Stearoyl CoA Desaturase-1 Silencing in Glioblastoma Cells: Phospholipid Remodeling and Cytotoxicity Enhanced upon Autophagy Inhibition.

Modulation of lipid metabolism is a well-established cancer hallmark, and SCD1 has been recognized as a key enzyme in promoting cancer cell growth, including in glioblastoma (GBM), the deadliest brain tumor and a paradigm of cancer resistance. The central goal of this work was to identify, by MS, the phospholipidome alterations resulting from the silencing of SCD1 in human GBM cells, in order to implement an innovative therapy to fight GBM cell resistance. With this purpose, RNAi technology was employed, and low serum-containing medium was used to mimic nutrient deficiency conditions, at which SCD1 is overexpressed. Besides the expected increase in the saturated to unsaturated fatty acid ratio in SCD1 silenced-GBM cells, a striking increase in polyunsaturated chains, particularly in phosphatidylethanolamine and cardiolipin species, was noticed and tentatively correlated with an increase in autophagy (evidenced by the increase in LC3BII/I ratio). The contribution of autophagy to mitigate the impact of SCD1 silencing on GBM cell viability and growth, whose modest inhibition could be correlated with the maintenance of energetically associated mitochondria, was evidenced by using autophagy inhibitors. In conclusion, SCD1 silencing could constitute an important tool to halt GBM resistance to the available treatments, especially when coupled with a mitochondria disrupter chemotherapeutic.

MiR-200c-based metabolic modulation in glioblastoma cells as a strategy to overcome tumor chemoresistance.

Glioblastoma (GB) is the most aggressive and common form of primary brain tumor characterized by fast proliferation, high invasion and resistance to current standard treatment. The average survival rate post-diagnosis is 14.6 months, despite the aggressive standard post-surgery radiotherapy concomitant with chemotherapy with temozolomide (TMZ). Currently, efforts are being endowed to develop new and more efficient therapeutic approaches capable to overcome chemoresistance, inhibit tumor progression and improve overall patient survival rate. Abnormal microRNA (miRNA) expression has been correlated with chemoresistance, proliferation and resistance to apoptosis, which result from their master regulatory role of gene expression. Altered cell metabolism, favoring glycolysis, was identified as an emerging cancer hallmark and has been described in GB, thus offering a new target for innovative GB therapies. In this work, we hypothesized that a gene therapy-based strategy consisting of the overexpression of a miRNA downregulated in GB and predicted to target crucial metabolic enzymes might promote a shift of GB cell metabolism, decreasing the glycolytic dependence of tumor cells and contributing to their sensitization to chemotherapy with TMZ. The increase of miR-200c levels in DBTRG cells resulted in downregulation of messenger RNA of enzymes involved in bioenergetics pathways and impaired cell metabolism and mobility. In addition, miR-200c overexpression prior to DBTRG cell exposure to TMZ resulted in cell cycle arrest. Overall, our results show that miR-200c overexpression could offer a way to overcome chemoresistance developed by GB cells in response to current standard chemotherapy, providing an improvement to current GB standard treatment, with benefit for patient outcome.

Improving pollutants environmental risk assessment using a multi model toxicity determination with in vitro, bacterial, animal and plant model systems: The case of the herbicide alachlor.

Several environmental pollutants, including pesticides, herbicides and persistent organic pollutants play an important role in the development of chronic diseases. However, most studies have examined environmental pollutants toxicity in target organisms or using a specific toxicological test, losing the real effect throughout the ecosystem. In this sense an integrative environmental risk of pollutants assessment, using different model organisms is necessary to predict the real impact in the ecosystem and implications for target and non-target organisms. The objective of this study was to use alachlor, a chloroacetanilide herbicide responsible for chronic toxicity, to understand its impact in target and non-target organisms and at different levels of biological organization by using several model organisms, including membranes of dipalmitoylphosphatidylcholine (DPPC), rat liver mitochondria, bacterial (Bacillus stearothermophilus), plant (Lemna gibba) and mammalian cell lines (HeLa and neuro2a). Our results demonstrated that alachlor strongly interacted with membranes of DPPC and interfered with mitochondrial bioenergetics by reducing the respiratory control ratio and the transmembrane potential. Moreover, alachlor also decreased the growth of B. stearothermophilus and its respiratory activity, as well as decreased the viability of both mammalian cell lines. The values of TC50 increased in the following order: Lemna gibba < neuro2a < HeLa cells < Bacillus stearothermophilus. Together, the results suggest that biological membranes constitute a putative target for the toxic action of this lipophilic herbicide and point out the risks of its dissemination on environment, compromising ecosystem equilibrium and human health.

