Systematic proteomics analysis revealed different expression of laminin interaction proteins in breast cancer: lower in luminal subtype and higher in claudin-low subtype.

Background: Breast cancer is a major public health concern. Proteomics enables identification of proteins with aberrant properties. Here, we identified proteins with abnormal expression levels in breast cancer tissues and systematically analyzed and validated the data to locate potential diagnostic and therapeutic targets. Methods: Protein expression level in breast cancer tissues and para-carcinoma tissues were detected by Isobaric Tags for Relative and Absolute Quantification (iTRAQ) technology and further screened through Gene Expression Profiling Interactive Analysis (GEPIA) database. Cellular components, protein domain and Reactome pathway analysis were performed to screen functional targets. Abnormal expression levels of functional targets were validated by Oncomine database, quantitative real time polymerase chain reaction (qRT-PCR) and proteomics detection. Protein correlation analysis was performed to explain the abnormal expression levels of potential targets in breast cancer. Results: Overall, 207 and 207 proteins were up- and down-regulated, respectively, in breast cancer tissues, and approximately 50% were also detected in the GEPIA database. The overlapping proteins were mainly extracellular proteins containing epidermal growth factor-like domain in leukocyte adhesion molecule (EGF-Lam) domain and enriched in laminin interaction pathway. Moreover, the downregulated laminin interaction proteins could be functional targets, which were also validated through Oncomine-Richardson and Oncomine-Curtis database. However, the lower expression level of laminin interaction proteins only fit for luminal breast cancer cells with no or low metastasis ability because the proteins achieved higher expression level in more invasive claudin-low breast cancer cells. In addition, when compared with corresponding in situ carcinoma tissues, above-mentioned proteins also showed higher expression levels in invasive carcinoma tissues. Finally, we have revealed the negative correlation between the laminin interaction proteins and the claudins. Conclusions: The laminin interaction protein, especially for laminins with ß1 and γ1 subunits and their integrin receptors with α1 and α6 subunits, showed lower expression levels in luminal breast cancer with no or lower metastatic ability, but showed higher expression levels in claudin-low breast cancer with higher metastatic ability; and their higher expression could be related to the low claudin expression.

Association of ATM and BMI-1 genetic variation with breast cancer risk in Han Chinese.

We tested the hypothesis that genetic variation in ATM and BMI-1 genes can alter the risk of breast cancer through genotyping 6 variants among 524 breast cancer cases and 518 cancer-free controls of Han nationality. This was an observational, hospital-based, case-control association study. Analyses of single variant, linkage, haplotype, interaction and nomogram were performed. Risk was expressed as odds ratio (OR) and 95% confidence interval (CI). All studied variants were in the Hardy-Weinberg equilibrium and were not linked. The mutant allele frequencies of rs1890637, rs3092856 and rs1801516 in ATM gene were significantly higher in cases than in controls (P = .005, <.001 and .001, respectively). Two variants, rs1042059 and rs201024480, in BMI-1 gene were low penetrant, with no detectable significance. After adjustment, rs189037 and rs1801516 were significantly associated with breast cancer under the additive model (OR: 1.37 and 1.52, 95% CI: 1.10-1.71 and 1.14-2.04, P: .005 and .005, respectively). In haplotype analysis, haplotypes A-C-G-G (in order of rs189037, rs3092856, rs1801516 and rs373759) and A-C-A-A in ATM gene were significantly associated with 1.98-fold and 6.04-fold increased risk of breast cancer (95% CI: 1.36-2.90 and 1.65-22.08, respectively). Nomogram analysis estimated that the cumulative proportion of 3 significant variants in ATM gene was about 12.5%. Our findings collectively indicated that ATM gene was a candidate gene in susceptibility to breast cancer in Han Chinese.

TOX3 protein expression is correlated with pathological characteristics in breast cancer.

TOX3 is a newly identified gene that has been observed to correlate with breast cancer by genome-wide association studies (GWAS) in recent years. In addition, it has been noted that single-nucleotide polymorphisms (SNPs) in the TOX3 gene have a strong correlation with estrogen receptor (ER)-positive tumors. However, the role of TOX3 in breast carcinoma development is still unclear. There are limited studies on the subject of TOX3 mRNA expression in breast tumors and little information on the variation of TOX3 protein expression in relation to the clinical pathological features in breast cancer and healthy tissues. In this study, we characterize the protein expression of TOX3 in breast tumors with respect to various clinical and pathological characteristics and explore the correlation between TOX3 protein expression and ER-positive tumors. A breast cancer tissue microarray containing 267 human breast tumors and 25 healthy controls, breast cancer cell lines (ZR-75-1, MDA-MB-231, MCF-7 and Bcap-37) with positive or negative ER expression, tumor tissues and matched controls were used to analyze the protein expression levels of TOX3 by immunohistochemistry, western blot analysis and quantitative polymerase chain reaction. Among the 267 breast tumor specimens, ER expression was detected in 66 tumor tissues. The expression levels of TOX3 increased in breast carcinoma tissue compared with controls, and were higher in advanced carcinoma (T3 and T4), lymph node metastases tissues (N2) and stage III tissues. Furthermore, TOX3 protein expression was more intense in ER-positive tumors, but did not demonstrate a statistical significance. However, it was significantly increased in ER-positive breast cancer cell lines (ZR-75-1, MCF-7 and Bcap-37) compared with the MDA-MB-231 cell line, which had ER-negative expression. Our findings provide support to the hypothesis that TOX3 has a strong correlation with the development of breast cancer. The current study is likely to assist in investigating the mechanisms involved in breast cancer development.

A novel small peptide as an epidermal growth factor receptor targeting ligand for nanodelivery in vitro.

The epidermal growth factor receptor (EGFR) serves an important function in the proliferation of tumors in humans and is an effective target for the treatment of cancer. In this paper, we studied the targeting characteristics of small peptides (AEYLR, EYINQ, and PDYQQD) that were derived from three major autophosphorylation sites of the EGFR C-terminus domain in vitro. These small peptides were labeled with fluorescein isothiocyanate (FITC) and used the peptide LARLLT as a positive control, which bound to putative EGFR selected from a virtual peptide library by computer-aided design, and the independent peptide RALEL as a negative control. Analyses with flow cytometry and an internalization assay using NCI-H1299 and K562 with high EGFR and no EGFR expression, respectively, indicated that FITC-AEYLR had high EGFR targeting activity. Biotin-AEYLR that was specifically bound to human EGFR proteins demonstrated a high affinity for human non-small-cell lung tumors. We found that AEYLR peptide-conjugated, nanostructured lipid carriers enhanced specific cellular uptake in vitro during a process that was apparently mediated by tumor cells with high-expression EGFR. Analysis of the MTT assay indicated that the AEYLR peptide did not significantly stimulate or inhibit the growth activity of the cells. These findings suggest that, when mediated by EGFR, AEYLR may be a potentially safe and efficient delivery ligand for targeted chemotherapy, radiotherapy, and gene therapy.