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1.
J Gene Med ; : e3396, 2021 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-34713552

RESUMO

BACKGROUND: We previously determined that polyplexes formed by linear H2K peptides were more effective in transfecting tumors in vivo than polyplexes formed by branched H2K4b-20 peptides. Based on trypsin digest and salt displacement studies, the linear H2K polyplexes were less stable than the branched H2K4b-20 polyplexes. Because binding and release of the polymer and DNA from the H2K4b-20 polyplex may account for the ineffectiveness, we investigated whether four-branched histidine-lysine (HK) peptides with varying numbers of amino acids in their branches would be more effective in their ability to increase gene expression in tumors in vivo. METHODS: Linear and branched peptides with multiple -KHHK- motifs were synthesized by solid-phase synthesis. The branched H2K4b-20, -18, -14 and 12 peptides had 20, 18, 14 and 12 amino acids in their branches, respectively. These peptides were examined for their ability to carry luciferase-expressing plasmids to human breast cancer xenografts in a mouse model. With gel retardation and in vivo transfection, the incorporation of a targeting ligand and an endosomal lysis peptide into these polyplexes was also examined. A blocking antibody was pre-injected prior to the polyplexes to determine the role of neuropilin 1 in the uptake of these polyplexes by the tumor. The size of the polyplexes was measured by dynamic light scattering. RESULTS: Of the four negative surface-charge polyplexes formed by the branched carriers, the H2K4b-14 polyplex was determined to be the most effective plasmid delivery platform to tumors. The incorporation of a targeting ligand and an endosomal lysis peptide into H2K4b-14 polyplexes further enhanced their ability to transfect tumors in vivo. Furthermore, after pre-injecting tumor-bearing mice with a blocking antibody to the neuropilin-1 receptor (NRP-1), there was a marked reduction of tumor gene expression with the modified H2K4b-14 polyplexes, suggesting that NRP-1 mediated their transport into the tumor. CONCLUSIONS: The present study established that branched peptides intermediate in length were very efficient in delivering plasmids to tumors in vivo.

2.
J Gene Med ; 23(2): e3295, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33171540

RESUMO

BACKGROUND: Previously, we determined that four-branched histidine-lysine (HK) peptides were effective carriers of plasmids and small interfering RNA. In the present study, we compared several branched HK carriers and, in particular, two closely-related H3K4b and H3K(+H)4b peptides for their ability as carriers of mRNA. The H3K(+H)4b peptide differed from its parent analogue, H3K4b, by only a single histidine in each branch. METHODS: A series of four-branched HK peptides with varied sequences was synthesized on a solid-phase peptide synthesizer. The ability of these peptides to carry mRNA expressing luciferase to MDA-MB-231 cells was investigated. With gel retardation and heparin displacement assays, the stability of HK polyplexes was examined. We determined the intracellular uptake of HK polyplexes by flow cytometry and fluorescence microscopy. The size and polydispersity index of the polyplexes in several media were measured by dynamic light scattering. RESULTS: MDA-MB-231 cells transfected by H3K(+H)4b-mRNA polyplexes expressed 10-fold greater levels of luciferase than H3K4b polyplexes. With gel retardation and heparin displacement assays, the H3K(+H)4b polyplexes showed greater stability than H3K4b. Intracellular uptake and co-localization of H3K(+H)4b polyplexes within acidic endosomes were also significantly increased compared to H3K4b. Similar to H3K(+H)4b, several HK analogues with an additional histidine in the second domain of their branches were effective carriers of mRNA. When combined with DOTAP liposomes, H3K(+H)4b was synergistic in delivery of mRNA. CONCLUSIONS: H3K(+H)4b was a more effective carrier of mRNA than H3K4b. Mechanistic studies suggest that H3K(+H)4b polyplexes were more stable than H3K4b polyplexes. Lipopolyplexes formed with H3K(+H)4b markedly increased mRNA transfection.

