Immune evasion: An imperative and consequence of MYC deregulation.

MYC has been implicated in the pathogenesis of a wide range of human tumors and has been described for many years as a transcription factor that regulates genes with pleiotropic functions to promote tumorigenic growth. However, despite extensive efforts to identify specific target genes of MYC that alone could be responsible for promoting tumorigenesis, the field is yet to reach a consensus whether this is the crucial function of MYC. Recent work shifts the view on MYC's function from being a gene-specific transcription factor to an essential stress resilience factor. In highly proliferating cells, MYC preserves cell integrity by promoting DNA repair at core promoters, protecting stalled replication forks, and/or preventing transcription-replication conflicts. Furthermore, an increasing body of evidence demonstrates that MYC not only promotes tumorigenesis by driving cell-autonomous growth, but also enables tumors to evade the host's immune system. In this review, we summarize our current understanding of how MYC impairs antitumor immunity and why this function is evolutionarily hard-wired to the biology of the MYC protein family. We show why the cell-autonomous and immune evasive functions of MYC are mutually dependent and discuss ways to target MYC proteins in cancer therapy.

Mammalian Cell Fusion Assays for the Study of Cell Cycle Progression by Functional Complementation.

Cell cycle progression, or its arrest upon checkpoint activation, is directed by a complex array of cellular processes dependent on the diffusion of chemical signals. These signals regulate the onset of each cell cycle phase and prevent undesired phase transitions. Functional complementation is a robust strategy to identify such signals, by which mutant phenotypes are rescued through complementation with candidate factors. Here we describe a method that reclaims a five-decade old mammalian cell-cell fusion strategy of functional complementation to study the molecular control of cell cycle progression. The generation of cell-cell fusions (heterokaryons) allows for the analysis, via immunofluorescence, of cell cycle regulator dynamics and evaluating the effective rescue of cell cycle progression in specific genetic settings.

ANT2 shRNA downregulates miR-19a and miR-96 through the PI3K/Akt pathway and suppresses tumor growth in hepatocellular carcinoma cells.

MicroRNAs (miRNAs) are negative regulators of gene expression, and miRNA deregulation is found in various tumors. We previously reported that suppression of adenine nucleotide translocase 2 (ANT2) by short hairpin RNA (shRNA) inhibits hepatocellular carcinoma (HCC) development by rescuing miR-636 expression. However, the tumor-suppressive mechanisms of ANT2 shRNA are still poorly understood in HCC. Here, we hypothesized that miRNAs that are specifically downregulated by ANT2 shRNA might function as oncomiRs, and we investigated the roles of ANT2 shRNA-regulated miRNAs in the pathogenesis of HCC. Our data show that miR-19a and miR-96, whose expression is regulated by ANT2 suppression, were markedly upregulated in HCC cell lines and clinical samples. Ectopic expression of miR-19a and miR-96 dramatically induced the proliferation and colony formation of hepatoma cells in vitro, whereas inhibition of miR-19a and miR-96 reduced these effects. To investigate the in vivo function, we implanted miR-96-overexpressing HepG2 cells in a xenograft model and demonstrated that the increase in miR-96 promoted tumor growth. We also found that miR-19a and miR-96 inhibited expression of tissue inhibitor of metalloproteinase-2. Taken together, our results suggest that ANT2-regulated miR-19a and miR-96 play an important role in promoting the proliferation of human HCC cells, and the knockdown of ANT2 directly downregulates miR-19a and miR-96, ultimately resulting in the suppression of tumor growth.

Exosome derived from epigallocatechin gallate treated breast cancer cells suppresses tumor growth by inhibiting tumor-associated macrophage infiltration and M2 polarization.

BACKGROUND: Tumor-associated macrophages (TAM) play an important role in tumor microenvironment. Particularly, M2 macrophages contribute to tumor progression, depending on the expression of NF-κB. Tumor-derived exosomes can modulate tumor microenvironment by transferring miRNAs to immune cells. Epigallocatechin gallate (EGCG) has well known anti-tumor effects; however, no data are available on the influence of EGCG on communication with cancer cells and TAM. METHODS: Murine breast cancer cell lines, 4T1, was used for in vivo and ex vivo studies. Exosome was extracted from EGCG-treated 4T1 cells, and the change of miRNAs was screened using microarray. Tumor cells or TAM isolated from murine tumor graft were incubated with exosomes derived from EGCG-treated and/or miR-16 inhibitor-transfected 4T1 cells. Chemokines for monocytes (CSF-1 and CCL-2), cytokines both with high (IL-6 and TGF-ß) and low (TNF-α) expression in M2 macrophages, and molecules in NF-κB pathway (IKKα and Iκ-B) were evaluated by RT-qPCR or western blot. RESULTS: EGCG suppressed tumor growth in murine breast cancer model, which was associated with decreased TAM and M2 macrophage infiltration. Expression of chemokine for monocytes (CSF-1 and CCL-2) were low in tumor cells from EGCG-treated mice, and cytokines of TAM was skewed from M2- into M1-like phenotype by EGCG as evidenced by decreased IL-6 and TGF-ß and increased TNF-α. Ex vivo incubation of isolated tumor cells with EGCG inhibited the CSF-1 and CCL-2 expression. Ex vivo incubation of TAM with exosomes from EGCG-treated 4T1 cells led to IKKα suppression and concomitant I-κB accumulation; increase of IL-6 and TGF-ß; and, decrease of TNF-α. EGCG up-regulated miR-16 in 4T1 cells and in the exosomes. Treatment of tumor cells or TAM with exosomes derived from EGCG-treated and miR-16-knock-downed 4T1 cells restored the above effects on chemokines, cytokines, and NF-κB pathway elicited by EGCG-treated exosomes. CONCLUSIONS: Our data demonstrate that EGCG up-regulates miR-16 in tumor cells, which can be transferred to TAM via exosomes and inhibits TAM infiltration and M2 polarization. We suggest a novel mechanism by which EGCG exerts anti-tumor activity via regulation of TAM in tumor microenvironment.

Exosomes derived from mesenchymal stem cells suppress angiogenesis by down-regulating VEGF expression in breast cancer cells.

Exosomes are small membrane vesicles released by a variety of cell types. Exosomes contain genetic materials, such as mRNAs and microRNAs (miRNAs), implying that they may play a pivotal role in cell-to-cell communication. Mesenchymal stem cells (MSCs), which potentially differentiate into multiple cell types, can migrate to the tumor sites and have been reported to exert complex effects on tumor progression. To elucidate the role of MSCs within the tumor microenvironment, previous studies have suggested various mechanisms such as immune modulation and secreted factors of MSCs. However, the paracrine effects of MSC-derived exosomes on the tumor microenvironment remain to be explored. The hypothesis of this study was that MSC-derived exosomes might reprogram tumor behavior by transferring their molecular contents. To test this hypothesis, exosomes from MSCs were isolated and characterized. MSC-derived exosomes exhibited different protein and RNA profiles compared with their donor cells and these vesicles could be internalized by breast cancer cells. The results demonstrated that MSC-derived exosomes significantly down-regulated the expression of vascular endothelial growth factor (VEGF) in tumor cells, which lead to inhibition of angiogenesis in vitro and in vivo. Additionally, miR-16, a miRNA known to target VEGF, was enriched in MSC-derived exosomes and it was partially responsible for the anti-angiogenic effect of MSC-derived exosomes. The collective results suggest that MSC-derived exosomes may serve as a significant mediator of cell-to-cell communication within the tumor microenvironment and suppress angiogenesis by transferring anti-angiogenic molecules.