Targeting TRAF3IP2, Compared to Rab27, is More Effective in Suppressing the Development and Metastasis of Breast Cancer.

Here we investigated the roles of Rab27a, a player in exosome release, and TRAF3IP2, an inflammatory mediator, in development and metastasis of breast cancer (BC) in vivo. Knockdown (KD) of Rab27a (MDAKDRab27a) or TRAF3IP2 (MDAKDTRAF3IP2) in triple negative MDA-MB231 cells reduced tumor growth by 70-97% compared to wild-type tumors (MDAw). While metastasis was detected in MDAw-injected animals, none was detected in MDAKDRab27a- or MDAKDTRAF3IP2-injected animals. Interestingly, micrometastasis was detected only in the MDAKDRab27a-injected group. In addition to inhibiting tumor growth and metastasis, silencing TRAF3IP2 disrupted inter-cellular inflammatory mediator-mediated communication with mesenchymal stem cells (MSCs) injected into contralateral mammary gland, evidenced by the lack of tumor growth at MSC-injected site. Of translational significance, treatment of pre-formed MDAw-tumors with a lentiviral-TRAF3IP2-shRNA not only regressed their size, but also prevented metastasis. These results demonstrate that while silencing Rab27a and TRAF3IP2 each inhibited tumor growth and metastasis, silencing TRAF3IP2 is more effective; targeting TRAF3IP2 inhibited tumor formation, regressed preformed tumors, and prevented both macro- and micrometastasis. Silencing TRAF3IP2 also blocked interaction between tumor cells and MSCs injected into the contralateral gland, as evidenced by the lack of tumor formation on MSCs injected site. These results identify TRAF3IP2 as a novel therapeutic target in BC.

Breast Tumor Microenvironment Can Transform Naive Mesenchymal Stem Cells into Tumor-Forming Cells in Nude Mice.

How mesenchymal stem cells (MSCs) interact with tumor cells and promote tumor growth is not well understood. In this study, we demonstrate that when naive MSCs and malignant breast cancer cells (MDA-MB231) were injected into opposing mammary glands of an immunodeficient nude mouse, both cell types formed tumor-like masses within 8 weeks at the injected site. Surprisingly, MDA-MB231 cells were detected in the opposing mammary gland injected with the naive MSCs, indicating migration and crosstalk between naive MSCs and MDA-MB231 cells. Furthermore, when naive MSCs preexposed to MDA-MB231-derived conditioned medium (CM; MSCCM) or purified exosomes (Exo; MSCExo) were injected into mammary glands of nude mice, they too formed a tumor-like mass with stromal tissue within 14 weeks. Interestingly, cells dissociated from these primary explants also formed tumor-like masses. Finally, injecting MSCCM or MSCExo and naive MSCs into opposing mammary glands formed tumor-like masses on the naive MSC-injected side, suggesting migration and crosstalk between MSCCM or MSCExo with naive MSCs, similar to that observed between malignant MDA-MB231 cells and naive MSCs. Importantly, molecular analysis of MSCCM and MSCExo revealed DNA hypermethylation. These data demonstrate that MSCs and breast cancer cells communicate, resulting in the transformation of naive MSCs into cells capable of forming explants in nude mice. Our data also suggest that DNA hypermethylation might have contribute to their migration. Understanding the crosstalk between MSCs and tumor cells, and identifying the players involved in their interaction, will help us develop novel therapeutics for breast cancer regression and elimination.

Intratracheal administration of cyclooxygenase-1-transduced adipose tissue-derived stem cells ameliorates monocrotaline-induced pulmonary hypertension in rats.

