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The targeting of eEF1A1-actin complex by GT75 DNA-aptamer fight androgen-independent human prostate adenocarcinoma cells.
Full text linkIntroduction: In prostate cancer, the second most common form of solid cancer in men worldwide, castration-resistant prostate cancer (CRPC) forms are responsible for high cancer-related death. Treatments are limited and often the resistance towards the second and third lines of treatment occurs. Thus, efforts for the identification of novel therapeutic strategies/targets remain an urgent need. The eukaryotic elongation factor 1A1 (eEF1A1) can play a role in sustaining the growth of different tumors as we found that it is overexpressed in high Gleason score (7-8) tumor tissues. DNA-aptamers can recognize targets with high specificity and they have been proposed as anti-tumor-specific agents. Our group has demonstrated that a DNA-aptamer, named GT75, can target eEF1A1 in hepatocarcinoma and chronic lymphocytic leukemia cells.
Materials and methods: PC-3 cell line (resembling the CRCP phenotype) and PZHPV-7 cells (control non-tumorigenic prostate cells), were transfected with GT75 or control CT75 (Eurofins MWG) by lipofectamine 3000 (Invitrogen); cell growth was measured by MTT or MTS (Promega) at different days after transfection. Autophagy was assessed by a consecutive cell staining with neutral red (NR), and crystal violet (CV), and by an independent MTS assay (Sigma Aldrich). In Cell Western assay (ICW) was used to measure eEF1A1 protein level (mouse monoclonal EF-Tu, sc-21758, Santa Cruz Biotechnologies) and autophagy LC3B marker; cell adhesion/spreading was measured by methylene blue assay. Confocal immunofluorescence microscopy was performed with FITC-conjugated GT75 or CT75 or with fluorescent-labeled IgG anti-eEF1A1 antibody (Invitrogen).
Results and discussion: In a panel of cancer cells (LNCaP, 22RV-1, DU-145, and PC-3), a single dose of 125 nM of GT75 was able to reduce the cell growth compared to CT75 control, but the highest effect was measured in the PC-3 cells (-59%). Notably, in the non-tumorigenic PZHPV-7 cells, GT75 did not alter cell growth, demonstrating the tumor-specific effect of GT75. Besides, in PC-3 cells the GT75 reduced eEF1A1 protein levels (p<0.05). The confocal microscopy analysis in PC3 showed that GT75 targeted the eEF1A1-actin complexes bound to the cytoskeleton. On the contrary, nonspecific co-localization of eEF1A1/actin was found in non-tumorigenic PZHPV-7 cells (p<0.05). Finally, in GT75-treated PC-3 cells, a higher rate of autophagy, a lower rate of cell adhesion and spreading compared to CT75 control were observed.
Conclusion: Our data indicate the targeting of GT75 to the eEF1A1-actin complex bound to the cytoskeleton in androgen-independent prostate cancer cells. This was paralleled by the reduction of cell viability, the activation of cell autophagy, the impairment of cell adhesion and spreading. Together our observations open new perspectives for the development of targeted therapies for CRPC
Sintesi e caratterizzazione di vettori polimerici a base di una poliidrossietilaspartammide ottenuti mediante ATRP per la veicolazione di SiRNA
No full textAptameric GT oligomers need to be complexed to ethoxylated polyethylenimine as pre-paired diplex to efficently exert their cytotoxic activity in human lynphoblastic cancer cells.
No full textThe aptameric oligonucleotides GT were found to exert a selective, specific and dose-dependent cell growth inhibition effect on a varietyof human cancer cells by recognising specific nuclear proteins and among these in particular an isoform of the eukaryotic elongation factor1A1 (EEF1A1). The potential development of these aptameric oligomers needs that they retain serum and intracellular stabilities. Polycationsare safe non-viral carriers of the nucleic acids.We demonstrated that a weakly basic polycation, the ethoxylated polyethylenimine (EPEI), canefficiently deliver cytotoxic GT oligomers when they were complexed as partial pre-paired duplex. In this way, nuclease-resistance of theoligomer was markedly improved and the administration of the duplex complexed with EPEI to lymphoblastic cancer cells caused a specificcytotoxic effect at concentrations lower than that of naked GT. However, the cytotoxic activity of the oligomer-EPEI complex resulted strictlyrelated to the GC content and Tm of the duplex region. The single-stranded GT and the duplex with high GC content and Tm, althoughcomplexed with EPEI failed to exert cytotoxicity. Overall results indicated that aptameric oligomers complexed with polycations can beefficiently delivered into the cells and display the desired biological effect designing a balanced partial duplex whose stability can allowoligomer release from the polycation under the physiological cellular conditions
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Increase in therapeutic index of doxorubicin and vinblastine by aptameric oligonucleotide in human T lymphoblastic drug-sensitive and multidrug-resistant cells
