Abstract P2-01-11: A peptide with the conserved amino acid residue of integrin α6 inhibits metastasis through disruption of complex formation in breast cancer cells

Abstract

Abstract Metastasis inhibition may improve survival of breast cancer patients. Our previous study revealed that a transcription factor SALL4 promotes cell migration for metastasis through up-regulation of integrin α6β1 in metastatic basal-like breast cancer cells. In these cells, integrin α6β1 modulates Rho GTPase activity, which enhances focal adhesion dynamics for cell migration. We therefore focused on integrin α6 as a therapeutic target for metastasis. We investigated the expression level of integrin α6 in breast cancer cell lines and a tissue microarray of breast cancer patients. In both experiments, we observed higher integrin α6 expression in basal-like breast cancer cells than in luminal ones. We performed shRNA-mediated integrin α6 knockdown in basal-like breast cancer cells and conducted Boyden chamber assays. In the results, reduced cell migration was observed in integrin α6 knocked-down cells. Reciprocally, we introduced integrin α6 overexpression in low-migratory luminal breast cancer cell lines and observed increased cell migration. These results indicate that integrin α6 promotes cell migration in breast cancer cells. To identify the functional residue of integrin α6, we analyzed its amino acid sequence by in silico analyses. First, we extracted integrin α6 specific residues in integrin α family in the human genome. Then, we compared integrin α6 sequences among vertebrates. In the result, we obtained an integrin α6-specific and vertebrate-conserved sequence, Asp-358, as a candidate for the functional residue. To inhibit integrin α6 function, we designed an 8-amino acid peptide with the sequence around Asp-358. Basal-like breast cancer cells treated with the peptide showed reduced cell migration in a dose-dependent manner. The peptide did not reduce cell migration in integrin α6 knocked-down cells, suggesting that the peptide inhibits integrin α6 function. To determine whether the peptide inhibits metastasis, we performed zebrafish metastasis assays, and observed reduced metastasis rate in the peptide-treated group. These results indicate that the peptide inhibits metastasis through reduction in cell migration. To understand the effect of the peptide, we performed chemical cross-liking assay in a basal-like breast cancer cell line. Protein samples from the cells treated with a cross-linker showed bands of integrin α6 complexes, and the intensities of these bands were reduced in the peptide-treated group. Moreover, we immunostained cells with an antibody for focal adhesion marker, phospho-paxillin, because integrins regulates focal adhesion formation for cell migration. In the results, the peptide reduced the number of focal adhesions. These observations indicate that the peptide inhibits integrin α6 function. In the future, our findings may contribute to development of a metastasis inhibition therapy. Citation Format: Itou J, Tanaka S, Senda N, Iida A, Sehara-Fujisawa A, Sato F, Toi M. A peptide with the conserved amino acid residue of integrin α6 inhibits metastasis through disruption of complex formation in breast cancer cells [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-01-11.