3.
Pharmaceutics ; 12(8)2020 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-32823960

RESUMO

Histidines incorporated into carriers of nucleic acids may enhance the extracellular stability of the nanoparticle, yet aid in the intracellular disruption of the nanoparticle, enabling the release of the nucleic acid. Moreover, protonation of histidines in the endosomes may result in endosomal swelling with subsequent lysis. These properties of histidine are based on its five-member imidazole ring in which the two nitrogen atoms may form hydrogen bonds or act as a base in acidic environments. A wide variety of carriers have integrated histidines or histidine-rich domains, which include peptides, polyethylenimine, polysaccharides, platform delivery systems, viral phages, mesoporous silica particles, and liposomes. Histidine-rich carriers have played key roles in our understanding of the stability of nanocarriers and the escape of the nucleic acids from endosomes. These carriers show great promise and offer marked potential in delivering plasmids, siRNA, and mRNA to their intracellular targets.

4.
Methods Mol Biol ; 1974: 161-180, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31099002

RESUMO

With the recent explosion of genomic information on the root causes of disease, there is an increased interest in nucleic acid therapeutics, including siRNA and gene therapy, all of which require delivery of highly charged nucleic acids from siRNA with a molecular weight of about 1.4 × 104 to plasmids with an approximate molecular weight of 2.0-3.0 × 106. This chapter describes the delivery of shRNA via plasmid or siRNA with a peptide-based carrier. We focus on the histidine-lysine peptide which serves as an example for other peptides and polymeric carrier systems. When the HK peptide and nucleic acids are mixed together and interact with one another through ionic and nonionic interactions, nanoplexes are formed. These nanoplexes, carrying either shRNA or siRNA that target oncogenes, provide promising options for the treatment of cancer. We describe methods of preparation and characterization of these nanoplexes using dynamic light scattering, zeta potential, and gel retardation assays. We also provide protocols for transfection in vitro and in vivo for these nanoplexes.


Assuntos
Terapia Genética/métodos , Nanotecnologia/métodos , Neoplasias/terapia , RNA Interferente Pequeno/genética , Animais , Humanos , Camundongos , Neoplasias/genética , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Biochem Biophys Res Commun ; 513(1): 242-247, 2019 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-30954222

RESUMO

Doxorubicin (Dox) has widespread use as a cancer chemotherapeutic agent, but Dox is limited by several side effects including irreversible cardiomyopathy. Although liposomal Dox formulations, such as Doxil, mitigate side effects, they do not prolong survival in many patients. As a result, efforts have continued to discover improved formulations of Dox. We previously found that a peptide-based nanoplex delivered plasmid DNA efficiently to tumors in murine models. Unlike the majority of nanoparticles that depend solely on enhanced permeability and retention (EPR) for their transport into the tumor, our peptide-based nanoplex has a potential advantage in that its uptake primarily depends on neuropilin-1 receptor targeting. Because Dox binds to DNA, we tested whether this delivery platform could effectively deliver Dox to tumors and reduce their size. The nanoplexes increased the levels of Dox in tumors by about 5.5-fold compared to aqueous (free) Dox controls. Consistent with enhanced levels in the tumor, the nanoplex-Dox treatment had significantly greater anti-tumor activity. Whereas low dose free Dox did not reduce the size of tumors compared to untreated controls, the low dose nanoplex-Dox reduced the size of tumors by nearly 55% (p < 0.001). The high dose nanoplex-Dox also inhibited the size of tumor significantly more than the comparable high-dose free Dox (p < 0.001). Furthermore, apoptosis and proliferation markers (Ki67) of tumors observed in the different treatment groups correlated with their ability to inhibit tumor size. This study shows the efficacy of an NRP-1 targeted nanoplexes to deliver Dox to tumors in vivo and lays the groundwork for more complex and effective formulations.


Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Portadores de Fármacos/metabolismo , Sistemas de Liberação de Medicamentos , Neoplasias/tratamento farmacológico , Peptídeos/metabolismo , Animais , Antibióticos Antineoplásicos/farmacocinética , Antibióticos Antineoplásicos/uso terapêutico , Apoptose , Linhagem Celular Tumoral , Doxorrubicina/farmacocinética , Doxorrubicina/uso terapêutico , Feminino , Humanos , Camundongos , Camundongos Nus , Nanopartículas/metabolismo , Neoplasias/metabolismo , Neuropilina-1/metabolismo
6.
Artigo em Inglês | MEDLINE | ID: mdl-30613436

RESUMO

Doxorubicin is a widely used chemotherapy agent. Despite its utility, several adverse side effects, especially its irreversible cardiotoxicity and reversible nephrotoxicity, have prompted the development of liposomal carriers, many of which are FDA approved. Antitumor efficacies of approved liposome-Dox preparations can equal or exceed that of conventional doxorubicin. Because these liposomes carriers accumulate in solid tumor tissues via an enhanced permeation and retention (EPR) effect, these carriers have an improved safety profile. Nevertheless, a significant problem with the current drug delivery preparations of doxorubicin is a lack of efficacy toward tumors that exhibit multidrug resistance. In this review, we consider the development of drug delivery systems for doxorubicin, which improve the therapeutic window (efficacy and safety) and which address limitations of the current FDA-approved doxorubicin formulations.

7.
J Drug Deliv ; 2017: 6971297, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29218233

RESUMO

Over the past 20 years, a diverse group of ligands targeting surface biomarkers or receptors has been identified with several investigated to target siRNA to tumors. Many approaches to developing tumor-homing peptides, RNA and DNA aptamers, and single-chain variable fragment antibodies by using phage display, in vitro evolution, and recombinant antibody methods could not have been imagined by researchers in the 1980s. Despite these many scientific advances, there is no reason to expect that the ligand field will not continue to evolve. From development of ligands based on novel or existing biomarkers to linking ligands to drugs and gene and antisense delivery systems, several fields have coalesced to facilitate ligand-directed siRNA therapeutics. In this review, we discuss the major categories of ligand-targeted siRNA therapeutics for tumors, as well as the different strategies to identify new ligands.

8.
Oncotarget ; 8(46): 80651-80665, 2017 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-29113333

RESUMO

Excessive skin scars due to elective operations or trauma represent a challenging clinical problem. Pathophysiology of hypertrophic scars entails a prolonged inflammatory and proliferative phase of wound healing. Over expression of TGF-ß1 and COX-2 play key regulatory roles of the aberrant fibrogenic responses and proinflammatory mediators. When we silenced TGF-ß1 and COX-2 expression simultaneously in primary human fibroblasts, a marked increase in the apoptotic cell population occurred in contrast to those only treated with either TGF-ß1 or COX-2 siRNA alone. Furthermore, using human hypertrophic scar and skin graft implant models in mice, we observed significant size reductions of the implanted tissues following intra-scar administration of TGF-ß1/COX-2 specific siRNA combination packaged with Histidine Lysine Polymer (HKP). Gene expression analyses of those treated tissues revealed silencing of the target gene along with down regulations of pro-fibrotic factors such as α-SMA, hydroxyproline acid, Collagen 1 and Collagen 3. Using TUNEL assay detection, we found that the human fibroblasts in the implanted tissues treated with the TGF-ß1/COX-2siRNAs combination exhibited significant apoptotic activity. Therefore we conclude that a synergistic effect of the TGF-ß1/COX-2siRNAs combination contributed to the size reductions of the hypertrophic scar implants, through activation of fibroblast apoptosis and re-balancing between scar tissue deposition and degradation.

9.
Drugs Future ; 42(2): 95-104, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28603338

RESUMO

Whereas uptake of low molecular weight agents is generally inhibited in tumors due to high interstitial pressure, tumor uptake of macromolecules is increased due to enhanced permeability and retention (EPR). Small molecule drugs alone or incorporated in nanoparticles (NP) have largely been dependent on such physical tumor uptake (passive) for therapeutic activity. Although passive targeted NP such as Stealth Liposomal Doxorubicin (Doxil ®) are effective with improved safety, drug delivery to tumors is still significantly limited. To improve tumor delivery and efficacy, tumor-penetrating peptides (TPP), which contain sequences that target the tumor and activate the neuropilin-1 receptor (NRP1), have either been co-administered with or conjugated to both small and large therapeutic molecules. In this review, we will discuss TPP-mediated therapeutics which target the NRP1 transport system of tumors.