No full textAptameric GT oligomers are a new class of potential anticancer molecules that inhibit the growth of human cancer cell lines by binding to specific nuclear proteins. We demonstrated that an aptameric GT oligonucleotide increased the therapeutic index of doxorubicin and vinblastine in T lymphoblastic drug-sensitive and multidrug-resistant (MDR) cells. The doxorubicin ID50 decreased 6.5-fold by coadministration of 1 μM GT to CCRF-CEM cells and by 24-fold by coadministration of 0.75 μM GT to CEM-VLB300 cells. In CEM-VLB300 cells, the vinblastine ID50 decreased 11-fold by coadministration of 0.5 μM GT. Control CT sequence did not potentiate the drugs in either CCRF-CEM or CEM-VLB300 cells. The ability of GT to bind to specific nuclear proteins in cancer cells related to the increase in the therapeutic index of doxorubicin and vinblastine. No cooperation was detected by the administration of GT oligomer together with doxorubicin to rat differentiated thyroid FRTL-5 cells and to normal human lymphocytes. These cells did not show binding of GT to the specific nuclear proteins, and they were not sensitive to the cytotoxic action of the GT sequence. Drug potentiation by GT not involving normal human lymphocytes might be exploited to develop a more selective treatment of drug-sensitive and MDR tumors
Effect of phosphorothioate modifications on the ability of GTn oligodeoxynucleotides to specifically recognize single-stranded DNA-binding proteins and to affect human cancer cellular growth
No full textSimultaneous release and ADME processes of poorly water soluble drugs: mathematical modelling
No full textThe importance of studying oral drug absorption is well recognized by both research facilities/institutions and the pharmaceutical industry. The use of mathematical models can represent a very profitable and indispensable tool to understand oral drug absorption. Indeed, mathematical models can verify the correctness of the mechanisms proposed to describe drug release, absorption, distribution and elimination thus reducing the number of expensive and time-consuming experiments. In this paper we develop a mathematical approach able to model both the polymeric particle mediated delivery and the gastrointestinal absorption−metabolism−excretion (ADME) of a given drug. As a model drug a poorly water-soluble drug (vinpocetine) in both the amorphous and nanocrystalline state is considered. The delivery system is obtained by drug cogrinding with a polymer (cross-linked polyvinilpyrrolidone). As the proposed mathematical model can properly fit the in vivo data on the basis of information obtained in vitro, it represents a powerful theoretical tool connecting in vitro and in vivo behavior
Drugs repurposing in high-grade serous ovarian cancer
No full textBackground: Ovary Carcinoma (OC) is the most lethal gynecological neoplasm due to the late diagnoses and to the common development of resistance to platinum-based chemotherapy. Thus, novel therapeutic approaches are urgently required. In this regard, the strategy of drug repurposing is becoming attractive. By this approach, the effectiveness of a drug originally developed for another indication is tested in a different pathology. The advan-tage is that data about pharmacokinetic properties and toxicity are already available. Thus, in principle, it is possible to reduce research costs and to speed up drug usage/marketing. Results: Here, some noticeable examples of repurposed drugs for OC, such as amiodarone, ruxolitinib, statins, disulfiram, ormeloxifenem, and Quinacrine, are reported. Amiodarone, an antiarrhythmic agent, has shown promising anti-OC activity, although the systemic toxicity should not be neglected. The JAK inhibitor, Ruxolitinib, may be employed particularly in co-administration with standard OC therapy as it synergistically interacts with platinum-based drugs. Particularly interesting is the use of statin which represent one of the most commonly administered drugs in aged population to treat hypercholesterolemia. Disulfiram, employed in the treatment of chronic alcoholism, has shown anti-OC properties. Ormeloxifene, commonly used for contraception, seems to be promising, especially due to the negligible side effects. Finally, Quinacrine used as an antimicrobial and anti-inflammatory drug, is able to downregu-late OC cell growth and promote cell death. Conclusion: Whereas further testing in patients are necessary to better clarify the therapeutic potential of repurposed drugs for OC, it is believed that their use, better if combined with OC targeted delivery systems, can significantly contribute to the development of novel and effective anti-OC treatments
An Overview of siRNA Delivery Strategies for Urological Cancers
Full text linkThe treatment of urological cancers has been significantly improved in recent years. However, for the advanced stages of these cancers and/or for those developing resistance, novel therapeutic options need to be developed. Among the innovative strategies, the use of small interfering RNA (siRNA) seems to be of great therapeutic interest. siRNAs are double-stranded RNA molecules which can specifically target virtually any mRNA of pathological genes. For this reason, siRNAs have a great therapeutic potential for human diseases including urological cancers. However, the fragile nature of siRNAs in the biological environment imposes the development of appropriate delivery systems to protect them. Thus, ensuring siRNA reaches its deep tissue target while maintaining structural and functional integrity represents one of the major challenges. To reach this goal, siRNA-based therapies require the development of fine, tailor-made delivery systems. Polymeric nanoparticles, lipid nanoparticles, nanobubbles and magnetic nanoparticles are among nano-delivery systems studied recently to meet this demand. In this review, after an introduction about the main features of urological tumors, we describe siRNA characteristics together with representative delivery systems developed for urology applications; the examples reported are subdivided on the basis of the different delivery materials and on the different urological cancers
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