Relevant bibliographies by topics / Oligonucleotidi antisenso

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Academic literature on the topic 'Oligonucleotidi antisenso'

Author: Grafiati
Published: 10 March 2023

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Journal articles on the topic "Oligonucleotidi antisenso"

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Goddard, Layla R., Charlotte E. Mardle, Hassan Gneid, Ciara G. Ball, Darren M. Gowers, Helen S. Atkins, Louise E. Butt, Jonathan K. Watts, Helen A. Vincent, and Anastasia J. Callaghan. "An Investigation into the Potential of Targeting Escherichia coli rne mRNA with Locked Nucleic Acid (LNA) Gapmers as an Antibacterial Strategy." Molecules 26, no. 11 (June 4, 2021): 3414. http://dx.doi.org/10.3390/molecules26113414.

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The increase in antibacterial resistance is a serious challenge for both the health and defence sectors and there is a need for both novel antibacterial targets and antibacterial strategies. RNA degradation and ribonucleases, such as the essential endoribonuclease RNase E, encoded by the rne gene, are emerging as potential antibacterial targets while antisense oligonucleotides may provide alternative antibacterial strategies. As rne mRNA has not been previously targeted using an antisense approach, we decided to explore using antisense oligonucleotides to target the translation initiation region of the Escherichia coli rne mRNA. Antisense oligonucleotides were rationally designed and were synthesised as locked nucleic acid (LNA) gapmers to enable inhibition of rne mRNA translation through two mechanisms. Either LNA gapmer binding could sterically block translation and/or LNA gapmer binding could facilitate RNase H-mediated cleavage of the rne mRNA. This may prove to be an advantage over the majority of previous antibacterial antisense oligonucleotide approaches which used oligonucleotide chemistries that restrict the mode-of-action of the antisense oligonucleotide to steric blocking of translation. Using an electrophoretic mobility shift assay, we demonstrate that the LNA gapmers bind to the translation initiation region of E. coli rne mRNA. We then use a cell-free transcription translation reporter assay to show that this binding is capable of inhibiting translation. Finally, in an in vitro RNase H cleavage assay, the LNA gapmers facilitate RNase H-mediated mRNA cleavage. Although the challenges of antisense oligonucleotide delivery remain to be addressed, overall, this work lays the foundations for the development of a novel antibacterial strategy targeting rne mRNA with antisense oligonucleotides.
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Božič, Tim, Matja Zalar, Boris Rogelj, Janez Plavec, and Primož Šket. "Structural Diversity of Sense and Antisense RNA Hexanucleotide Repeats Associated with ALS and FTLD." Molecules 25, no. 3 (January 25, 2020): 525. http://dx.doi.org/10.3390/molecules25030525.

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The hexanucleotide expansion GGGGCC located in C9orf72 gene represents the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). Since the discovery one of the non-exclusive mechanisms of expanded hexanucleotide G4C2 repeats involved in ALS and FTLD is RNA toxicity, which involves accumulation of pathological sense and antisense RNA transcripts. Formed RNA foci sequester RNA-binding proteins, causing their mislocalization and, thus, diminishing their biological function. Therefore, structures adopted by pathological RNA transcripts could have a key role in pathogenesis of ALS and FTLD. Utilizing NMR spectroscopy and complementary methods, we examined structures adopted by both guanine-rich sense and cytosine-rich antisense RNA oligonucleotides with four hexanucleotide repeats. While both oligonucleotides tend to form dimers and hairpins, the equilibrium of these structures differs with antisense oligonucleotide being more sensitive to changes in pH and sense oligonucleotide to temperature. In the presence of K+ ions, guanine-rich sense RNA oligonucleotide also adopts secondary structures called G-quadruplexes. Here, we also observed, for the first time, that antisense RNA oligonucleotide forms i-motifs under specific conditions. Moreover, simultaneous presence of sense and antisense RNA oligonucleotides promotes formation of heterodimer. Studied structural diversity of sense and antisense RNA transcripts not only further depicts the complex nature of neurodegenerative diseases but also reveals potential targets for drug design in treatment of ALS and FTLD.
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Dung, Tran Huu, Seung-Rok Lee, Suhk-Dong Han, Seon-Jeong Kim, Yeon-Mi Ju, Myong-Soo Kim, and Hoon Yoo. "Chitosan-TPP Nanoparticle as a Release System of Antisense Oligonucleotide in the Oral Environment." Journal of Nanoscience and Nanotechnology 7, no. 11 (November 1, 2007): 3695–99. http://dx.doi.org/10.1166/jnn.2007.041.

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Antisense oligonucleotide loaded chitosan nanoparticles were prepared and the release of oligonucleotide from chitosan-TPP/oligonucleotide nanoparticles was investigated. Morphological property, zeta potential and particle size of the prepared chitosan/oligonucleotide nanoparticles were investigated using Field Emission-Scanning Electron Microscope (FE-SEM) and particle size analyzer. The interaction between chitosan and oligonucleotide was confirmed by using capillary zone electrophoresis (CZE), and the released oligonucleotides were determined by spectrophotometric method. Oligonucleotides formed the complexes with chitosan with a unique morphological property. The release of oligonucleotides from nanoparticles was dependent on loading methods and pH conditions. Chitosan/oligomer-TPP nanoparticles, which was prepared by adding TPP after the formation of chitosan/oligonucleotide complex, showed the lowest release percent of oligonucleotides with 41.3% at pH 7.0 among the loading methods. The percent release of oligonucleotide from oligonucleotide loaded chitosan nanoparticle at pH 10 was higher than the one in acidic condition (pH 5.0). The released oligonucleotides from chitosan/oligonucleotide nanoparticles were stable enough for 12 h under the 20% saliva solution. Our results suggest that the sustained release of oligonucleotide from chitosan nanoparticles may be suitable for the local therapeutic application in periodontal diseases.
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CHAKRABORTY, Rila, Dalia DASGUPTA, Samit ADHYA, and Mukul K. BASU. "Cationic liposome-encapsulated antisense oligonucleotide mediates efficient killing of intracellular Leishmania." Biochemical Journal 340, no. 2 (May 25, 1999): 393–96. http://dx.doi.org/10.1042/bj3400393.

