Space travel has always been the man's ultimate destination. With the ability of spaceflight though, came the realization that exposure to microgravity has lasting effects on the human body. To ...counteract these, many studies were and are undertaken, on multiple levels. Changes in cell growth, gene, and protein expression have been described in different models on Earth and in space. Extracellular vesicles, and in particular exosomes, are important cell-cell communicators, being secreted from almost all the cells and therefore, are a perfect target to further investigate the underlying reasons of the organism's adaptations to microgravity. Here, we studied supernatants harvested from the CellBox-1 experiment, which featured human thyroid cancer cells flown to the International Space Station during the SpaceX CRS-3 cargo mission. The initial results show differences in the number of secreted exosomes, as well as in the distribution of subpopulations in regards to their surface protein expression. Notably, alteration of their population regarding the tetraspanin surface expression was observed. This is a promising step into a new area of microgravity research and will potentially lead to the discovery of new biomarkers and pathways of cellular cross-talk.
Although Bcr-Abl kinase inhibitors have proven effective in the treatment of chronic myeloid leukemia (CML), they generally fail to eradicate Bcr-Abl(+) leukemia cells. To identify genes whose ...inhibition sensitizes Bcr-Abl(+) leukemias to killing by Bcr-Abl inhibitors, we performed an RNAi-based synthetic lethal screen with imatinib mesylate in CML cells. This screen identified numerous components of a Wnt/Ca(2+)/NFAT signaling pathway. Antagonism of this pathway led to impaired NFAT activity, decreased cytokine production, and enhanced sensitivity to Bcr-Abl inhibition. Furthermore, NFAT inhibition with cyclosporin A facilitated leukemia cell elimination by the Bcr-Abl inhibitor dasatinib and markedly improved survival in a mouse model of Bcr-Abl(+) acute lymphoblastic leukemia (ALL). Targeting this pathway in combination with Bcr-Abl inhibition could improve treatment of Bcr-Abl(+) leukemias.
Non-small cell lung cancer (NSCLC) is the leading cause of cancer death worldwide. Chemotherapy, the treatment of choice in non-operable cases, achieves a dismal success rate, raising the need for ...new therapeutic options. In about 25% of NSCLC, the activating mutations of the
oncogene define a subclass that cannot benefit from tyrosine kinase inhibitors (TKIs). The tumor suppressor miR-16 is downregulated in many human cancers, including NSCLC. The main objectives of this study were to evaluate miR-16 treatment to restore the TKI sensitivity and compare its efficacy to MEK inhibitors in
-mutated NSCLC.
We performed in vitro and in vivo studies to investigate whether miR-16 could be exploited to overcome TKI resistance in KRAS-mutated NSCLC. We had three goals: first, to identify the KRAS downstream effectors targeted by mir-16, second, to study the effects of miR-16 restoration on TKI resistance in KRAS-mutated NSCLC both in vitro and in vivo, and finally, to compare miR-16 and the MEK inhibitor selumetinib in reducing KRAS-mutated NSCLC growth in vitro and in vivo.
We demonstrated that miR-16 directly targets the three
downstream effectors
,
, and
in NSCLC, restoring the sensitivity to erlotinib in
-mutated NSCLC both in vitro and in vivo. We also provided evidence that the miR-16-erlotinib regimen is more effective than the selumetinib-erlotinib combination in
-mutated NSCLC.
Our findings support the biological preclinical rationale for using miR-16 in combination with erlotinib in the treatment of NSCLC with
-activating mutations.
FTY720 (Fingolimod, Gilenya) is a sphingosine analog used as an immunosuppressant in multiple sclerosis patients. FTY720 is also a potent protein phosphatase 2A (PP2A)–activating drug (PAD). PP2A is ...a tumor suppressor found inactivated in different types of cancer. We show here that PP2A is inactive in polycythemia vera (PV) and other myeloproliferative neoplasms characterized by the expression of the transforming Jak2V617F oncogene. PP2A inactivation occurs in a Jak2V617F dose/kinase-dependent manner through the PI-3Kγ-PKC–induced phosphorylation of the PP2A inhibitor SET. Genetic or PAD-mediated PP2A reactivation induces Jak2V617F inactivation/downregulation and impairs clonogenic potential of Jak2V617F cell lines and PV but not normal CD34+ progenitors. Likewise, FTY720 decreases leukemic allelic burden, reduces splenomegaly, and significantly increases survival of Jak2V617F leukemic mice without adverse effects. Mechanistically, we show that in Jak2V617F cells, FTY720 antileukemic activity requires neither FTY720 phosphorylation (FTY720-P) nor SET dimerization or ceramide induction but depends on interaction with SET K209. Moreover, we show that Jak2V617F also utilizes an alternative sphingosine kinase-1–mediated pathway to inhibit PP2A and that FTY720-P, acting as a sphingosine-1-phosphate-receptor-1 agonist, elicits signals leading to the Jak2-PI-3Kγ-PKC-SET–mediated PP2A inhibition. Thus, PADs (eg, FTY720) represent suitable therapeutic alternatives for Jak2V617F MPNs.