10.
Biochem Biophys Res Commun ; 477(4): 957-962, 2016 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-27387239

RESUMO

Previously, a four branched histidine-lysine rich peptide, H3K4b, was shown to demonstrate selective antifungal activity with minimal antibacterial activity. Due to the potential breakdown from proteases, H3K4b was further evaluated in the current study by varying the D- and l-amino acid content in its branches. Whereas analogues of H3K4b that selectively replaced l-amino acids (H3k4b, h3K4b) had improved antifungal activity, the all d-amino acid analogue, h3k4b, had reduced activity, suggesting that partial breakdown of the peptide may be necessary. Moreover, because histidines form coordination bonds with the silver ion, we examined whether silver adducts can be formed with these branched histidine-lysine peptides, which may improve antifungal activity. For Candida albicans, the silver adduct of h3K4b or H3k4b reduced the MIC compared to peptide and silver ions alone by 4- and 5-fold, respectively. For Aspergillus fumigatus, the silver adducts showed even greater enhancement of activity. Although the silver adducts of H3k4b or h3K4b showed synergistic activity, the silver adduct with the all l-amino acid H3K4b surprisingly showed the greatest synergistic and growth inhibition of A. fumigatus: the silver adduct of H3K4b reduced the MIC compared to the peptide and silver ions alone by 30- and 26-fold, respectively. Consistent with these antifungal efficacy results, marked increases in free oxygen radicals were produced with the H3K4b and silver combination. These studies suggest that there is a balance between stability and breakdown for optimal antifungal activity of the peptide alone and for the peptide-silver adduct.


Assuntos
Antifúngicos/administração & dosagem , Aspergillus fumigatus/citologia , Aspergillus fumigatus/efeitos dos fármacos , Histidina/administração & dosagem , Peptídeos/administração & dosagem , Prata/administração & dosagem , Antifúngicos/síntese química , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Sinergismo Farmacológico , Histidina/química , Peptídeos/química , Prata/química
11.
J Gene Med ; 18(7): 134-44, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27257039

RESUMO

BACKGROUND: Promising plasmid-based treatments have limited value without an effective delivery system. Recently, the linear H2K with a repeating -KHHK- pattern was determined to be an effective plasmid carrier to tumor xenografts in vivo. Although unpacking of the H2K polyplex within the tumor may have a role, the mechanism for the enhanced efficacy remains unclear. METHODS: After solid-phase synthesis of linear and branched histidine-lysine (HK) peptide carriers of plasmids, the peptides were compared for their ability to lyse endosomes with a red blood cell model and to transfect MDA-MB-435 xenografts in the presence or absence of neuropilin-1 receptor (NRP-1) antibodies. To examine stability, polyplexes were incubated with trypsin or NaCl and then analyzed by electrophoresis. RESULTS: After screening peptides with a model for endosomal lysis at two pHs, the 33-mer H3K peptide lysed red blood cells effectively at the lower pH. Combining H3K and H2K peptides as carriers of plasmids expressing luciferase were more effective than H2K alone. Based on the repeating -KHHK- sequences of H2K, we studied whether the widespread gene expression in the tumor may be mediated by NRP-1. By blocking NRP-1 in tumor-bearing mice, luciferase activity in tumors delivered by HK polyplexes was reduced by 96%, whereas activity in normal tissues was minimally reduced. CONCLUSIONS: Combining an endosomolytic peptide, H3K, with H2K polyplexes as a carrier further enhanced transfection in vivo. Moreover, the widespread distribution of H2K polyplexes is mediated by NRP-1, suggesting that transcytosis of these polyplexes through the tumor endothelium may lead to efficient transfection. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Neoplasias/genética , Neuropilina-1/genética , Peptídeos/genética , Transfecção/métodos , Sequência de Aminoácidos , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Linhagem Celular Tumoral , Feminino , Terapia Genética/métodos , Histidina/genética , Histidina/metabolismo , Humanos , Luciferases/genética , Luciferases/metabolismo , Lisina/genética , Lisina/metabolismo , Camundongos Nus , Neoplasias/metabolismo , Neoplasias/terapia , Neuropilina-1/imunologia , Neuropilina-1/metabolismo , Peptídeos/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
13.
J Gene Med ; 16(9-10): 317-28, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25303767