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Antisense oligonucleotides have been considered as inhibitors of growth of intracellular parasites such as Leishmania, but only limited inhibition has been observed in vitro. We have encapsulated an antisense oligonucleotide, complementary to the Leishmania universal miniexon sequence, in cationic liposomes. Low concentrations (4 μM) of encapsulated oligonucleotides specifically reduced the amastigote burden within cultured macrophages by 80%. This result illustrates the importance of effective delivery for efficient antiparasitic activity of antisense oligonucleotides.
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Zhao, Q., X. Song, T. Waldschmidt, E. Fisher, and AM Krieg. "Oligonucleotide uptake in human hematopoietic cells is increased in leukemia and is related to cellular activation." Blood 88, no. 5 (September 1, 1996): 1788–95. http://dx.doi.org/10.1182/blood.v88.5.1788.1788.

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Abstract The use of antisense oligonucleotides as tools for modulating gene expression represents a novel strategy for designing drugs to treat a variety of diseases. Several factors, including cellular uptake and internalization of the oligonucleotides, are important parameters in determining the effectiveness of antisense agents such as therapeutic drugs. We have studied oligonucleotides uptake in normal and leukemic human hematopoietic cells, such as peripheral blood, bone marrow (BM), and HL-60 cell line; and have found that, in normal human blood and BM, myeloid cells and B cells preferably took up more oligonucleotides than T cells. There was no marked difference in oligonucleotide uptake between CD4+ helper T cells and CD8+ cytolytic T cells. Leukemic cells had greater oligonucleotide uptake than their normal counterparts. Furthermore, oligonucleotide uptake was closely related to cell activation status and can be modulated by growth factors or inhibitors. These studies provide a basis for using oligonucleotides as therapeutic drugs both in vitro and in vivo.
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Zhao, Q., X. Song, T. Waldschmidt, E. Fisher, and AM Krieg. "Oligonucleotide uptake in human hematopoietic cells is increased in leukemia and is related to cellular activation." Blood 88, no. 5 (September 1, 1996): 1788–95. http://dx.doi.org/10.1182/blood.v88.5.1788.bloodjournal8851788.

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The use of antisense oligonucleotides as tools for modulating gene expression represents a novel strategy for designing drugs to treat a variety of diseases. Several factors, including cellular uptake and internalization of the oligonucleotides, are important parameters in determining the effectiveness of antisense agents such as therapeutic drugs. We have studied oligonucleotides uptake in normal and leukemic human hematopoietic cells, such as peripheral blood, bone marrow (BM), and HL-60 cell line; and have found that, in normal human blood and BM, myeloid cells and B cells preferably took up more oligonucleotides than T cells. There was no marked difference in oligonucleotide uptake between CD4+ helper T cells and CD8+ cytolytic T cells. Leukemic cells had greater oligonucleotide uptake than their normal counterparts. Furthermore, oligonucleotide uptake was closely related to cell activation status and can be modulated by growth factors or inhibitors. These studies provide a basis for using oligonucleotides as therapeutic drugs both in vitro and in vivo.
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Studzińska, Sylwia, Ewelina Zawadzka, Szymon Bocian, and Michał Szumski. "Synthesis and application of stationary phase for DNA-affinity chromatographic analysis of unmodified and antisense oligonucleotide." Analytical and Bioanalytical Chemistry 413, no. 20 (June 24, 2021): 5109–19. http://dx.doi.org/10.1007/s00216-021-03473-7.

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AbstractThe goal of the research was the synthesis and application of an oligonucleotide immobilized stationary phase for the analysis of unmodified and antisense oligonucleotides. The method for attaching these molecules to aminopropyl silica modified with pentanedioic acid was developed. Each step of the synthesis was carefully controlled with the application of spectroscopic, elemental, and chromatographic analyses. The oligonucleotide-based stationary phase was applied for the retention studies. Unmodified oligonucleotides of different complementarity to the molecule attached as a stationary phase, as well as antisense oligonucleotides, were tested. The comparative study upon complex optimization of oligonucleotide analysis in different liquid chromatography modes was performed. Results have shown that this stationary phase may be applied for oligonucleotide analysis in hydrophilic interaction liquid chromatography and ion exchange chromatography, but no unique sequence-based selectivity was obtained. Contrary results were observed for affinity chromatography, which allowed for specific separation of the complementary strands based on hydrogen bonding and stacking interactions, where the temperature was the main factor influencing the selectivity of the separation. Furthermore, the oligonucleotide-based stationary phase may be applied for comparative antisense oligonucleotide hybridization studies to a specific RNA sequence. All of the results have shown that affinity chromatography with oligonucleotide-based stationary phases is a powerful technique for the specific base recognition of polynucleotides. Graphical abstract
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Pandiri, Kavya, Mohammed Abdul Samad, Nadeem Abbas Gulamus, and Hajera Khanam. "Medicinal Applications of Antisense Oligonucleotides: A Review." INTERNATIONAL JOURNAL OF APPLIED PHARMACEUTICAL SCIENCES AND RESEARCH 5, no. 02 (April 1, 2020): 30–36. http://dx.doi.org/10.21477/ijapsr.5.2.2.

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Antisense technology has emerged as a fast and conceivably high-throughput method for repressing genes due to advancement in knowledge about DNA and RNA physiology. The limitations of antisense oligonucleotide therapy in delivery strategies have been overcome in recent years. Antisense oligonucleotide treatment was effectively applied towards targeting a wide range of therapeutic areas. With ongoing approvals of antisense oligonucleotides, there is an expanding enthusiasm for increasing the utilization of these compounds for curing various infections. This short survey gives a far-reaching comprehension of applications of antisense technology, how they can be utilized therapeutically, and current endeavors to grow new antisense oligonucleotide treatments that will add a forthcoming therapeutic approach for the treatment of various diseases.
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Yamamoto, T., R. P. Moerschell, L. P. Wakem, S. Komar-Panicucci, and F. Sherman. "Strand-specificity in the transformation of yeast with synthetic oligonucleotides." Genetics 131, no. 4 (August 1, 1992): 811–19. http://dx.doi.org/10.1093/genetics/131.4.811.

