Exosomes derived from mesenchymal stem cells enhance radiotherapy-induced cell death in tumor and metastatic tumor foci.

BACKGROUND: We have recently shown that radiotherapy may not only be a successful local and regional treatment but, when combined with MSCs, may also be a novel systemic cancer therapy. This study aimed to investigate the role of exosomes derived from irradiated MSCs in the delay of tumor growth and metastasis after treatment with MSC + radiotherapy (RT). METHODS: We have measured tumor growth and metastasis formation, of subcutaneous human melanoma A375 xenografts on NOD/SCID-gamma mice, and the response of tumors to treatment with radiotherapy (2 Gy), mesenchymal cells (MSC), mesenchymal cells plus radiotherapy, and without any treatment. Using proteomic analysis, we studied the cargo of the exosomes released by the MSC treated with 2 Gy, compared with the cargo of exosomes released by MSC without treatment. RESULTS: The tumor cell loss rates found after treatment with the combination of MSC and RT and for exclusive RT, were: 44.4% % and 12,1%, respectively. Concomitant and adjuvant use of RT and MSC, increased the mice surviving time 22,5% in this group, with regard to the group of mice treated with exclusive RT and in a 45,3% respect control group. Moreover, the number of metastatic foci found in the internal organs of the mice treated with MSC + RT was 60% less than the mice group treated with RT alone. We reasoned that the exosome secreted by the MSC, could be implicated in tumor growth delay and metastasis control after treatment. CONCLUSIONS: Our results show that exosomes derived form MSCs, combined with radiotherapy, are determinant in the enhancement of radiation effects observed in the control of metastatic spread of melanoma cells and suggest that exosome-derived factors could be involved in the bystander, and abscopal effects found after treatment of the tumors with RT plus MSC. Radiotherapy itself may not be systemic, although it might contribute to a systemic effect when used in combination with mesenchymal stem cells owing the ability of irradiated MSCs-derived exosomes to increase the control of tumor growth and metastasis.

Vasculogenic mimicry signaling revisited: focus on non-vascular VE-cadherin.

Vasculogenic mimicry (VM) is a blood supply system independent of endothelial vessels in tumor cells from different origins. It reflects the plasticity of aggressive tumor cells that express vascular cell markers and line tumor vasculature. The presence of VM is associated with a high tumor grade, short survival, invasion and metastasis. Endothelial cells (ECs) express various members of the cadherin superfamily, in particular vascular endothelial (VE-) cadherin, which is the main adhesion receptor of endothelial adherent junctions. Aberrant extra-vascular expression of VE-cadherin has been observed in certain cancer types associated with VM. In this review we focus on non-endothelial VE-cadherin as a prominent factor involved in the acquisition of tubules-like structures by aggressive tumor cells and we summarize the specific signaling pathways, the association with trans-differentiation and stem-like phenotype and the therapeutic opportunities derived from the in-depth knowledge of the peculiarities of the biology of VE-cadherin and other key components of VM.

Enhancer Clusters Drive Type I Interferon-Induced TRAIL Overexpression in Cancer, and Its Intracellular Protein Accumulation Fails to Induce Apoptosis.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytokine produced and secreted by immune cells in response to an infection, often in response to interferon (IFN) stimulation. In cancer, it has also been shown that IFN stimulates the production of TRAIL, and it has been proposed that this TRAIL can induce apoptosis in an autocrine or paracrine manner in different cancer cells. Yet, the mechanism mediating TRAIL upregulation and the implications of TRAIL as an apoptotic molecule in cancer cells are still poorly understood. We show here that in certain cancer cells, TRAIL is upregulated by enhancer clusters, potent genomic regulatory regions containing densely packed enhancers that have combinatorial and additive activity and that are usually found to be associated with cancer-promoting genes. Moreover, we found that TRAIL upregulation by IFNα is mediated by these enhancer clusters in breast and lung cancer cells. Surprisingly, IFNα stimulation leads to the intracellular accumulation of TRAIL protein in these cancer cells. Consequently, this TRAIL is not capable of inducing apoptosis. Our study provides novel insights into the mechanism behind the interferon-mediated upregulation of TRAIL and its protein accumulation in cancer cells. Further investigation is required to understand the role of intracellular TRAIL or depict the mechanisms mediating its apoptosis impairment in cancer cells.

The Multifactorial Role of PARP-1 in Tumor Microenvironment.