Anti-inflammatory activity of novel thiosemicarbazone compounds indole-based as COX inhibitors.

BACKGROUND: In this article, a series of 20 new thiosemicarbazone derivatives containing indole were synthesized and evaluated for their anti-inflammatory potential. METHODS: The compounds were obtained through a synthetic route of only two steps, with yields that varied between 33.6 and 90.4%, and characterized by spectroscopic and spectrometric techniques. RESULTS: An initial screening through the lymphoproliferation assay revealed that compounds LT76, LT81, and LT87 were able to inhibit lymphocyte proliferation, with CC50 of 0.56 ± 0.036, 0.9 ± 0.01 and 0.5 ± 0.07 µM, respectively, better results than indomethacin (CC50 > 12 µM). In addition, these compounds were able to suppress the in-vitro production of TNF-α and NO, in addition to stimulating the production of IL-4. Reinforcing in-vitro assays, the compounds were able to inhibit COX-2 similar to Celecoxib showing greater selectivity for this isoform (LT81 SI: 23.06 versus Celecoxib SI: 11.88). Animal studies showed that compounds LT76 (64.8% inhibition after 6 h), LT81 (89% inhibition after 6 h) and LT87 (100% inhibition after 4 h) were able to suppress edema in mice after inoculation carrageenan with greater potency than indomethacin, and immunohistochemistry revealed that the groups treated with LT76, LT81 and LT87 reduced the expression of COX-2, similar or better results when compared to indomethacin. Complementarily, in-silico studies have shown that these compounds have a good pharmacokinetic profile, for respecting the parameters of Lipinski and Veber, showing their good bioavailability. CONCLUSIONS: These results demonstrate the potency of thiosemicarbazone derivatives containing indole and confirm their importance as scaffolds of molecules with notorious anti-inflammatory activity.

Differentiation of glioblastoma stem cells promoted by miR-128 or miR-302a overexpression enhances senescence-associated cytotoxicity of axitinib.

Despite the intense global efforts towards an effective treatment of glioblastoma (GB), current therapeutic options are unsatisfactory with a median survival time of 12-15 months after diagnosis, which has not improved significantly over more than a decade. The high tumoral heterogeneity confers resistance to therapies, which has hindered a successful clinical outcome, GB remaining among the deadliest cancers. A hallmark of GB is its high recurrence rate, which has been attributed to the presence of a small subpopulation of tumor cells called GB stem-like cells (GSC). In the present work, the efficacy of a multimodal strategy combining microRNA (miRNA) modulation with new generation multitargeted tyrosine kinase inhibitors (imatinib and axitinib) was investigated aiming at tackling this subpopulation of GB cells. MiR-128 and miR-302a were selected as attractive therapeutic candidates on the basis of previous findings reporting that reestablishment of their decreased expression levels in GSC resulted in cell differentiation, which could represent a possible strategy to sensitize GSC to chemotherapy. Our results show that overexpression of miR-128 or miR-302a induced GSC differentiation, which enhanced senescence mediated by axitinib treatment, thus further impairing GSC proliferation. We also provided evidence for the capacity of GSC to efficiently internalize functionalized stable nucleic acid lipid particles, previously developed and successfully applied in our laboratory to target GB. Taken together, our findings will be important in the future design of a GB-targeted multimodal miRNA-based gene therapy, combining overexpression of miR-128 or miR-302a with axitinib treatment, endowed with the ability to overcome drug resistance.

Downregulation of long non-protein coding RNA MVIH impairs glioblastoma cell proliferation and invasion through an miR-302a-dependent mechanism.