RESUMO

BACKGROUND: Selecting nonviral carriers for in vivo gene delivery is often dependent on determining the optimal carriers from transfection assays in vitro. The rationale behind this in vitro strategy is to cast a net sufficiently wide to identify the few effective carriers of plasmids for in vivo studies. Nevertheless, many effective in vivo carriers may be overlooked by this strategy because of the marked differences between in vitro and in vivo assays. METHODS: After solid-phase synthesis of linear and branched histidine/lysine (HK) peptides, the two peptide carriers were compared for their ability to transfect MDA-MB-435 tumor cells in vitro and then in vivo. RESULTS: By contrast to their transfection activity in vitro, the linear H2K carrier of plasmids was far more effective in vivo compared to the branch H2K4b. Surprisingly, negatively-charged polyplexes formed by the linear H2K peptide gave higher transfection in vivo than did those with a positive surface charge. To examine the distribution of plasmid expression within the tumor from H2K polyplexes, we found widespread expression by immunohistochemical staining. With a fluorescent tdTomato expressing-plasmid, we confirmed a pervasive distribution and gene expression within the tumor mediated by the H2K polyplex. CONCLUSIONS: Although mechanisms underlying the efficiency of gene expression are probably multifactorial, unpacking of the H2K polyplex within the tumor appears to have a significant role. Further development of these H2K polyplexes represents an attractive approach for plasmid-based therapies of cancer.


Assuntos
Biopolímeros/química , Regulação Neoplásica da Expressão Gênica , Histidina/química , Plasmídeos/genética , Animais , Linhagem Celular Tumoral , Feminino , Técnicas de Transferência de Genes , Genes Reporter , Terapia Genética , Humanos , Lisina/química , Camundongos , Camundongos Nus , Tamanho da Partícula , Transfecção , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Angew Chem Int Ed Engl ; 53(40): 10631-5, 2014 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-25146232

RESUMO

Gene delivery is a promising way to treat hereditary diseases and cancer; however, there is little understanding of DNA:carrier complex mechanical properties, which may be critical for the protection and release of nucleic acids. We applied optical tweezers to directly measure single-molecule mechanical properties of DNA condensed using 19-mer poly-L-lysine (PLL) or branched histidine-lysine (HK) peptides. Force-extension profiles indicate that both carriers condense DNA actively, showing force plateaus during stretching and relaxation cycles. As the environment such as carrier concentration, pH, and the presence of zinc ions changes, DNA:HK complexes showed dynamically regulated mechanical properties at multiple force levels. The fundamental knowledge from this study can be applied to design a mechanically tailored complex which may enhance transfection efficiency by controlling the stability of the complex temporally and spatially.


Assuntos
DNA/administração & dosagem , DNA/química , Técnicas de Transferência de Genes , Peptídeos/química , Polilisina/química , Sequência de Aminoácidos , Cátions Bivalentes/química , Histidina , Lisina/química , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Pinças Ópticas , Zinco/química
15.
J Drug Target ; 22(6): 536-42, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24731059

RESUMO

The treatment of invasive candidiasis associated with growing numbers of immunocompromised patients remains a major challenge complicated by increasing drug resistance. A novel class of branched histidine-lysine (bHK) peptides has promising antifungal activity, and exhibits a mechanism similar to natural histatins, and thus may avoid drug resistance. The present studies evaluate ligand targeting of bHK peptides to fungal surface integrins by determining whether a cyclic RGD (cRGD) peptide with a large PEG linker could enhance bHK peptide antifungal activity. Whereas conjugates containing only the PEG linker reduced bHK peptide activity, conjugates with the cRGD-PEG ligand resulted in marked enhancement of activity against Candida albicans. This study provides the first demonstration of benefit from ligand targeting of antifungal agents to fungal surface receptors.