Poly(ADP-ribose) polymerases (PARPs), represent a family of 17 proteins implicated in a variety of cell functions; some of them possess the enzymatic ability to synthesize and attach poly (ADP-ribose) (also known as PAR) to different protein substrates by a post-translational modification; PARPs are key components in the cellular response to stress with consequences for different physiological and pathological events, especially during neoplasia. In recent years, using PARP inhibitors as antitumor agents has raised new challenges in understanding their role in tumor biology. Notably, the function of PARPs and PAR in the dynamic of tumor microenvironment is only starting to be understood. In this review, we summarized the conclusions arising from recent studies on the interaction between PARPs, PAR and key features of tumor microenvironment such as hypoxia, autophagy, tumor initiating cells, angiogenesis and cancer-associated immune response.

VE-cadherin promotes vasculogenic mimicry by modulating kaiso-dependent gene expression.

Aberrant extra-vascular expression of VE-cadherin (VEC) has been observed in metastasis associated with vasculogenic mimicry (VM); however, the ultimate reason why non-endothelial VEC favors the acquisition of this phenotype is not established. In this study, we show that human malignant melanoma cells have a constitutively high expression of phoshoVEC (pVEC) at Y658; pVEC is a target of focal adhesion kinase (FAK) and forms a complex with p120-catenin and the transcriptional repressor kaiso in the nucleus. FAK inhibition enabled kaiso to suppress the expression of its target genes and enhanced kaiso recruitment to KBS-containing promoters. Finally we have found that ablation of kaiso-repressed genes WNT11 and CCDN1 abolished VM. Thus, identification of pVEC as a component of the kaiso transcriptional complex establishes a molecular paradigm that links FAK-dependent phosphorylation of VEC as a major mechanism by which ectopical VEC expression exerts its function in VM.

PIM kinases mediate resistance of glioblastoma cells to TRAIL by a p62/SQSTM1-dependent mechanism.

Glioblastoma (GBM) is the most common and aggressive brain tumor and is associated with poor prognosis. GBM cells are frequently resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and finding new combinatorial therapies to sensitize glioma cells to TRAIL remains an important challenge. PIM kinases are serine/threonine kinases that promote cell survival and proliferation and are highly expressed in different tumors. In this work, we studied the role of PIM kinases as regulators of TRAIL sensitivity in GBM cells. Remarkably, PIM inhibition or knockdown facilitated activation by TRAIL of a TRAIL-R2/DR5-mediated and mitochondria-operated apoptotic pathway in TRAIL-resistant GBM cells. The sensitizing effect of PIM knockdown on TRAIL-induced apoptosis was mediated by enhanced caspase-8 recruitment to and activation at the death-inducing signaling complex (DISC). Interestingly, TRAIL-induced internalization of TRAIL-R2/DR5 was significantly reduced in PIM knockdown cells. Phospho-proteome profiling revealed a decreased phosphorylation of p62/SQSTM1 after PIM knockdown. Our results also showed an interaction between p62/SQSTM1 and the DISC that was reverted after PIM knockdown. In line with this, p62/SQSTM1 ablation increased TRAIL-R2/DR5 levels and facilitated TRAIL-induced caspase-8 activation, revealing an inhibitory role of p62/SQSTM1 in TRAIL-mediated apoptosis in GBM. Conversely, upregulation of TRAIL-R2/DR5 upon PIM inhibition and apoptosis induced by the combination of PIM inhibitor and TRAIL were abrogated by a constitutively phosphorylated p62/SQSTM1S332E mutant. Globally, our data represent the first evidence that PIM kinases regulate TRAIL-induced apoptosis in GBM and identify a specific role of p62/SQSTM1Ser332 phosphorylation in the regulation of the extrinsic apoptosis pathway activated by TRAIL.

PARP targeting counteracts gliomagenesis through induction of mitotic catastrophe and aggravation of deficiency in homologous recombination in PTEN-mutant glioma.

Glioblastoma multiforme (GBM) is the most common primary brain tumour in adults and one of the most aggressive cancers. PARP-1 is a nuclear protein involved in multiple facets of DNA repair and transcriptional regulation. In this study we dissected the action of PARP inhibition in different GBM cell lines with either functional or mutated PTEN that confers resistance to diverse therapies. In PTEN mutant cells, PARP inhibition induced a severe genomic instability, exacerbated homologous recombination repair (HR) deficiency and down-regulated the Spindle Assembly Checkpoint (SAC) factor BUBR1, leading to mitotic catastrophe (MC). EGFR gene amplification also represents a signature of genetic abnormality in GBM. To more effectively target GBM cells, co-treatment with a PARP inhibitor and an EGFR blocker, erlotinib, resulted in a strong suppression of ERK1/2 activation and in vivo the combined effect elicited a robust reduction in tumour development. In conclusion, PARP inhibition targets PTEN-deficient GBM cells through accentuation of SAC repression and aggravation of HR deficiency, leading to the induction of genomic instability and eventually deriving to mitotic catastrophe (MC); the inhibition of PARP and co-treatment with an inhibitor of pro-survival pathways strongly retarded in vivo gliomagenesis.