Overexpression of EZH2 in estrogen receptor negative (ER-) breast cancer promotes metastasis. EZH2 has been mainly studied as the catalytic component of the Polycomb Repressive Complex 2 (PRC2) that mediates gene repression by trimethylating histone H3 at lysine 27 (H3K27me3). However, how EZH2 drives metastasis despite the low H3K27me3 levels observed in ER- breast cancer is unknown. Here we show that in human invasive carcinomas and distant metastases, cytoplasmic EZH2 phosphorylated at T367 is significantly associated with ER- disease and low H3K27me3 levels. p38-mediated EZH2 phosphorylation at T367 promotes EZH2 cytoplasmic localization and potentiates EZH2 binding to vinculin and other cytoskeletal regulators of cell migration and invasion. Ectopic expression of a phospho-deficient T367A-EZH2 mutant is sufficient to inhibit EZH2 cytoplasmic expression, disrupt binding to cytoskeletal regulators, and reduce EZH2-mediated adhesion, migration, invasion, and development of spontaneous metastasis. These results point to a PRC2-independent non-canonical mechanism of EZH2 pro-metastatic function.

Collectively, these data demonstrate that salivary gland mucoepidermoid carcinomas contain a small population of cancer stem cells with enhanced tumorigenic potential and that are characterized by high ALDH activity and CD44 expression. These results suggest that patients with mucoepidermoid carcinoma might benefit from therapies that ablate these highly tumorigenic cells.

Cancer-stromal cell interaction is a critical process in tumorigenesis. Conventional dish-based assays, which simply mix two cell types, are limited in three aspects: 1) Limited control of cell microenvironment; 2) Inability of studying cell behavior in a single-cell manner, and; 3) Difficulties in characterizing single cell behavior within a highly heterogeneous cell population (e.g. tumor). An innovative use of microfluidic technology is centered on improving the spatial resolution for single cell assays. However, it is challenging to isolate the paired interacting cells, while maintaining nutrient renewal. In this work, two-phase flow was used as the simple isolation method, separating the microenvironments of each individual chamber. As nutrients in an isolated chamber are consumed by cells, media exchange is required. To connect cell culture chamber to the media exchange layer, we demonstrated a 3D microsystem integration technique using vertical connections fabricated by deep reactive-ion etching (DRIE). Compared to previous approaches, the presented process allows the area reduction in vertical connections by an order of magnitude, enabling compact 3D integration. A semi-permeable membrane was sandwiched between cell culture layer and media exchange layer. The selectivity of the semi-permeable membrane can retain the signaling proteins within the chamber, while allowing free diffusion of nutrients (e.g., glucose and amino acids). Thus, paracrine signals are accumulated inside the chamber without cross-talk with cells in other chambers. Utilizing these innovations, we demonstrated co-culture of UM-SCC-1 (head and neck squamous cell carcinoma) cells and endothelial cells to recapitulate tumor proliferation enhancement in the vascular endothelial niche.

The emergence of colistin or tigecycline resistance as well as imipenem resistance in Acinetobacter baumannii poses a great therapeutic challenge. The bactericidal and synergistic effects of several combinations of antimicrobial agents against imipenem-, colistin-or tigecyclineresistant A. baumannii isolates were investigated by in vitro time-kill experiments. Six imipenemresistant A. baumannii blood isolates were examined in this study, including colistin-and tigecycline-susceptible, colistin-resistant but tigecycline-susceptible, and colistin-susceptible but tigecycline-resistant isolates. Time-kill studies were performed using five antimicrobial agents singly or in combinations (imipenem plus colistin, imipenem plus ampicillin-sulbactam, colistin plus rifampicin, colistin plus tigecycline, and tigecycline plus rifampicin) at concentrations of 0.5¾ and 1¾ their MICs. Only imipenem was consistently effective as a single agent against all six A. baumannii isolates. Although the effectiveness of combinations of 0.5¾ MIC antimicrobial agents was inconsistent, combination regimens using 1¾ MIC of the antimicrobial agents displayed excellent bactericidal activities against all six A. baumannii isolates. Among the combinations of 0.5¾ MIC antimicrobial agents, the combination of colistin and tigecycline showed synergistic or bactericidal effects against four of the isolates. This in vitro time-kill analysis suggests that antimicrobial combinations are effective for killing imipenem-resistant A. baumannii isolates, even if they are simultaneously resistant to either colistin or tigecycline. However, the finding that the combinations of 0.5¾ MIC antimicrobial agents were effective on only some isolates may warrant further investigation of the doses of combination agents needed to kill resistant A. baumannii.