•The tumor suppressor PP2A is repressed in Jak2V617F-driven myleoproliferative neoplasms by a Jak2/PI3K/PKC/SET signaling pathway.•PP2A-activating (eg, FTY720, OSU-2S) but not sphingosine-1-phosphate agonistic (eg, FTY720-P) drugs selectively kill Jak2V617F+ cells.
Altered mRNA metabolism is a feature of many cancers including blast crisis chronic myelogenous leukemia. Indeed, loss of
function of many tumor suppressors regulating cell proliferation, survival, ...and differentiation results from aberrant mRNA
processing, nuclear export, and/or translation. Here, we summarize the effects of increased BCR/ABL oncogenic activity on
the expression and function of RNA binding proteins (e.g., FUS, hnRNP A1, hnRNP E2, hnRNP K, and La/SSB) with posttranscriptional
and translational regulatory activities and their importance for the phenotype of BCR/ABL-transformed hematopoietic progenitors.
We also provide evidence that these studies not only advance our understanding on the molecular mechanisms contributing to
tumor/leukemia emergence, maintenance, and/or progression but they also serve for the identification of novel molecular targets
useful for the development of alternative therapies for imatinib-resistant and blast crisis chronic myelogenous leukemia and,
perhaps, for other cancers characterized by similar alterations in the mRNA metabolism.
The oncogenic BCR/ABL kinase activity induces and maintains chronic myelogenous leukemia (CML). We show here that, in BCR/ABL-transformed cells and CML blast crisis (CML-BC) progenitors, the ...phosphatase activity of the tumor suppressor PP2A is inhibited by the BCR/ABL-induced expression of the PP2A inhibitor SET. In imatinib-sensitive and -resistant (T315I included) BCR/ABL
+ cell lines and CML-BC progenitors, molecular and/or pharmacological activation of PP2A promotes dephosphorylation of key regulators of cell proliferation and survival, suppresses BCR/ABL activity, and induces BCR/ABL degradation. Furthermore, PP2A activation results in growth suppression, enhanced apoptosis, restored differentiation, impaired clonogenic potential, and decreased in vivo leukemogenesis of imatinib-sensitive and -resistant BCR/ABL
+ cells. Thus, functional inactivation of PP2A is essential for BCR/ABL leukemogenesis and, perhaps, required for blastic transformation.
Blast crisis chronic myelogenous leukemia (CML-BC) and Philadelphia chromosome-positive (Ph1-positive) acute lymphocytic leukemia (ALL) are 2 fatal BCR/ABL-driven leukemias against which Abl kinase ...inhibitors fail to induce a long-term response. We recently reported that functional loss of protein phosphatase 2A (PP2A) activity is important for CML blastic transformation. We assessed the therapeutic potential of the PP2A activator FTY720 (2-amino-2-2-(4-octylphenyl)ethyl-1,3-propanediol hydrochloride), an immunomodulator in Phase III trials for patients with multiple sclerosis or undergoing organ transplantation, in CML-BC and Ph1 ALL patient cells and in in vitro and in vivo models of these BCR/ABL+ leukemias. Our data indicate that FTY720 induces apoptosis and impairs clonogenicity of imatinib/dasatinib-sensitive and -resistant p210/p190(BCR/ABL) myeloid and lymphoid cell lines and CML-BC(CD34+) and Ph1 ALL(CD34+/CD19+) progenitors but not of normal CD34+ and CD34+/CD19+ bone marrow cells. Furthermore, pharmacologic doses of FTY720 remarkably suppress in vivo p210/p190(BCR/ABL)-driven including p210/p190(BCR/ABL)(T315I) leukemogenesis without exerting any toxicity. Altogether, these results highlight the therapeutic relevance of rescuing PP2A tumor suppressor activity in Ph1 leukemias and strongly support the introduction of the PP2A activator FTY720 in the treatment of CML-BC and Ph1 ALL patients.
