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Comparison of Chemical Structure for PNA and DNA: In the structure of PNA, bases (A, T, G, C) are linked with polyamide backbone as against sugar phosphate backbone in DNA.
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Since the discovery and synthesis of a novel DNA mimic, peptide nucleic acid (PNA) in 1991, PNAs have attracted tremendous interest and have shown great promise as potential antisense drugs. They have been used extensively as tools for specific modulation of gene expression by targeting translation or transcription processes. This review discusses...
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... expression, could be, individually or collectively, potential targets for the next generation of drug design and development. In this context, peptide nucleic acids (PNAs) have recently been explored as potential antisense agents for targeting these regulatory regions on the HIV-1 genome. PNAs are DNA mimics in which sugar phosphate backbone ( Fig. 1) is replaced with N-(2-amino ethyl) glycine (aeg) units [2]. This modification makes them highly stable in the cellular compartment. In addition, PNAs show stronger binding affinity to their complementary RNA or DNA than do the respective RNA/DNA or DNA/DNA hybridizations ...
Citations
... 45 The standard of care for treating HIV consists of treatment mechanisms that target the enzymes protease and reverse transcriptase. 48 HIV is a virus prone to rapid genome variation leading to development of strains resistant to available treatments. Newer drugs being explored include those targeting accessory and regulatory proteins of HIV. ...
... One established potential target for anti-HIV PNA is the HIV-1 transactivator response (TAR) regulatory region of the genome contained within the long terminal repeat region. 48 CEM-line cells were first infected with HIV-1 virions, which possessed a bioluminescence luciferase gene to enable detection of HIV-1. The anti-TAR PNA was incubated with the CEM cells, and HIV-1 levels were quantified via analyzing luciferase activity. ...
... Anti-TAR PNA effectively inhibited reverse transcription of TAR-RNA, decreasing HIV-1 replication. 48,49 When conjugated to carrier peptides to assist with cellular entry, the anti-TAR PNA exhibited potent virucidal properties, leaving HIV cells non-infectious. 48,49 None of the CPPs resulted in decreased affinity of PNA to target RNA. ...
Peptide nucleic acids (PNAs) are synthetic nucleic acid analogs with a neutral N-(2-aminoethyl) glycine backbone. PNAs possess unique physicochemical characteristics such as increased resistance to enzymatic degradation, ionic strength and stability over a wide range of temperatures and pH, and low intrinsic electrostatic repulsion against complementary target oligonucleotides. PNA has been widely used as an antisense oligonucleotide (ASO). Despite the favorable characteristics of PNA, in comparison with other ASO technologies, the use of antisense PNA for novel therapeutics has lagged. This review provides a brief overview of PNA, its antisense mechanisms of action, delivery strategies, and highlights successful applications of PNA, focusing on anti-pathogenic, anti-neurodegenerative disease, anti-cancer, and diagnostic agents. For each application, several studies are discussed focusing on the different target sites of the PNA, design of different PNAs and the therapeutic outcome in different cell lines and animal models. Thereafter, persisting limitations slowing the successful integration of antisense PNA therapeutics are discussed in order to highlight actionable next steps in the development and optimization of PNA as an ASO.
... Peptide nucleic acid (PNA), a novel DNA mimic, has attracted tremendous interest as potential antisense strategy as a tool for specific modulation of gene expression by targeting translation or transcription processes. PNA compounds have therapeutic potential as anti-HIV-1 drugs (Pandey et al 2009). PR2, îs a PNA complementary to a sequence of the viral protease-encoding gene, is effective in blocking HIV release, when used at high doses and targeting to the macrophage compartment is facilitated by use of RBCs (Fraternale et al 2009). ...
This report reviews the current state-of-art of antiviral approaches including vaccines, pharmaceuticals and innovative technologies for delivery of therapeutics. The introduction starts with a practical classification of viral diseases according to their commercial importance. Various antiviral approaches are described including pharmaceuticals and molecular biological therapies such as gene therapy and RNA interference (RNAi) as well as vaccines for virus infections. Expert opinion is given about the current problems and needs in antiviral therapy. SWOT (strengths, weaknesses, opportunities and threats) analysis of antiviral approaches is presented against the background of concept of an ideal antiviral agent.