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Abstract Cyc1 mutants of the yeast Saccharomyces cerevisiae were directly transformed with both sense and antisense oligonucleotides to examine the involvement of the two genomic DNA strands in transformation. Sense oligonucleotides yielded approximately 20-fold more transformants than antisense oligonucleotides. This differential effect was observed with oligonucleotides designed to make alterations at six different sites along the gene and was independent of the oligonucleotide sequence and length, number of mismatches and the host strain. Competition studies showed that antisense oligonucleotides did not inhibit transformation. Although the mechanism for this strand specificity is unknown, this difference was maintained even when CYC1 transcription was diminished to approximately 2% of the normal level.
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Tanaka, T., C. C. Chu, and W. E. Paul. "An antisense oligonucleotide complementary to a sequence in I gamma 2b increases gamma 2b germline transcripts, stimulates B cell DNA synthesis, and inhibits immunoglobulin secretion." Journal of Experimental Medicine 175, no. 2 (February 1, 1992): 597–607. http://dx.doi.org/10.1084/jem.175.2.597.

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An antisense phosphorothioate (S)-oligonucleotide to a sequence in the intervening (I) region of the gamma 2b immunoglobulin (Ig) heavy chain gene inhibits Ig secretion by B cells stimulated with lipopolysaccharide (LPS) or LPS plus interleukin 4. It is also a striking stimulant of DNA synthesis by resting B cells. The antisense S-oligonucleotide causes a 10-20-fold increase in the expression of the gamma 2b germline transcript. Among mutants of the antisense S-oligonucleotide, some show all the effects whereas others are inactive. A similar hierarchy exists in the quantitative biological activities of mutant S-oligonucleotides and in their capacity to hybridize to the sense oligonucleotide, strongly suggesting that an I gamma 2b sequence in the RNA transcript or in the noncoding strand of the DNA is the target of the antisense S-oligonucleotide. The possible relationship of the overexpression of the germline gamma 2b transcript to the biological functions of the I gamma 2b antisense S-oligonucleotide is discussed.
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Dissertations / Theses on the topic "Oligonucleotidi antisenso"

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Lombardini, Lorenza <1982&gt. "Valutazione preclinica di oligonucleotidi antisenso come nuovo approccio terapeutico specifico per le leucemie acute con riarrangiamenti di MLL." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/3238/1/Lombardini_Lorenza_Tesi.pdf.

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Le Leucemie Acute Mieloidi di sottotipo FAB M4 e M5, le Leucemie Acute Linfoblastiche e le Leucemie Bifenotipiche sono frequentemente caratterizzate da traslocazioni del gene 11q23/MLL con formazione di oncogeni di fusione e produzione di oncoproteine che inducono la trasformazione neoplastica. Tali leucemie con riarrangiamenti di 11q23/MLL sono caratterizzate da prognosi infausta e scarsa responsività alle terapie convenzionali. Data la necessità di trovare terapie efficaci per le leucemie con traslocazione di MLL, in questo lavoro di ricerca sono stati progettati, caratterizzati e validati siRNA per il silenziamento genico degli oncogeni di fusione di MLL, con lo scopo di valutare il ripristino delle normali funzionalità di differenziamento cellulare e l’arresto della proliferazione neoplastica. Sono stati progettati siRNA specifici per gli oncogeni di fusione di MLL, sia per le regioni conservate nei diversi oncogeni di fusione, sia a livello del punto di fusione (breakpoint), sia per le regioni sui geni partner. I siRNA sono stati valutati su linee cellulari contenenti diverse traslocazioni del gene MLL. Il silenziamento è stato valutato sia a livello cellulare in termini di riduzione della capacità proliferativa e del numero delle cellule leucemiche, sia a livello molecolare tramite l’analisi della diminuzione dell’mRNA degli oncogeni di fusione di MLL. E’ stata valutata la diminuzione delle oncoproteine di fusione di MLL in seguito a trattamento con siRNA. E’ stata analizzata la variazione dell’espressione di geni dipendenti da MLL in seguito a trattamento con siRNA. Sono stati messi a punto modelli murini bioluminescenti di leucemie acute con traslocazioni di MLL innanzitutto per studiare il trafficking in vivo e la progressione leucemica delle leucemie acute con traslocazione di MLL. Successivamente sono stati utilizzati i modelli murini per lo studio in vivo dell’efficienza e della tossicità dei siRNA progettati e validati in vitro, valutando diversi sistemi di delivery per i siRNA in vivo.
Chromosomal translocations involving 11q23/MLL gene represent frequent abnormalities in Acute Myeloid Leukemias (AML) subtype FAB M5 and M4, Acute Lymphoblastic Leukemias (ALL) and Biphenotypic Leukemias. MLL-related leukemias are generally associated with aggressive disease and poor prognosis. MLL-related leukemias need validation of new therapies, so we designed and validated anti-MLL siRNA to downregulate MLL-fusion oncogenes. We tested anti-MLL siRNA in MLL-carrying acute leukemias cell lines, and we evaluated both MLL mRNA downregulation, cellular viability and proliferation, and protein inhibition. We also evaluated gene expression of MLL-dependent genes. We generated bioluminescent acute leukemias xenograft mouse models of the most frequent MLL fusion genes. We evaluated leukemias trafficking and progression, and we evaluated delivery methods for siRNA delivery in vivo.
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Lombardini, Lorenza <1982&gt. "Valutazione preclinica di oligonucleotidi antisenso come nuovo approccio terapeutico specifico per le leucemie acute con riarrangiamenti di MLL." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/3238/.