The effect of intratracheal administration of cyclooxygenase-1 (COX-1)-modified adipose stem cells (ASCs) on monocrotaline-induced pulmonary hypertension (MCT-PH) was investigated in the rat. The COX-1 gene was cloned from rat intestinal cells, fused with a hemagglutanin (HA) tag, and cloned into a lentiviral vector. The COX-1 lentiviral vector was shown to enhance COX-1 protein expression and inhibit proliferation of vascular smooth muscle cells without increasing apoptosis. Human ASCs transfected with the COX-1 lentiviral vector (ASCCOX-1) display enhanced COX-1 activity while exhibiting similar differentiation potential compared with untransduced (native) ASCs. PH was induced in rats with MCT, and the rats were subsequently treated with intratracheal injection of ASCCOX-1 or untransduced ASCs. The intratracheal administration of ASCCOX-1 3 × 10(6) cells on day 14 after MCT treatment significantly attenuated MCT-induced PH when hemodynamic values were measured on day 35 after MCT treatment whereas administration of untransduced ASCs had no significant effect. These results indicate that intratracheally administered ASCCOX-1 persisted for at least 21 days in the lung and attenuate MCT-induced PH and right ventricular hypertrophy. In addition, vasodilator responses to the nitric oxide donor sodium nitroprusside were not altered by the presence of ASCCOX-1 in the lung. These data emphasize the effectiveness of ASCCOX-1 in the treatment of experimentally induced PH.

Molecular characterization of exosome-like vesicles from breast cancer cells.

BACKGROUND: Membrane vesicles released by neoplastic cells into extracellular medium contain potential of carrying arrays of oncogenic molecules including proteins and microRNAs (miRNA). Extracellular (exosome-like) vesicles play a major role in cell-to-cell communication. Thus, the characterization of proteins and miRNAs of exosome-like vesicles is imperative in clarifying intercellular signaling as well as identifying disease markers. METHODS: Exosome-like vesicles were isolated using gradient centrifugation from MCF-7 and MDA-MB 231 cultures. Proteomic profiling of vesicles using liquid chromatography-mass spectrometry (LC-MS/MS) revealed different protein profiles of exosome-like vesicles derived from MCF-7 cells (MCF-Exo) than those from MDA-MB 231 cells (MDA-Exo). RESULTS: The protein database search has identified 88 proteins in MDA-Exo and 59 proteins from MCF-Exo. Analysis showed that among all, 27 proteins were common between the two exosome-like vesicle types. Additionally, MDA-Exo contains a higher amount of matrix-metalloproteinases, which might be linked to the enhanced metastatic property of MDA-MB 231 cells. In addition, microarray analysis identified several oncogenic miRNA between the two types vesicles. CONCLUSIONS: Identification of the oncogenic factors in exosome-like vesicles is important since such vesicles could convey signals to non-malignant cells and could have an implication in tumor progression and metastasis.

Prospective dual role of mesenchymal stem cells in breast tumor microenvironment.

Breast cancer tissue is a heterogeneous cellular milieu comprising cancer and host cells. The interaction between breast malignant and non-malignant cells takes place in breast tumor microenvironment (TM), and has a crucial role in breast cancer progression. In addition to cellular component of TM, it mainly consists of cytokines released by tumor cells. The tumor-tropic capacity of mesenchymal stem cells (MSCs) and their interaction with breast TM is an active area of investigation. In the present communication, the interplay between the breast resident adipose tissue-derived MSCs (B-ASCs) and breast TM was studied. It was found that a distinct subset of B-ASCs display a strong affinity for conditioned media (CM) from two breast cancer cell lines, MDA-MB 231 (MDA-CM) and MCF-7 (MCF-CM). The expressions of several cytokines including angiogenin, GM-CSF, IL-6, GRO-α and IL-8 in MDA-CM and MCF-CM have been identified. Upon functional analysis a crucial role for GRO-α and IL-8 in B-ASCs migration was detected. The B-ASC migration was found to be via negative regulation of RECK and enhanced expression of MMPs. Furthermore, transcriptome analysis showed that migratory subpopulation express both pro- and anti-tumorigenic genes and microRNAs (miRNA). Importantly, we observed that the migratory cells exhibit similar gene and miRNA attributes as those seen in B-ASCs of breast cancer patients. These findings are novel and suggest that in breast cancer, B-ASCs migrate to the proximity of tumor foci. Characterization of the molecular mechanisms involved in the interplay between B-ASCs and breast TM will help in understanding the probable role of B-ASCs in breast cancer development, and could pave way for anticancer therapies.