A novel feature of this report is the use of nanotechnology in virology and its potential for antiviral therapeutics. Interaction of nanoparticles with viruses are described. NanoViricides are polymeric micelles, which act as nanomedicines to destroy viruses. Various methods for local as well as systemic delivery of antiviral agents and vaccines are described. Nanobiotechnology plays an important role in improving delivery of antivirals. Advantages and limitations of delivery of gene-based, antisense and RNAi antiviral therapeutics are discussed.
Anti-influenza measures applicable to human as well as avian forms are described including the recent epidemic of swine flu. Resistance can develop against neuraminidase inhibitors although it is less than that with adamantanes. Considering these problems, there is need for a more effective agent. Investigations into alternative anti-influenza target will probably expand in the coming years. These include the development of mechanisms to inhibit fusion between the virus envelope and the cell membrane.
After a discussion of current therapies of AIDS/HIV and their limitations, new strategies in development of antiviral agents are described. Drug resistance and toxicities are emerging as major treatment challenges. Based on a review of technologies and drugs in development, it can be stated that there are good prospects are of finding a cure for HIV/AIDS in the next decade.
Hepatitis viruses are described with focus on hepatitis C virus (HCV) and hepatitis B virus (HBV). Despite the presence of numerous drug candidates in the anti-HCV pipeline, and the commitment of major R&D resources by many pharmaceutical companies, it might still take several years for any new anti-HCV drugs to reach the market. Although many companies are focusing their efforts on developing viral inhibitors, cellular targets in the host are beginning to emerge as attractive possibilities because they might enable the development of broad-spectrum antiviral drugs with less chance for developing viral resistance.
Various commercially important viruses include herpes simplex (HSV) and human papilloma virus (HPV). There a number of treatments but HSV is not destroyed completely and remains dormant and activates from time to time to cause various clinical manifestations. There is discussion about the role of HPV in cervical cancer and vaccines available now seem to be adequate in preventing HSV-induced cervical cancer. There is no effective vaccine for respiratory syncytial virus (RSV) although monoclonal antibody (MAb) treatment is useful for prophylaxis and reducing the clinical manifestations. There is a need for an agent to eliminate this virus.
Various viruses that either occur in epidemics or in tropics and some naturally emerging infectious diseases are described, e.g. viral hemorrhagic fevers such as dengue and West Nile virus infection. These are a constant threat and impossible to anticipate. Some of these lack antiviral agents or vaccines for prevention. Although these include some of the most serious viral disorders, the development of antiviral agents for these is not commercially attractive. Current research and approaches to these virus infections, particularly the current pandemic of COVID-19, are discussed. There are over 110 drugs and vaccine candidates in development of which 3 have been approved by the FDA and other health authorities around the world. Vaccination is being carried out in several countries
Markets for antivirals are considered according to viruses and diseases caused by them and also according to management approaches: antiviral drugs, vaccines, MAbs and innovative approaches that include immunological and use of other technologies such as gene therapy, antisense, RNAi and nanobiotechnology. Antiviral markets are estimated starting with 2020 with projections up to the year 2030.
Profiles of 197 companies that are involved in developing various technologies and products are profiled and with 182 collaborations. These include major pharmaceutical companies (12), Biopharmaceutical companies with antiviral products (87), Antiviral drug companies (26) as well as viral vaccine companies (71). The report is supplemented with 58 tables, 17 figures and 550 references from the literature.
... In this study, after PNA administration to the animal, the functions of all organs were normal, and no side effects was seen in the studied mice. The researchers reported PNA as non-toxic at all therapeutic doses (42). Several therapeutic approaches and novel antimicrobial agents are limited due to their potential toxicity and side effects. ...