Glioblastoma (GB) is the most frequent and malignant type of brain tumor, for which no effective therapy exists. The high proliferative and invasive nature of GB, as well as its acquired resistance to chemotherapy, makes this type of cancer extremely lethal shortly after diagnosis. Long non-protein coding RNAs (lncRNA) are a class of regulatory RNAs whose levels can be dysregulated in the context of diseases, unbalancing several physiological processes. The lncRNA associated with microvascular invasion in hepatocellular carcinoma (lncRNA-MVIH), overexpressed in several cancers, was described to co-precipitate with phosphoglycerate kinase 1 (PGK1), preventing secretion of this enzyme to the extracellular environment and promoting cell migration and invasion. We hypothesized that, by silencing the expression of lncRNA-MVIH, the secretion of PGK1 would increase, reducing GB cell migration and invasion capabilities. We observed that lncRNA-MVIH silencing in human GB cells significantly decreased glycolysis, cell growth, migration, and invasion and sensitized GB cells to cediranib. However, no increase in extracellular PGK1 was observed as a consequence of lncRNA-MVIH silencing, and therefore, we investigated the possibility of a mechanism of miRNA sponge of lncRNA-MVIH being in place. We found that the levels of miR-302a loaded onto RISC increased in GB cells after lncRNA-MVIH silencing, with the consequent downregulation of several miR-302a molecular targets. Our findings suggest a new mechanism of action of lncRNA-MVIH as a sponge of miR-302a. We suggest that lncRNA-MVIH knockdown may be a promising strategy to address GB invasiveness and chemoresistance, holding potential towards its future application in a clinical context.

Lauroylated Histidine-Enriched S413-PV Peptide as an Efficient Gene Silencing Mediator in Cancer Cells.

PURPOSE: This study aimed to endow the cell-penetrating peptide (CPP) S413-PV with adequate features towards a safe and effective application in cancer gene therapy. METHODS: Peptide/siRNA complexes were prepared with two new derivatives of the CPP S413-PV, which combine a lauroyl group attached to the N- or C-terminus with a histidine-enrichment in the N-terminus of the S413-PV peptide, being named C12-H5-S413-PV and H5-S413-PV-C12, respectively. Physicochemical characterization of siRNA complexes was performed and their cytotoxicity and efficiency to mediate siRNA delivery and gene silencing in cancer cells were assessed in the absence and presence of serum. RESULTS: Peptide/siRNA complexes prepared with the C12-H5-S413-PV derivative showed a nanoscale (ca. 100 nm) particle size, as revealed by TEM, and efficiently mediated gene silencing (37%) in human U87 glioblastoma cells in the presence of 30% serum. In addition, the new C12-H5-S413-PV-based siRNA delivery system efficiently downregulated stearoyl-CoA desaturase-1, a key-enzyme of lipid metabolism overexpressed in cancer, which resulted in a significant decrease in the viability of U87 cells. Importantly, these complexes were able to spare healthy human astrocytes. CONCLUSIONS: These encouraging results pave the way for a potential application of the C12-H5-S413-PV peptide as a promising tool in cancer gene therapy.

Lysosomal Storage Disease-Associated Neuropathy: Targeting Stable Nucleic Acid Lipid Particle (SNALP)-Formulated siRNAs to the Brain as a Therapeutic Approach.

More than two thirds of Lysosomal Storage Diseases (LSDs) present central nervous system involvement. Nevertheless, only one of the currently approved therapies has an impact on neuropathology. Therefore, alternative approaches are under development, either addressing the underlying enzymatic defect or its downstream consequences. Also under study is the possibility to block substrate accumulation upstream, by promoting a decrease of its synthesis. This concept is known as substrate reduction therapy and may be triggered by several molecules, such as small interfering RNAs (siRNAs). siRNAs promote RNA interference, a naturally occurring sequence-specific post-transcriptional gene-silencing mechanism, and may target virtually any gene of interest, inhibiting its expression. Still, naked siRNAs have limited cellular uptake, low biological stability, and unfavorable pharmacokinetics. Thus, their translation into clinics requires proper delivery methods. One promising platform is a special class of liposomes called stable nucleic acid lipid particles (SNALPs), which are characterized by high cargo encapsulation efficiency and may be engineered to promote targeted delivery to specific receptors. Here, we review the concept of SNALPs, presenting a series of examples on their efficacy as siRNA nanodelivery systems. By doing so, we hope to unveil the therapeutic potential of these nanosystems for targeted brain delivery of siRNAs in LSDs.

