as previously described (Nakayama et al, 2017).…”

Metastasis-Specific Gene Expression in Autochthonous and Allograft Mouse Mammary Tumor Models: Stratification and Identification of Targetable Signatures

“…Nakayama and colleagues compared gene expression of xenograft models using the human breast cancer cell line MDA-MB-231 in immune-deficient mice. They identified chemotaxis and cell adhesion gene expression enriched in pulmonary metastases generated by the orthotopic implantation model, and antigen recognition and cell adhesion in metastases from tail vein-derived metastases (40). Similarly, Pillar and colleagues have found a significant difference between the miRNA expression profiles of metastases generated by orthotopic or intravenous injection of 4T1 cells in a syngeneic model (41).…”

Generation of Rat Monoclonal Antibodies Specific for Human Stromal Cell-Derived Factor-2

“…17224; Addgene), HIV-based vectors including a cDNA expression cassette, and the pPACK packaging plasmids (System Biosciences, Palo Alto, CA) as previously described. (13) Plasmid DNAs were transfected into 293T cells by the calcium phosphate transfection method. At 32 hours post-transfection, the culture supernatants were harvested and stored as lentivirus stock.…”