For miR-335 experiments, when tumors became palpable (mean tumor volume ~ 100 mm3 ) on day 7 post-tumor inoculation, using the in vivo polyethylenimine (in vivo-jetPEI®) carrier or delivery medium, we established miR335 (miR-NC, miR-335 mimic or miR-335 inhibitor)/in vivo-jetPEI® complexes, according to manufacturer’s instruction. Treatment with the miR-335/PEI complexes consisting of 10 μg miR-335 and 1.2 μl in 100 μl of 5% glucose in vivo-jetPEI® reagent per injection, was done by intratumoral injection 3 times per day, every 72 h.

BACKGROUND: Metastasis is a leading cause of breast cancer mortality. The induction of epithelial-to-mesenchymal transition (EMT) and complex oncogenic signaling is a vital step in the evolution of highly metastatic and therapeutically-intractable breast cancer; necessitating novel target discovery or development of therapeutics that target metastatic breast cells (MBCs). METHODS: To achieve this, this study employs a combination of in silico bioinformatics analyses, protein and transcript analyses, drug sensitivity assays, functional assays and animal studies. RESULTS: The present study identified CDH11 as an inductor and/or facilitator of metastatic signaling, and biomarker of poor prognosis in MBCs. Furthermore, we showed that in the presence of CDH11-rich cancer-associated fibroblasts (CAFs), MCF7 and MDA-MB-231 MBC cell lines acquired enhanced metastatic phenotype with increased CDH11, beta-catenin, vimentin, and fibronectin (FN) expression. We also demonstrated, for the first time to the best of our knowledge that exposure to anti-CDH11 antibody suppresses metastasis, reduces CDH11, FN and beta-catenin expression, and abrogate the cancer stem cell (CSC)-like traits of MBC cells. Interestingly, ectopic expression of miR-335 suppressed CDH11, beta-catenin and vimentin expression, in concert with attenuated metastatic and CSC potentials of the MBC cells; conversely, inhibition of miR-335 resulted in increased metastatic potential. Finally, corroborating the in silica and in vitro findings, in vivo assays showed that the administration of anti-CDH11 antibody or miR-335 mimic suppressed tumorigenesis and inhibited cancer metastasis. CONCLUSIONS: These findings validate our hypotheses that miR-335 mediates anti-CDH11 antibody therapy response and that an enhanced miR-335/CDH11 ratio elicits marked suppression of the MBC CSC-like and metastatic phenotypes, thus revealing a therapeutically-exploitable inverse correlation between CDH11-enhanced CSC-like and metastatic phenotype and miR-335 expression in MBCs. Thus, we highlight the therapeutic promise of humanized anti-CDH11 antibodies or miR-335-mimic, making a case for their clinical application as efficacious therapeutic option in patients with MBC.