Glucosylceramide synthase silencing combined with the receptor tyrosine kinase inhibitor axitinib as a new multimodal strategy for glioblastoma.

A great deal of evidence revealing that lipid metabolism is drastically altered during tumorigenesis has been accumulated. In this work, glucosylceramide synthase (GCS) was targeted, using RNA interference technology (siRNAs), in U87 and DBTRG human glioblastoma (GBM) cells, as in both cell types GCS showed to be overexpressed with respect to normal human astrocytes. The efficacy of a combined therapy to tackle GBM, allying GCS silencing to the new generation chemotherapeutics sunitinib and axitinib, or to the alkylating drugs etoposide and temozolomide, is evaluated here for the first time. With this purpose, studies addressing GBM cell viability and proliferation, cell cycle and apoptosis were performed, which revealed that combination of GCS silencing with axitinib treatment represents a promising therapeutic approach. The reduction of cell viability induced by this combined therapy is proposed to be mediated by excessive production of reactive oxygen species. This work, identifying GCS as a key molecular target to increase GBM susceptibility to a new generation chemotherapeutic, opens windows to the development of innovative strategies to halt GBM recurrence after surgical resection.

Suicide Gene Therapy for Oral Squamous Cell Carcinoma.

Suicide gene therapy has been tested for the treatment of a variety of cancers, including oral cancer. Among the various suicide gene therapy approaches that have been reported, the Herpes Simplex Virus thymidine kinase (HSV-tk)/ganciclovir (GCV) system is one of the most extensively studied systems, holding great promise in cancer therapy. In this chapter, we describe methods to use the HSV-tk/GCV system to achieve antitumor activity, both in cultured oral cancer cells and in orthotopic and subcutaneous murine models of oral squamous cell carcinoma, using ligand-associated lipoplexes for enhancing therapeutic delivery.

Acylation of the S413-PV cell-penetrating peptide as a means of enhancing its capacity to mediate nucleic acid delivery: Relevance of peptide/lipid interactions.

BACKGROUND: Cell-penetrating peptides (CPPs) have been extensively exploited in gene therapy approaches as vectors for intracellular delivery of bioactive molecules. The ability of CPPs to be internalized into cells and their capacity to complex nucleic acids depend on their molecular structure, both primary and secondary, namely regarding hydrophobicity/hydrophilicity. CPP acylation has been used as a strategy to improve this structural feature. METHODS: Acyl groups (from 6 to 18 carbon atoms) were attached to the S413-PV peptide and their effects on the peptide competence to complex siRNAs and to mediate gene silencing in glioblastoma (GBM) cells were studied. A systematic characterization of membrane interactions with S413-PV acyl-derivatives was also conducted, using different biophysical techniques (surface pressure-area isotherms in Langmuir monolayers, DSC and 31P NMR) to unravel a relationship between CPP biological activity and CPP effects on membrane stability and lipid organization. RESULTS: A remarkable concordance was noticed between acylated-S413-PV peptide competence to promote gene silencing in GBM cells and disturbance induced in membrane models, the lauroyl- and myristoyl-S413-PV peptides being the most effective. A cut-off effect was described for the first time regarding the influence of acyl-chain length on CPP bioactivity. CONCLUSIONS: C12-S413-PV showed high capacity to destabilize lipid bilayers, to escape from lysosomal degradation and to mediate gene silencing without promoting cytotoxicity. GENERAL SIGNIFICANCE: Besides unraveling a new CPP with high potential to be employed as a gene delivery vector, this work emphasizes the benefit from allying biophysical and biological studies towards a proper CPP structural refinement for successful pre-clinical/clinical application.

Multimodal highly fluorescent-magnetic nanoplatform to target transferrin receptors in cancer cells.