The transcribed ultraconserved regions (T-UCRs) encode long non-coding RNAs implicated in human carcinogenesis. Their mechanisms of action and the factors regulating their expression in cancers are ...poorly understood. Here we show that high expression of uc.339 correlates with lower survival in 210 non-small cell lung cancer (NSCLC) patients. We provide evidence from cell lines and primary samples that TP53 directly regulates uc.339. We find that transcribed uc.339 is upregulated in archival NSCLC samples, functioning as a decoy RNA for miR-339-3p, -663b-3p, and -95-5p. As a result, Cyclin E2, a direct target of all these microRNAs is upregulated, promoting cancer growth and migration. Finally, we find that modulation of uc.339 affects microRNA expression. However, overexpression or downregulation of these microRNAs causes no significant variations in uc.339 levels, suggesting a type of interaction for uc.339 that we call "entrapping". Our results support a key role for uc.339 in lung cancer.
The capture of tumour‐derived extracellular vesicles (TEVs) by cells in the tumour microenvironment (TME) contributes to metastasis and notably to the formation of the pre‐metastatic niche (PMN). ...However, due to the challenges associated with modelling release of small EVs in vivo, the kinetics of PMN formation in response to endogenously released TEVs have not been examined. Here, we have studied the endogenous release of TEVs in mice orthotopically implanted with metastatic human melanoma (MEL) and neuroblastoma (NB) cells releasing GFP‐tagged EVs (GFTEVs) and their capture by host cells to demonstrate the active contribution of TEVs to metastasis. Human GFTEVs captured by mouse macrophages in vitro resulted in transfer of GFP vesicles and the human exosomal miR‐1246. Mice orthotopically implanted with MEL or NB cells showed the presence of TEVs in the blood between 5 and 28 days after implantation. Moreover, kinetic analysis of TEV capture by resident cells relative to the arrival and outgrowth of TEV‐producing tumour cells in metastatic organs demonstrated that the capture of TEVs by lung and liver cells precedes the homing of metastatic tumour cells, consistent with the critical roles of TEVs in PMN formation. Importantly, TEV capture at future sites of metastasis was associated with the transfer of miR‐1246 to lung macrophages, liver macrophages, and stellate cells. This is the first demonstration that the capture of endogenously released TEVs is organotropic as demonstrated by the presence of TEV‐capturing cells only in metastatic organs and their absence in non‐metastatic organs. The capture of TEVs in the PMN induced dynamic changes in inflammatory gene expression which evolved to a pro‐tumorigenic reaction as the niche progressed to the metastatic state. Thus, our work describes a novel approach to TEV tracking in vivo that provides additional insights into their role in the earliest stages of metastatic progression.
Oncogenic mutations of the receptor tyrosine kinase c-KIT play an important role in the pathogenesis of gastrointestinal stromal tumors, systemic mastocytosis, and some acute myeloid leukemias (AML). ...Although juxtamembrane mutations commonly detected in gastrointestinal stromal tumor are sensitive to tyrosine kinase inhibitors, the kinase domain mutations frequently encountered in systemic mastocytosis and AML confer resistance and are largely unresponsive to targeted inhibition by the existing agent imatinib. In this study, we show that myeloid cells expressing activated c-KIT mutants that are imatinib sensitive (V560G) or imatinib resistant (D816V) can inhibit the tumor suppressor activity of protein phosphatase 2A (PP2A). This effect was associated with the reduced expression of PP2A structural (A) and regulatory subunits (B55alpha, B56alpha, B56gamma, and B56delta). Overexpression of PP2A-Aalpha in D816V c-KIT cells induced apoptosis and inhibited proliferation. In addition, pharmacologic activation of PP2A by FTY720 reduced proliferation, inhibited clonogenic potential, and induced apoptosis of mutant c-KIT(+) cells, while having no effect on wild-type c-KIT cells or empty vector controls. FTY720 treatment caused the dephosphorylation of the D816V c-KIT receptor and its downstream signaling targets pAkt, pSTAT5, and pERK1/2. Additionally, in vivo administration of FTY720 delayed the growth of V560G and D816V c-KIT tumors, inhibited splenic and bone marrow infiltration, and prolonged survival. Our findings show that PP2A inhibition is essential for c-KIT-mediated tumorigenesis, and that reactivating PP2A may offer an attractive strategy to treat drug-resistant c-KIT(+) cancers.
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