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Le Leucemie Acute Mieloidi di sottotipo FAB M4 e M5, le Leucemie Acute Linfoblastiche e le Leucemie Bifenotipiche sono frequentemente caratterizzate da traslocazioni del gene 11q23/MLL con formazione di oncogeni di fusione e produzione di oncoproteine che inducono la trasformazione neoplastica. Tali leucemie con riarrangiamenti di 11q23/MLL sono caratterizzate da prognosi infausta e scarsa responsività alle terapie convenzionali. Data la necessità di trovare terapie efficaci per le leucemie con traslocazione di MLL, in questo lavoro di ricerca sono stati progettati, caratterizzati e validati siRNA per il silenziamento genico degli oncogeni di fusione di MLL, con lo scopo di valutare il ripristino delle normali funzionalità di differenziamento cellulare e l’arresto della proliferazione neoplastica. Sono stati progettati siRNA specifici per gli oncogeni di fusione di MLL, sia per le regioni conservate nei diversi oncogeni di fusione, sia a livello del punto di fusione (breakpoint), sia per le regioni sui geni partner. I siRNA sono stati valutati su linee cellulari contenenti diverse traslocazioni del gene MLL. Il silenziamento è stato valutato sia a livello cellulare in termini di riduzione della capacità proliferativa e del numero delle cellule leucemiche, sia a livello molecolare tramite l’analisi della diminuzione dell’mRNA degli oncogeni di fusione di MLL. E’ stata valutata la diminuzione delle oncoproteine di fusione di MLL in seguito a trattamento con siRNA. E’ stata analizzata la variazione dell’espressione di geni dipendenti da MLL in seguito a trattamento con siRNA. Sono stati messi a punto modelli murini bioluminescenti di leucemie acute con traslocazioni di MLL innanzitutto per studiare il trafficking in vivo e la progressione leucemica delle leucemie acute con traslocazione di MLL. Successivamente sono stati utilizzati i modelli murini per lo studio in vivo dell’efficienza e della tossicità dei siRNA progettati e validati in vitro, valutando diversi sistemi di delivery per i siRNA in vivo.
Chromosomal translocations involving 11q23/MLL gene represent frequent abnormalities in Acute Myeloid Leukemias (AML) subtype FAB M5 and M4, Acute Lymphoblastic Leukemias (ALL) and Biphenotypic Leukemias. MLL-related leukemias are generally associated with aggressive disease and poor prognosis. MLL-related leukemias need validation of new therapies, so we designed and validated anti-MLL siRNA to downregulate MLL-fusion oncogenes. We tested anti-MLL siRNA in MLL-carrying acute leukemias cell lines, and we evaluated both MLL mRNA downregulation, cellular viability and proliferation, and protein inhibition. We also evaluated gene expression of MLL-dependent genes. We generated bioluminescent acute leukemias xenograft mouse models of the most frequent MLL fusion genes. We evaluated leukemias trafficking and progression, and we evaluated delivery methods for siRNA delivery in vivo.
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Albouz, Soulaf. "Evaluation de l’efficacité de l’atovaquone encapsulée associée à des oligonucléotides antisens anti-ARNm de topoisomérase II chez Plasmodium falciparum." Thesis, Université Paris-Saclay (ComUE), 2017. http://www.theses.fr/2017SACLS079.

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Selon les estimations de l'OMS, le bilan mondial du paludisme a atteint 212 millions de cas et 429 000 décèsen 2015 (OMS, 2016). Cette gravité est principalement due à Plasmodium falciparum. A l’heure actuelle, P. falciparum présente des résistances à tous les antipaludiques donnés en monothérapie.Par conséquent, pour réduire le risque d’échec thérapeutique, l'OMS a recommandé depuis 2001 l’utilisation de bithérapie, notamment d'Artemisinin Combination Therapy (ACT), comme traitement de première intention.Les ACT sont composés essentiellement d’un dérivé d’artéminisine, à demi-vie courte et un autre antipaludique à demi-vie longue, connu en monothérapie.Le parasite a également montré des signes de résistance aux ACT, principalement en Asie du Sud-est, menaçant les programmes d’éradications contre le paludisme.La découverte de nouveaux composés à activité antipaludique ou de nouvelles procédures de traitement sont urgentes.La valorisation d’anciennes molécules est également au cœur des études afin d’améliorer notamment leur biodisponibilité et réverser les mécanismes de résistance du parasite. Ainsi, des études prouvent l’intérêt de l’utilisation de nanotechnologies pour l’amélioration de l’efficacité d’antipaludiques. L’atovaquone en est un exemple, cette modification a notamment permis d’améliorer sa biodisponibilité. Notre étude a porté sur une de ces formulations, de l’atovaquone encapsulée dans une nanoémulsion cationique (NE) appelée ATQ. Une deuxième génération a ensuite été testée par association d’oligonucléotides antisens anti-ARNm de topoisomérase II(AST) de P. falciparum. En effet, des stratégies antisens thérapeutiques font leur preuve en santé humaine et présentent un intérêt croissant en parasitologie. Les NE/AST ont montré une activité anti-palustre spécifique contre P. falciparum in vitro. Leur spécificité a permis d’aboutir à l’arrêt du cycle cellulaire et une forte diminution du taux d’ARNm de la topoisomérase II. Ce phénomène a montré être dépendant de l’action de la RNase H. Un effet synergique de ces NE/AST a également été montré en association avec la chloroquine, l’atovaquone et la dihydroartémisinine sur une souche sensible de P. falciparum et des souches résistantes aux antipaludiques précédemment cités.L’ATQ a également montré une forte efficacité sur une souche résistante à l’atovaquone d’un facteur 5. En présence d’ATQ, la mitochondrie a rapidement été altérée conduisant à une mort précoce du parasite. Un traitement à l’ATQ a abouti à la guérison de souris Swiss infectée par P. berghei après deux injections en i.v. en 5 jours. Enfin, l’ATQ/AST a montré une efficacité in vitro contre P. falciparum et P. berghei in vivo. Un test de cytoadhérance des hématies parasitées a des cellules endothéliales a révélé un fort pourvoir d’inhibition de la cytoadhérance de l’ATQ/AST.Un résultat prometteur dans le cadre de traitement du neuropaludisme
According to the estimations of theWHO, in 2015, 212million cases ofmalariahave been reported(WHO,2016). These figuresmakemalariathe most deadlyparasitic diseasein the world, with429.000deaths per year. Some treatments against Plasmodium falciparum exist. However, no really good treatment option can be found in monotherapy due to the resistance emergency. Therefore To reduce the risk of resistance, WHO has recommended since 2001 combination therapies, which is basically an Artemisinin Combined Therapy (ACT), as first-line treatment. The main problem of commercialized bi-therapy is that they are composed of two molecules with individual resistance which leaded to the emergence of resistance to the latest ACTs such as a dihydroartemisinin /piperaquine combinationmainly in South-East Asia.Thus the use of new therapeutic combination strategy that can bypass the parasites' mechanisms of resistance is urgent to effectively treat malaria. As the pathway from drug discovery to drug commercialization is both long and very expensive, it is essential to develop ways to improve existing antimalarial treatments. In the first place it’s necessary to find a new antimalarial formulation based on an already commercialized drug to modify its biodisponibility and its mechanism of action in order to revert the resistance. In the second place its necessary to associate this formulation with a novel none commercialized antimalarial strategy such as the antisens oligonucleotides already usedinhumanhealth. In our lab we have developed nanoemulsions containing atovaquone and antisense oligonucleotides anti topoisomerase II against P. falciparum.Nanoemulsionsvectoringantisens oligonucleotidesandused againstP. falciparum topoisomerase II(NE/AST) showed encouraging anti-parasite killing results.Additionalresultshave showna synergistic in vitro effectwithantimalarial drugs(chloroquine, dihydroartemisinin and atovaquone) in sensitive and resistances strains. Moreover NE/ASTrestricted Topoisomerase II gene expression and blocked the cell cycle in G2/M phase leading to parasite’s death by mitophagy.As Drug delivery systemscan improve the efficacy ofcommon antimalarial drugs by delivering the drug to its target, while protecting it from degradation in biological environment and increasing its biodisponibility, our nanoemulsions containing atovaquone (ATQ) leaded to reversion of atovaquone resistance with 5 fold decrease in its IC50. Observations made with confocal microscopy have shown mitochondrial alteration after ATQ treatment.Our novel and original bi-therapy is focused on the association ofATQ with NE/AST (ATQ/AST).We obtained an IC50 8-fold lower than atovaquone’s IC50with total inhibition of parasites’ capacity to reinfect new red blood cells. A cytoadherence test of parasitized erythrocytes to endothelial cells revealed a strong capacity of cytoadherence inhibition of ATQ / AST, a promising result in the treatment of cerebral malaria
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Yannopoulos, Constantin G. "Synthesis and targeting of antisense oligonucleotides." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/tape16/PQDD_0014/NQ37032.pdf.