BACKGROUND: Site-specific multimodal nanoplatforms with fluorescent-magnetic properties have great potential for biological sciences. For this reason, we developed a multimodal nanoprobe (BNPs-Tf), by covalently conjugating an optical-magnetically active bimodal nanosystem, based on quantum dots and iron oxide nanoparticles, with the human holo-transferrin (Tf). METHODS: The Tf bioconjugation efficiency was evaluated by the fluorescence microplate assay (FMA) and the amount of Tf immobilized on BNPs was quantified by fluorescence spectroscopy. Moreover, relaxometric and fluorescent properties of the BNPs-Tf were evaluated, as well as its ability to label specifically HeLa cells. Cytotoxicity was also performed by Alamar Blue assay. RESULTS: The FMA confirmed an efficient bioconjugation and the fluorescence spectroscopy analysis indicated that 98% of Tf was immobilized on BNPs. BNPs-Tf also presented a bright fluorescence and a transversal/longitudinal relaxivities ratio (r2/r1) of 65. Importantly, the developed BNPs-Tf were able to label, efficiently and specifically, the Tf receptors in HeLa cells, as shown by fluorescence and magnetic resonance imaging assays. Moreover, this multimodal system did not cause noteworthy cytotoxicity. CONCLUSIONS: The prepared BNPs-Tf hold great promise as an effective and specific multimodal, highly fluorescent-magnetic, nanoplatform for fluorescence analyses and T2-weighted images. GENERAL SIGNIFICANCE: This study developed an attractive and versatile multimodal nanoplatform that has potential to be applied in a variety of in vitro and in vivo studies, addressing biological processes, diagnostic, and therapeutics. Moreover, this work opens new possibilities for designing other efficient multimodal nanosystems, considering other biomolecules in their composition able to provide them important functional properties.

High-throughput screening uncovers miRNAs enhancing glioblastoma cell susceptibility to tyrosine kinase inhibitors.

Glioblastoma (GBM) is a deadly and therapy resistant malignant brain tumour, characterized by an aggressive and diffuse growth pattern, which prevents complete surgical resection. Despite advances in the identification of genomic and molecular alterations that fuel the tumour, average patient survival post-diagnosis remains very low (∼14.6-months). In addition to being highly heterogeneous, GBM tumour cells exhibit high adaptive capacity to targeted molecular therapies owing to an established network of signalling cascades with functional redundancy, which provides them with robust compensatory survival mechanisms. Here, we investigated whether a multimodal strategy combining multitargeted tyrosine kinase inhibitors (MTKIs) and microRNA (miRNA) modulation could overcome the signalling pathway redundancy in GBM and, hence, promote tumour cell death. By performing a high-throughput screening, we identified a myriad of miRNAs, including those belonging to the miR-302-3p/372-3p/373-3p/520-3p family, which coordinately act with the MTKI sunitinib to decrease GBM cell viability. Two members of this family, hsa-miRNA-302a-3p and hsa-miRNA-520 b, were found to modulate the expression of receptor tyrosine kinase mediators (including AKT1, PIK3CA and SOS1) in U87 and DBTRG human GBM cells. Importantly, administration of mimics of these miRNAs with sunitinib or axitinib resulted in decreased tumour cell proliferation and enhanced cell death, whereas no significant effect was observed when coupling miRNA modulation with temozolomide, the first-line drug for GBM therapy. Overall, our results provide evidence that combining the 'horizontal' inhibition of signalling pathways promoted by MTKIs with the 'vertical' inhibition of the downstream signalling cascade promoted by hsa-miR-302a-3p and hsa-miR-520 b constitutes a promising approach towards GBM treatment.

Resveratrol stimulates the metabolic reprogramming of human CD4+ T cells to enhance effector function.