Assuntos
Antifúngicos/farmacologia , Sistemas de Liberação de Medicamentos , Histidina/análise , Integrinas/efeitos dos fármacos , Peptídeos/farmacologia , Candida albicans/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Peptídeos/química , Polietilenoglicóis/química
16.
Biomaterials ; 35(2): 846-55, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24161165

RESUMO

Branched peptides containing histidines and lysines (HK) have been shown to be effective carriers for DNA and siRNA. We anticipate that elucidation of the binding mechanism of HK with siRNA will provide greater insight into the self-assembly and delivery of the HK:siRNA polyplex. Non-covalent bonds between histidine residues and nucleic acids may enhance the stability of siRNA polyplexes. We first compared the polyplex biophysical properties of a branched HK with those of branched asparagine-lysine peptide (NK). Consistent with siRNA silencing experiments, gel electrophoresis demonstrated that the HK siRNA polyplex maintained its integrity with prolonged incubation in serum, whereas siRNA in complex with NK was degraded in a time-dependent manner. Isothermal titration calorimetry of various peptides binding to siRNA at pH 7.3 showed that branched polylysine, interacted with siRNA was initially endothermic, whereas branched HK exhibited an exothermic reaction at initial binding. The exothermic interaction indicates formation of non-ionic bonds between histidines and siRNA; purely electrostatic interaction is entropy-driven and endothermic. To investigate the type of non-ionic bond, we studied the protonation state of imidazole rings of a selectively (15)N labeled branched HK by heteronuclear single quantum coherence NMR. The peak of Nδ1-H tautomers of imidazole shifted downfield (in the direction of deprotonation) by 0.5-1.0 ppm with addition of siRNA, providing direct evidence that histidines formed hydrogen bonds with siRNA at physiological pH. These results establish that histidine-rich peptides form hydrogen bonds with siRNA, thereby enhancing the stability and biological activity of the polyplex in vitro and in vivo.


Assuntos
Inativação Gênica , Histidina/metabolismo , RNA Interferente Pequeno/química , Asparagina/química , Linhagem Celular Tumoral , Humanos , Ligação de Hidrogênio , Concentração de Íons de Hidrogênio , Lisina/química , Espectroscopia de Ressonância Magnética , Peptídeos/química , Polilisina/química , Polímeros/química , Conformação Proteica , Transfecção
17.
Biomacromolecules ; 14(3): 752-60, 2013 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-23360232

RESUMO

We characterized in this study the pharmacokinetics and antitumor efficacy of histidine-lysine (HK):siRNA nanoplexes modified with PEG and a cyclic RGD (cRGD) ligand targeting αvß3 and αvß5 integrins. With noninvasive imaging, systemically administered surface-modified HK:siRNA nanoplexes showed nearly 4-fold greater blood levels, 40% higher accumulation in tumor tissue, and 60% lower luciferase activity than unmodified HK:siRNA nanoplexes. We then determined whether the surface-modified HK:siRNA nanoplex carrier was more effective in reducing MDA-MB-435 tumor growth with an siRNA targeting Raf-1. Repeated systemic administration of the selected surface modified HK:siRNA nanoplexes targeting Raf-1 showed 35% greater inhibition of tumor growth than unmodified HK:siRNA nanoplexes and 60% greater inhibition of tumor growth than untreated mice. The improved blood pharmacokinetic results and tumor localization observed with the integrin-targeting surface modification of HK:siRNA nanoplexes correlated with greater tumor growth inhibition. This investigation reveals that through control of targeting ligand surface display in association with a steric PEG layer, modified HK: siRNA nanoplexes show promise to advance RNAi therapeutics in oncology and potentially other critical diseases.