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Lin, Zhaoru. "Characterisation of antisense oligonucleotide-stimulated ribosomal frameshifting." Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609380.

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GODARD, GERARD. "Oligonucleotides antisens modifies : potentialites et limites." Paris 6, 1994. http://www.theses.fr/1994PA066374.

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La stabilite des oligonucleotides antisens vis-a-vis des nucleases, et leur penetration dans les cellules sont deux limitations majeures a leur efficacite. De nombreuses modifications chimiques ont ete realisees pour rendre les oligonucleotides plus stables vis-a-vis des nucleases. Les oligonucleotides font partie de ces molecules chimiquement modifiees. De tels oligonucleotides sont incapables de former des hybrides avec l'arn qui soient substrats de la ribonuclease h, enzyme vraisemblablement implique dans l'activite antisens des oligonucleotides. L'activite d'oligonucleotides ou b couples a des molecules reactives (1,10 phenanthroline ou psoralene) dans le but de pallier a cette deficience, a ete etudiee sur des substrats adn ou arn. La plus faible reactivite de ces molecules observee sur l'arn reflete probablement, au moins en partie, la difference de structure arn/adn et la difference de stabilite des hybrides oligo/arn-oligo/adn. Des oligonucleotides resistants aux nucleases (oligonucleotides contenant des 2,4-dideoxy-b-d-erythro-hexopyranosyls, ou couples a la lithocholamide) ont montre des proprietes interessantes. Il se trouve que certain de ces oligonucleotides peuvent induire une activite de coupure de l'arn par la rnase h plus importante qu'un oligonucleotide non modifie. Un des moyens mis en uvre afin d'ameliorer la penetration des oligonucleotides dans les cellules est de les associer a des transporteurs. L'association d'oligonucleotides a des nanoparticules de polyisohexylcyanoacrylate peut etre realise par l'intermediaire d'un cation hydrophobe, tel que le ctab. Neanmoins, ce cation pose des problemes de toxicite. Nous avons pu associer directement des oligonucleotides couples au cholesterol aux nanoparticules via la molecule de cholesterol et ainsi nous affranchir de la presence du ctab. Ces oligonucleotides, associes au ctab, ont une activite biologique specifique a des concentrations de l'ordre de 200 a 400 nm
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Rouleau, Samuel. "Oligonucléotides comme modulateurs de l'expression génique." Thèse, Université de Sherbrooke, 2017. http://hdl.handle.net/11143/11570.