The polyphenol resveratrol activates the deacetylase Sirt1, resulting in various antioxidant, chemoprotectant, neuroprotective, cardioprotective, and anti-inflammatory properties. We found that at high concentrations of resveratrol, human CD4+ T cells showed defective antigen receptor signaling and arrest at the G1 stage of the cell cycle, whereas at low concentrations, cells were readily activated and exhibited enhanced Sirt1 deacetylase activity. Nevertheless, low-dose resveratrol rapidly stimulated genotoxic stress in the T cells, which resulted in engagement of a DNA damage response pathway that depended on the kinase ATR [ataxia telangiectasia-mutated (ATM) and Rad3-related], but not ATM, and subsequently in premitotic cell cycle arrest. The concomitant activation of p53 was coupled to the expression of gene products that regulate cell metabolism, leading to a metabolic reprogramming that was characterized by decreased glycolysis, increased glutamine consumption, and a shift to oxidative phosphorylation. These alterations in the bioenergetic homeostasis of CD4+ T cells resulted in enhanced effector function, with both naïve and memory CD4+ T cells secreting increased amounts of the inflammatory cytokine interferon-γ. Thus, our data highlight the wide range of metabolic adaptations that CD4+ T lymphocytes undergo in response to genomic stress.

MicroRNA deregulation and chemotaxis and phagocytosis impairment in Alzheimer's disease.

INTRODUCTION: Mononuclear phagocytes play a critical role during Alzheimer's disease (AD) pathogenesis due to their contribution to innate immune responses and amyloid beta (Aß) clearance mechanisms. METHODS: Blood-derived monocytes (BDMs) and monocyte-derived macrophages (MDMs) were isolated from blood of AD, mild cognitive impairment (MCI) patients, and age-matched healthy controls for molecular and phenotypic comparisons. RESULTS: The chemokine/chemokine receptor CCL2/CCR2 axis was impaired in BDMs from AD and MCI patients, causing a deficit in cell migration. Changes were also observed in MDM-mediated phagocytosis of Aß fibrils, correlating with alterations in the expression and processing of the triggering receptor expressed on myeloid cells 2 (TREM2). Finally, immune-related microRNAs (miRNAs), including miR-155, -154, -200b, -27b, and -128, were found to be differentially expressed in these cells. DISCUSSION: This work provides evidence that chemotaxis and phagocytosis, two crucial innate immune functions, are impaired in AD and MCI patients. Correlations with miRNA levels suggest an epigenetic contribution to systemic immune dysfunction in AD.

Enhancing glioblastoma cell sensitivity to chemotherapeutics: A strategy involving survivin gene silencing mediated by gemini surfactant-based complexes.

Glioblastoma (GBM), the highest grade astrocytoma, is one of the most aggressive and challenging cancers to treat. The standard treatment is usually limited due to the intrinsic resistance of GBM to chemotherapy and drug non-specific effects. Therefore, new therapeutic strategies need to be developed to target tumor cells, sparing healthy tissues. In this context, the inhibitor-of-apoptosis protein (IAP) survivin emerges as an ideal target for a gene silencing approach, since it is sharply differentially expressed in cancer tissues. In this work, two different families of cationic gemini surfactants (bis-quat conventional and serine-derived) were tested regarding their efficiency to deliver small interfering RNAs (siRNAs) in a human GBM cell line (U87), in order to select an effective siRNA anti-survivin carrier. Importantly, survivin downregulation combined with administration of the chemotherapeutic agents temozolomide or etoposide resulted in a synergistic cytotoxic effect, thus revealing to be a promising strategy to reduce the chemotherapeutic doses for GBM treatment.

CdTe quantum dots as fluorescent probes to study transferrin receptors in glioblastoma cells.

BACKGROUND: Overexpression of transferrin receptors (TfRs), which are responsible for the intracellular uptake of ferric transferrin (Tf), has been described in various cancers. Although molecular biology methods allow the identification of different types of receptors in cancer cells, they do not provide features about TfRs internalization, quantification and distribution on cell surface. This information can, however, be accessed by fluorescence techniques. In this work, the quantum dots (QDs)' unique properties were explored to strengthen our understanding of TfRs in cancer cells. METHODS: QDs were conjugated to Tf by covalent coupling and QDs-(Tf) bioconjugates were applied to quantify and evaluate the distribution of TfRs in two human glioblastoma cells lines, U87 and DBTRG-05MG, and also in HeLa cells by using flow cytometry and confocal microscopy. RESULTS: HeLa and DBTRG-05MG cells showed practically the same TfR labeling profile by QDs-(Tf), while U87 cells were less labeled by bioconjugates. Furthermore, inhibition studies demonstrated that QDs-(Tf) were able to label cells with high specificity. CONCLUSIONS: HeLa and DBTRG-05MG cells presented a similar and a higher amount of TfR than U87 cells. Moreover, DBTRG-05MG cells are more efficient in recycling the TfR than the other two cells types. GENERAL SIGNIFICANCE: This is the first study about TfRs in human glioblastoma cells using QDs. This new fluorescent tool can contribute to our understanding of the cancer cell biology and can help in the development of new therapies targeting these receptors.

