Additional file 1 of ADAMTS18 deficiency associates extracellular matrix dysfunction with a higher risk of HER2-positive mammary tumorigenesis and metastasis
Creators: name: Nie, Jiahui
name: Dang, Suying
name: Zhu, Rui
name: Lu, Tiantian
name: Zhang, Wei
Publisher: name: figshare
Year: 2024
Subjects: subject: Medicine
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Biological Sciences not elsewhere classified
subject: Developmental Biology
subject: Cancer
subject: Plant Biology
subject: Computational Biology

description: Additional file 1. Supplemental information file. Figure S1. Generation and characterization of Her2t/w/Adamts18+/+ mice and Her2t/w/Adamts18−/− mice with C57BL/6-FVB mixed background. (A) The breeding strategy of Her2t/w transgenic mouse model in the presence or absence of ADAMTS18 was described in method. Her2t/w/Adamts18+/+ mice (n = 36); Her2t/w/Adamts18−/− mice (n = 32). (B) Monitoring of mouse body weight during the period of observation. (C) The survival rate of mice until 30-month-old. P values were determined through Log-rank (Mantel-Cox) test. Figure S2. Average tumor numbers in tumor-bearing Her2t/w/Adamts18+/+ mice and Her2t/w/Adamts18−/− mice. Each dot or square represents an individual. Figure S3. Representative images and HE staining of metastatic tumors in the peritoneal cavity, liver, and kidney of Her2t/w/Adamts18−/− mice. Scale bars, 5 mm (left panel) and 100 μm (right panel). Figure S4. ADAMTS18 deficiency induces early proliferation of mammary epithelial cells (A) Representative immunohistochemical staining of cross-sections of mammary glands from the indicated genotypes using Ki-67 antibody. The red arrows are Ki-67 positive signals that were detected in Her2t/w/Adamts18−/− mice at 10 months of age. Scale bars = 100 μm. (B) Quantification of Ki-67-positive cells (%) (n = 4/group). Data are expressed as mean ± SD. **p &lt; 0.01; Student’s t-test, two tailed. Figure S5. Enhanced ERK1/2 and PI3K/AKT signaling activity and epithelial-mesenchymal transitions (EMT) process in mammary glands of 30-month-old Her2t/w/Adamts18−/− mice. (A) Western blot analysis of the protein levels of total ERK1/2 (t-ERK1/2), phosphorylated ERK1/2 (p-ERK1/2), AKT, phosphorylated AKT, E-cadherin, Ncadherin, and fibronectin in mammary glands of 30-month-old virgin Her2t/w/Adamts18+/+ and Her2t/w/Adamts18−/− mice. (B-F) Relative expression levels of the proteins are represented as p-ERK1/2/t-ERK1/2, p-AKT/AKT or protein/GAPDH. Each dot or square represents an individual. Data are expressed as means ± SD (n = 4). *p &lt; 0.05; **p &lt; 0.01; ***p &lt; 0.001; Student’s t-test, two tailed. Figure S6. Low ADAMTS18 expression in distant metastases of HER2+ tumors relapsed posttrastuzumab treatment. (A) HER2-positive breast cancer-associated GEO dataset (GSE191230). In this dataset, RNA sequencing (RNA-seq) analysis was performed on 13 treatment-naïve HER2+ breast tumors (n = 13) and 7 distant metastases of HER2+ tumors relapsed posttrastuzumab treatment (n = 7), including 2 lung metastases and 5 brain metastases. (B) Analysis of ADAMTS18 expression. RNA expression was quantified with transcripts per kilobase of exon model per million mapped reads (TPM). Data are expressed as mean ± SD. *p &lt; 0.05; two-tailed Student’s ttest. Table S1. Antibodies used in this study. Table S2. Total tumor burden at endpoint. Table S3. Determination of mRNA levels of main ECM molecules in BM and stroma as well as the mammary epithelial cell receptors by qRT-PCR. Table S4. The sample information of HER2-positive breast cancer patients.