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L’ARN est sans aucun doute la molécule biologique la plus versatile qui soit. Tout comme l’ADN, il peut contenir et transmettre de l’information génétique. Tout comme les protéines, il peut accomplir une multitude de fonctions biologiques. De plus, son rôle le plus connu demeure celui d’intermédiaire entre l’ADN et les protéines. L’ARN est donc au cœur d’un bon nombre de processus biologiques. Ceci lui confère un immense potentiel thérapeutique qui jusqu’à présent demeure largement inexploité. Pour accomplir ses fonctions, l’ARN doit adopter une structure tridimensionnelle précise qui est dépendante à la fois de sa séquence et de son environnement. Ainsi, en modifiant la structure d’un ARN, il est possible d’en moduler sa fonction. C’est l’objectif global des travaux présentés dans cette thèse. Pour y parvenir, de courts oligonucléotides antisens (OA) ont été utilisés. Cette stratégie revêt plusieurs avantages. Comme les OA s’apparient à leur cible en formant des paires de bases Watson-Crick, ils offrent une grande spécificité et leur design est facile. De plus, en se fiant aux données structurales et aux logiciels de prédictions de structures des ARN, on peut aisément identifier les régions à cibler avec les OA. Enfin, cette technique est versatile puisqu’on peut cibler différents motifs d’ARN. La première cible a été le ribozyme du virus de l’hépatite D. Cet ARN, qui catalyse une réaction d’auto-coupure, a été modifié afin que son activité devienne dépendante à la liaison d’OA. Plusieurs modules ont ainsi été créés et combinés afin d’obtenir des ribozymes qui répondaient à la présence d’un ou plusieurs OA. En insérant ces interrupteurs moléculaires dans les régions non traduites d’un ARNm, nous avons ainsi modulé l’expression de ce gène avec les OA. Cet outil a des applications intéressantes pour la régulation de gènes en biologie synthétique. Un autre motif ciblé a été le G-quadruplex (G4). Cette structure non canonique exerce de nombreuses fonctions biologiques et représente donc une cible thérapeutique intéressante. Lorsque présent dans la région 5’ non traduite d’un ARNm, le G4 mène généralement à une diminution de la traduction. En utilisant des OA qui empêchent la formation du G4, nous avons été en mesure d’augmenter la traduction du gène ciblé. De plus, il a été possible de développer des OA qui favorisent la formation d’un G4 dans le but de diminuer l’expression de la cible. Finalement, dans le dernier chapitre de cette thèse, il est démontré que les G4 présents dans les microARN primaires influencent leur maturation en microARN matures. Des OA ciblant ces G4 ont été utilisés afin de favoriser la maturation de microARN suppresseurs de tumeurs, ce qui présente un potentiel thérapeutique intéressant. En bref, les travaux présentés dans cette thèse démontrent clairement que les OA sont un outil de choix pour cibler et modifier la structure de motifs d’ARN spécifiques.
Abstract : RNA is a versatile biological molecule. Like DNA, it can contain and transmit genetic information. Like proteins, it can accomplish multiple biological functions. Also, its most known role remains that of intermediary between DNA and proteins. RNA is thus a key player in many biological processes. This gives it an immense therapeutic potential which remains largely untapped. To fulfill its functions, RNA must adopt a precise threedimensional structure that is dependent on both its sequence and its environment. Thus, by modifying the structure of an RNA, it is possible to modulate its function. This is the overall objective of the work presented in this thesis. To achieve this, small antisense oligonucleotides (ASO) have been used. This strategy has several advantages. As ASO bind their target with Watson-Crick base pairs, they offer great specificity and their design is easy. Moreover, reliance on structural data and RNA structure prediction softwares makes it easy to identify the regions to be targeted with ASO. Finally, this technique is versatile since it is possible to target different RNA motifs. The first target was the HDV self-cleaving motif. This RNA, which catalyzes a self-cleaving reaction, has been modified so that its activity became dependent on the binding of ASO. Several modules were thus created and combined in order to obtain ribozymes which responded to the presence of one or more ASO. By inserting these molecular switches into an mRNA’s UTR, the expression of this gene was modulated with the ASO. This has interesting applications for the regulation of genes in synthetic biology. Another target motif was the G-quadruplex (G4). This non-canonical structure exerts many biological functions and therefore represents an interesting therapeutic target. When present in the mRNA’s 5’UTR, G4 generally lead to a decrease in translation. Using ASO that prevent G4 formation, we were able to increase the translation of the target gene. In addition, it has been possible to develop ASO which promote the formation of a G4 in order to decrease the expression of the target. Finally, in the last chapter of this thesis, it is demonstrated that the G4 present in the primary microRNAs influence their maturation in mature microRNAs. ASO targeting these G4 have been used in order to promote the maturation of tumor suppressor microRNAs, which has an interesting therapeutic potential. The work presented in this thesis clearly demonstrates that ASO are ideal for targeting and altering the structure of specific RNA motifs.
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Chalk, Alistair. "Computational prediction of antisense oligonucleotides and siRNAs /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-376-0/.

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Gill, Taylor Elizabeth. "Selective targeting of MYC by antisense oligonucleotides." Thesis, Massachusetts Institute of Technology, 2018. http://hdl.handle.net/1721.1/115602.

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Thesis: Ph. D. in Biomedical Engineering, Harvard-MIT Program in Health Sciences and Technology, 2018.
This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references (pages 181-208).
MYC is one of the most commonly dysregulated genes across all cancers. As a master transcription factor with greater than 10,000 binding sites throughout the genome, the MYC oncoprotein coordinates a transcriptional regulatory network consisting of approximately 15% of all genes, controlling cancer hallmark expression programs responsible for cellular proliferation, growth, metabolism, and evasion from apoptosis. MYC dysregulation occurs genetically, epigenetically, and post-transcriptionally through a wide variety of mechanisms. Despite its well-characterized properties as a proto-oncogene, direct potent and selective inhibition of MYC remains a significant challenge. Models of systemic MYC inhibition utilizing inducible genetic constructs in mice have revealed that inhibition of MYC activity leads to potent tumor regression with an evident therapeutic window, suggesting that pharmacologic MYC inhibition may be a viable cancer therapeutic strategy. Small molecule inhibitors designed to block MYC protein activity exhibit low potency, display poor selectivity, and lack antitumor efficacy, which has led MYC to be historically classified as 'undruggable.' Efforts aimed at indirectly targeting MYC transcription often lead to development of resistance characterized by reinforced expression of MYC. Clearly, alternate strategies are needed to achieve selective and potent inhibition of MYC. The goals of this research were to develop antisense oligonucleotides specifically targeted against the MYC mRNA to achieve potent inhibition of MYC translation, and to characterize the activity of these molecules as specific modulators of MYC expression and as prototypical MYC-directed therapeutics. We designed and synthesized a library of MYC-targeting antisense oligonucleotides (MYCASOs) containing several chemical synthetic features to increase target affinity and stability. Treatment of MYC-expressing cancer cells with MYCASOs leads to RNase H-mediated cleavage of MYC mRNA and a potent decrease in MYC protein levels. MYC knockdown is accompanied by significant effects on cellular viability and inhibition of cellular proliferation. Furthermore, MYCASO treatment specifically perturbs MYC-driven gene expression signatures. In a MYC-induced murine model of hepatocellular carcinoma, MYCASO treatment leads to cleavage of the MYC transcript, decreased MYC protein levels within tumors, and reduced tumor burden. MYCASOs represent a new chemical tool for in vitro and in vivo modulation of MYC activity, and promising therapeutic agents for MYC-addicted tumors.
by Taylor Elizabeth Gill.
Ph. D. in Biomedical Engineering
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Åström, Hans. "Studies on phosphate ester cleavage and development of oligonucleotide based artificial nucleases (OBAN's) /." Stockholm, 2004. http://diss.kib.ki.se/2004/91-7349-935-8/.

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Books on the topic "Oligonucleotidi antisenso"

1

A, Stein Cy, and Krieg Arthur M, eds. Applied antisense oligonucleotide technology. New York: Wiley-Liss, 1998.