Role of microRNAs in the regulation of innate immune cells under neuroinflammatory conditions.

MiRNAs are short, evolutionary conserved noncoding RNA molecules with the ability to control the magnitude of inflammation. The immunosuppressive nature of the brain is sustained by miRNA-dependent regulation of microglial cells, which become activated under neuroinflammatory conditions, such as brain injury and neurodegeneration. The pro-inflammatory and suppressive role of the most studied neuroimmune miRNAs, miR-155 and miR-146a, has been recently challenged. Although the molecular targets of these miRNAs remain unchanged across brain diseases, different kinetics of miRNA expression and degradation can produce different immune outcomes and change microglia phenotypes. Here, we discuss current knowledge regarding the implications of disruption of miRNA networks in neuroinflammation and in the pathophysiology of acute and chronic CNS diseases.

MiRNA-21 silencing mediated by tumor-targeted nanoparticles combined with sunitinib: A new multimodal gene therapy approach for glioblastoma.

Malignant brain tumors, including glioblastoma (GBM), are among the most lethal human cancers, due to their tremendous invasive capacity and limited therapeutic options. Despite remarkable advances in cancer theranostics, which resulted in significant improvement of clinical outcomes, GBM relapse is very frequent and patient survival remains under one year. The elucidation of the role of abnormally-expressed miRNAs in different steps of GBM pathogenesis and in tumor resistance to therapy paved the way for the development of new miRNA-based therapeutic approaches targeting this disease, aiming at increasing specific tumor cell killing and, ultimately, cancer eradication. Here, we demonstrate that intravenously-administered chlorotoxin (CTX)-coupled (targeted) stable nucleic acid lipid particle (SNALP)-formulated anti-miR-21 oligonucleotides accumulate preferentially within brain tumors and promote efficient miR-21 silencing, which results in increased mRNA and protein levels of its target RhoB, while showing no signs of systemic immunogenicity. Decreased tumor cell proliferation and tumor size, as well as enhanced apoptosis activation and, to a lesser extent, improvement of animal survival, were also observed in GBM-bearing mice upon systemic delivery of targeted nanoparticle-formulated anti-miR-21 oligonucleotides and exposure to the tyrosine kinase inhibitor sunitinib. Overall, our results provide evidence that CTX-coupled SNALPs are a reliable and efficient system for systemic delivery of anti-miRNA oligonucleotides. Moreover, although further studies are still necessary to demonstrate a therapeutic benefit in a clinical context, our findings suggest that miRNA modulation by the targeted nanoparticles combined with anti-angiogenic chemotherapy may hold promise as an attractive approach towards GBM treatment.

Gemini surfactants mediate efficient mitochondrial gene delivery and expression.

Gene delivery targeting mitochondria has the potential to transform the therapeutic landscape of mitochondrial genetic diseases. Taking advantage of the nonuniversal genetic code used by mitochondria, a plasmid DNA construct able to be specifically expressed in these organelles was designed by including a codon, which codes for an amino acid only if read by the mitochondrial ribosomes. In the present work, gemini surfactants were shown to successfully deliver plasmid DNA to mitochondria. Gemini surfactant-based DNA complexes were taken up by cells through a variety of routes, including endocytic pathways, and showed propensity for inducing membrane destabilization under acidic conditions, thus facilitating cytoplasmic release of DNA. Furthermore, the complexes interacted extensively with lipid membrane models mimicking the composition of the mitochondrial membrane, which predicts a favored interaction of the complexes with mitochondria in the intracellular environment. This work unravels new possibilities for gene therapy toward mitochondrial diseases.