Assuntos
Histidina/química , Lisina/química , Nanoestruturas/química , Neoplasias/tratamento farmacológico , RNA Interferente Pequeno/farmacocinética , Animais , Linhagem Celular Tumoral , Feminino , Expressão Gênica , Inativação Gênica , Humanos , Camundongos , Proteínas Proto-Oncogênicas c-raf/genética , Proteínas Proto-Oncogênicas c-raf/metabolismo , RNA Interferente Pequeno/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Mol Ther ; 20(12): 2282-90, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23032972

RESUMO

Induction of cytokines by small interfering RNA (siRNA) polyplexes has been a significant concern of researchers attempting to minimize the toxicity of this promising therapy. Although cationic carriers of siRNA are known to increase cytokine levels, few systematic studies have been done to determine what properties of the carrier are important to modulate cytokines. Because branched histidine-lysine (HK) peptides are effective carriers of siRNA and their sequence can be readily modified, we selected this class of carrier to determine which sequences of the peptide were important for cytokine induction. With the use of peripheral blood mononuclear cells (PBMCs), the HK peptide with a higher number of histidines (H3K(+H)4b) in complex with siRNA induced lower levels of cytokines compared with other HK (e.g., H2K4b, H3K4b, H3K(+N)4b) siRNA nanoplexes. Notably, these peptides' siRNA polyplexes showed a similar pattern of cytokine induction when injected intravenously in a mouse model, i.e., the HK with higher content of histidines induced cytokines the least. As indicated by the pH-sensitive dye within acidic endosomes, the greater pH-buffering capacity of H3K(+H)4b compared with other HK peptides may explain why cytokine levels were reduced. In addition to buffering capacity, the size of HK polyplexes markedly influenced cytokine production.


Assuntos
Citocinas/metabolismo , RNA Interferente Pequeno/genética , Animais , Células Cultivadas , Citometria de Fluxo , Histidina/química , Humanos , Concentração de Íons de Hidrogênio , Leucócitos Mononucleares/metabolismo , Lisina/química , Camundongos , Camundongos Endogâmicos BALB C , Peptídeos/química
19.
Future Med Chem ; 1(9): 1671-81, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21425985

RESUMO

While the discovery of RNA interference (RNAi) has been considered one of the most significant breakthroughs in biomedicine, its prospects for novel therapeutic applications are even more exciting. The high specificity, exquisite selectivity and chemical homogeneity of small interfering RNAs (siRNA; intermediates in RNAi activity), provide unique advantages for these moieties as multi-targeted inhibitory drugs. Many such applications have demonstrated significant benefit compared with single gene-targeted siRNA inhibitors. In this article, we will review the current status of using a multi-targeted siRNA cocktail for novel therapeutic development in the treatment of cancer and viral infections. We will also propose the characteristics of various types of siRNA cocktails and their design, while recognizing the potential future impact of and challenges facing this unique therapeutic modality.


Assuntos
Interferência de RNA , RNA Interferente Pequeno/uso terapêutico , Humanos , Neoplasias/genética , Neoplasias/terapia , Viroses/genética , Viroses/terapia
20.
Drugs Future ; 34(9): 721, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20161621

RESUMO

Sequence-specific gene silencing with small interfering RNA (siRNA) has transformed basic science research, and the efficacy of siRNA therapeutics toward a variety of diseases is now being evaluated in pre-clinical and clinical trials. Despite its potential value, the highly negatively charged siRNA has the classic delivery problem of requiring transport across cell membranes to the cytosol. Consequently, carrier development for siRNA delivery is one of the most important problems to solve before siRNA can achieve widespread clinical use. An assortment of non-viral carriers including liposomes, peptides, polymers, and aptamers are being evaluated for their ability to shepherd siRNA to the target tissue and cross the plasma membrane barrier into the cell. Several promising carriers with low toxicity and increased specificity for disease targets have emerged for siRNA-based therapeutics. This review will discuss non-viral approaches for siRNA therapeutics, with particular focus on synthetic carriers for in vivo systemic delivery of siRNA.

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