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1956-, Sanghvi Yogesh S., Cook P. Dan 1944-, American Chemical Society. Division of Carbohydrate Chemistry., and American Chemical Society Meeting, eds. Carbohydrate modifications in antisense research. Washington, DC: American Chemical Society, 1994.

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Saghir, Akhtar, ed. Delivery strategies for antisense oligonucleotide therapeutics. Boca Raton: CRC Press, 1995.

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Thomas, Tuschl, Rossi John J, and New York Academy of Sciences, eds. Oligonucleotide therapeutics. Boston, Mass: Blackwell on behalf of the New York Academy of Sciences, 2006.

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J, Rossi John, Gait, M. J. (Michael J.), Eckstein Fritz 1932-, and New York Academy of Sciences, eds. Oligonucleotide therapeutics: Fourth annual meeting. Boston, Mass: Published by Blackwell Pub. on behalf of the New York Academy of Sciences, 2009.

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Sudhir, Agrawal, ed. Antisense therapeutics. Totowa, N.J: Humana Press, 1996.

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Ugent, Steven Jay. Antisense oligonucleotides: Psoralen photoreactivity and enzymatic resistance. [S.l: s.n.], 1991.

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Crooke, Stanley T. Therapeutic applications of oligonucleotides. New York: Springer-Verlag, 1995.

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Ian, Phillips M., ed. Antisense therapeutics. 2nd ed. Totowa, N.J: Humana Press, 2005.

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Ian, Phillips M., ed. Antisense therapeutics. 2nd ed. Totowa, N.J: Humana Press, 2005.

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Book chapters on the topic "Oligonucleotidi antisenso"

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Bremer, Jeroen, and Peter C. van den Akker. "In Vivo Models for the Evaluation of Antisense Oligonucleotides in Skin." In Methods in Molecular Biology, 315–20. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2010-6_21.

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AbstractHere, we describe an in vivo model in which antisense oligonucleotides were preclinically evaluated in reconstituted patient and healthy control skin. The aim was to investigate the effect of antisense oligonucleotides upon local or systemic administration. This allows for clinically relevant evaluation of antisense oligonucleotides in an in vivo setting. In this model, primary human keratinocytes and fibroblasts were placed into silicone grafting chambers, implanted onto the back of athymic nude mice. After sufficient cells were expanded, within a few weeks, human skin grafts were generated with a high success rate. These mice bearing grafts were subsequently treated with antisense oligonucleotides targeting exon 105 of the COL7A1 gene which encodes type VII collagen. Patients completely lacking expression of type VII collagen develop severe blistering of skin and mucosa, i.e., recessive dystrophic epidermolysis bullosa. In this chapter, we describe the in vivo model used for the preclinical evaluation of antisense oligonucleotides as therapeutic approach for recessive dystrophic epidermolysis bullosa.
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Scherrmann, Jean-Michel, Kim Wolff, Christine A. Franco, Marc N. Potenza, Tayfun Uzbay, Lisiane Bizarro, David C. S. Roberts, et al. "Antisense Oligonucleotides." In Encyclopedia of Psychopharmacology, 122–27. Berlin, Heidelberg: Springer Berlin Heidelberg, 2010. http://dx.doi.org/10.1007/978-3-540-68706-1_384.

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Neumann, Inga D., and Peter J. Flor. "Antisense Oligonucleotides." In Encyclopedia of Psychopharmacology, 162–67. Berlin, Heidelberg: Springer Berlin Heidelberg, 2015. http://dx.doi.org/10.1007/978-3-642-36172-2_384.

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Crooke, Stanley T. "Antisense Oligonucleotides." In Cancer Therapeutics, 299–336. Totowa, NJ: Humana Press, 1997. http://dx.doi.org/10.1007/978-1-59259-717-8_15.

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Shah, Neel Jayesh. "Antisense Oligonucleotides." In Introduction to Basics of Pharmacology and Toxicology, 407–11. Singapore: Springer Singapore, 2019. http://dx.doi.org/10.1007/978-981-32-9779-1_33.

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Neumann, Inga D., and Peter J. Flor. "Antisense Oligonucleotides." In Encyclopedia of Psychopharmacology, 1–7. Berlin, Heidelberg: Springer Berlin Heidelberg, 2013. http://dx.doi.org/10.1007/978-3-642-27772-6_384-2.

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López-Martínez, Andrea, Patricia Soblechero-Martín, and Virginia Arechavala-Gomeza. "Evaluation of Exon Skipping and Dystrophin Restoration in In Vitro Models of Duchenne Muscular Dystrophy." In Methods in Molecular Biology, 217–33. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2010-6_14.

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AbstractSeveral exon skipping antisense oligonucleotides (eteplirsen, golodirsen, viltolarsen, and casimersen) have been approved for the treatment of Duchenne muscular dystrophy, but many more are in development targeting an array of different DMD exons. Preclinical screening of the new oligonucleotide sequences is routinely performed using patient-derived cell cultures, and evaluation of their efficacy may be performed at RNA and/or protein level. While several methods to assess exon skipping and dystrophin expression in cell culture have been developed, the choice of methodology often depends on the availability of specific research equipment.In this chapter, we describe and indicate the relevant bibliography of all the methods that may be used in this evaluation and describe in detail the protocols routinely followed at our institution, one to evaluate the efficacy of skipping at RNA level (nested PCR) and the other the restoration of protein expression (myoblot), which provide good results using equipment largely available to most research laboratories.
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Nyce, Jonathan W. "Respirable Antisense Oligonucleotides." In New Drugs for Asthma, Allergy and COPD, 361–64. Basel: KARGER, 2001. http://dx.doi.org/10.1159/000062198.

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Zhou, Haiyan. "Design of Bifunctional Antisense Oligonucleotides for Exon Inclusion." In Methods in Molecular Biology, 53–62. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2010-6_3.

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AbstractBifunctional antisense oligonucleotide (AON) is a specially designed AON to regulate pre-messenger RNA (pre-mRNA) splicing of a target gene. It is composed of two domains. The antisense domain contains sequences complementary to the target gene. The tail domain includes RNA sequences that recruit RNA binding proteins which may act positively or negatively in pre-mRNA splicing. This approach can be designed as targeted oligonucleotide enhancers of splicing, named TOES, for exon inclusion; or as targeted oligonucleotide silencers of splicing, named TOSS, for exon skipping. Here, we provide detailed methods for the design of TOES for exon inclusion, using SMN2 exon 7 splicing as an example. A number of annealing sites and the tail sequences previously published are listed. We also present methodology of assessing the effects of TOES on exon inclusion in fibroblasts cultured from a SMA patient. The effects of TOES on SMN2 exon 7 splicing were validated at RNA level by PCR and quantitative real-time PCR, and at protein level by western blotting.
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Krieg, A. M. "Immune Stimulation by Oligonucleotides." In Antisense Research and Application, 243–62. Berlin, Heidelberg: Springer Berlin Heidelberg, 1998. http://dx.doi.org/10.1007/978-3-642-58785-6_8.

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Conference papers on the topic "Oligonucleotidi antisenso"

1

Brown, Paige K., Ammar T. Qureshi, Daniel J. Hayes, and W. Todd Monroe. "Targeted Gene Silencing With Light and a Silver Nanoparticle Antisense Delivery System." In ASME 2011 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2011. http://dx.doi.org/10.1115/sbc2011-53647.

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Targeted delivery and controlled release of oligonucleotide therapeutics in vivo are essential aspects of an ideal delivery vehicle. Here we demonstrate the synthesis and in vitro/intracellular characterization of silver nanoparticle (SNP) photolabile nucleic acid conjugates, with the aim of developing a nanoparticulate platform for inducible gene silencing. Due to unique size related properties, nanostructures are being increasingly utilized for intracellular diagnostics and delivery applications. While most nanoscale delivery platforms are polymeric in composition, studies of metallic nanoparticles have highlighted their suitability for delivery of therapeutic agents such as antisense oligonucleotides [1]. The potential benefits of noble metal nanoparticles in delivery applications include tunable size and shape, ease of bulk synthesis and functionalization via ‘wet chemistry’ techniques, and enhanced stability of tethered DNA [2]. Silver is one of the best surface-enhancing substrates available for nanostructure synthesis [3]. SNP composites afford external control over surface-tethered drug release via external triggers.
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Puzanova, E. V. "Prospects for the use of unmodified antisense oligonucleotides in the regulation of the synthesis of secondary metabolites of essential oil plants." In 2nd International Scientific Conference "Plants and Microbes: the Future of Biotechnology". PLAMIC2020 Organizing committee, 2020. http://dx.doi.org/10.28983/plamic2020.202.

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A scheme and basic principles for the use of unmodified antisense oligonucleotides in the agricultural sector as a regulator of the synthesis of secondary metabolites of essential oil plants are developed.
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Thomas, Sufi M., and Danith Ly. "Abstract 3301: Guanidinium antisense oligonucleotides for cancer therapy." In Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1538-7445.am2013-3301.

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Van Aerschot, Arthur, Mark Vandermeeren, Johan Geysen, Walter Luyten, Marc Miller, David Atkins, Sonia Preveral, Ester Saison-Behmoaras, and Piet Herdewijn. "In vitro evaluation of hexitol nucleic acid antisense oligonucleotides." In XIth Symposium on Chemistry of Nucleic Acid Components. Prague: Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, 1999. http://dx.doi.org/10.1135/css199902151.

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Zheng, Shi-ying, Dong Jiang, Jin-feng Ge, and Zhen-ya Shen. "T Cell Apoptosis Was Inhibited by Fas of Antisense Oligonucleotide." In 2010 4th International Conference on Bioinformatics and Biomedical Engineering (iCBBE). IEEE, 2010. http://dx.doi.org/10.1109/icbbe.2010.5516974.

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Zhang, M., J. R. Crosby, D. Bai, C. Zhao, A. R. Macleod, and S. Guo. "Antisense Oligonucleotide Mediated Reduction of TG2 Attenuates Pulmonary Fibrosis in Mice." In American Thoracic Society 2019 International Conference, May 17-22, 2019 - Dallas, TX. American Thoracic Society, 2019. http://dx.doi.org/10.1164/ajrccm-conference.2019.199.1_meetingabstracts.a7438.

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Kuijper, Elsa, Maurice Overzier, Ernst Suidgeest, Louise van der Weerd, Lodewijk Toonen, and Willeke van Roon-Mom. "I02 Therapeutic effect of antisense oligonucleotide treatment in YAC128 Huntington mice." In EHDN 2022 Plenary Meeting, Bologna, Italy, Abstracts. BMJ Publishing Group Ltd, 2022. http://dx.doi.org/10.1136/jnnp-2022-ehdn.228.

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Ripple, Michael J., Dahui You, Joseph Giamo, Andrew Sewell, David M. Becnel, and Stephania A. Cormier. "Inhaled IL-4 Receptor A Antisense Oligonucleotide Prevents RSV-Mediated Asthma." In American Thoracic Society 2010 International Conference, May 14-19, 2010 • New Orleans. American Thoracic Society, 2010. http://dx.doi.org/10.1164/ajrccm-conference.2010.181.1_meetingabstracts.a6185.

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Logan, Andrew E., Timothy R. Wilson, Patrick G. Johnston, and Daniel B. Longley. "Abstract C12: Preclinical characterization of a novel FLIP‐targeted antisense phosphorothioate oligonucleotide." In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics--Nov 15-19, 2009; Boston, MA. American Association for Cancer Research, 2009. http://dx.doi.org/10.1158/1535-7163.targ-09-c12.

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Sakurai, Kazuo, Shinichi Mochizuki, and Jusaku Minari. "Antisense Oligonucleotides Delivery to the Antigen Presenting Cells by using Schizophyllan." In 2008 MRS Fall Meetin. Materials Research Society, 2008. http://dx.doi.org/10.1557/proc-1140-hh05-17.

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Reports on the topic "Oligonucleotidi antisenso"

1

Liu, Yong-Yu. Antisense Oligonucleotides to Glucosylceramide Synthase Can Reverse Multidrug Resistance in Breast Cancer. Fort Belvoir, VA: Defense Technical Information Center, June 2002. http://dx.doi.org/10.21236/ada407440.

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Chakraborty, Srijani. The Dawn of RNA Therapeutics. Spring Library, December 2020. http://dx.doi.org/10.47496/sl.blog.19.

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