Background and objectives:
Helicobacter pylori causes several gastrointestinal diseases, including asymptomatic gastritis, chronic peptic ulcer, duodenal ulcer, lymphoma of the mucosa-associated lymphoid tissue (MALT), and gastric adenocarcinoma. In recent years, failure to eradicate H. pylori infections has become an alarming problem for physicians. It is now clear that the current treatment strategies may become ineffective, necessitating the development of innovative antimicrobial compounds as alternative treatments.
Materials and methods:
In this experimental study, a cell-penetrating peptide-conjugated peptide nucleic acid (CPP-PNA) was used to target the cagA expression. cagA expression was evaluated using RT-qPCR assay after treatment by the CPPPNA in cell culture and animal model. Additionally, immunogenicity and toxicity of the CPP-PNA were assessed in both cell culture and animal models.
Results:
Our analysis showed that cagA mRNA levels reduced in H. pylori-infected HT29 cells after treatment with CPPPNA in a dose-dependent manner. Also, cagA expression in bacterial RNA extracted from stomach tissue of mice treated with PNA was reduced compared to that of untreated mice. The expression of inflammatory cytokines also decreased in cells and tissue of H. pylori-infected mice after PNA treatment. The tested CPP-PNA showed no significant adverse effects on cell proliferation of cultured cells and no detectable toxicity and immunogenicity were observed in mice.
Conclusion:
These results suggest the effectiveness of CPP-PNA in targeting CagA for various research and therapeutic purposes, offering a potential antisense therapy against H. pylori infections.
... The considered biomolecule are based on Peptide Nucleic Acids, DNA analogues described for the first time by Nielsen et al. [45], in which the sugar-phosphate backbone has been replaced by N-(2-aminoethyl)glycine units [15][16][17]46] as depicted in Figure B in S1 File. PNAs have been demonstrated to be very efficient tools for pharmacologically-mediated alteration of gene expression, both in vitro and in vivo [47][48][49], in consideration of the possibility to be used as antisense molecules targeting mRNAs, triple-helix forming molecules targeting eukaryotic gene promoters, artificial promoters, decoy molecules targeting transcription factors [15][16][17][46][47][48][49]. Relevant in the context of the proposed practical laboratory exercise, PNAs have been demonstrated to be able of altering miRNA functions, both in vitro and in vivo [50][51][52][53][54][55][56][57][58]. ...
... The considered biomolecule are based on Peptide Nucleic Acids, DNA analogues described for the first time by Nielsen et al. [45], in which the sugar-phosphate backbone has been replaced by N-(2-aminoethyl)glycine units [15][16][17]46] as depicted in Figure B in S1 File. PNAs have been demonstrated to be very efficient tools for pharmacologically-mediated alteration of gene expression, both in vitro and in vivo [47][48][49], in consideration of the possibility to be used as antisense molecules targeting mRNAs, triple-helix forming molecules targeting eukaryotic gene promoters, artificial promoters, decoy molecules targeting transcription factors [15][16][17][46][47][48][49]. Relevant in the context of the proposed practical laboratory exercise, PNAs have been demonstrated to be able of altering miRNA functions, both in vitro and in vivo [50][51][52][53][54][55][56][57][58]. ...
Practical laboratory classes teaching molecular pharmacology approaches employed in the development of therapeutic strategies are of great interest for students of courses in Biotechnology, Applied Biology, Pharmaceutic and Technology Chemistry, Translational Oncology. Unfortunately, in most cases the technology to be transferred to learning students is complex and requires multi-step approaches. In this respect, simple and straightforward experimental protocols might be of great interest. This study was aimed at presenting a laboratory exercise focusing (a) on a very challenging therapeutic strategy, i.e. microRNA therapeutics, and (b) on the employment of biomolecules of great interest in applied biology and pharmacology, i.e. peptide nucleic acids (PNAs). The aims of the practical laboratory were to determine: (a) the possible PNA-mediated arrest in RT-qPCR, to be eventually used to demonstrate PNA targeting of selected miRNAs; (b) the possible lack of activity on mutated PNA sequences; (c) the effects (if any) on the amplification of other unrelated miRNA sequences. The results which can be obtained support the following conclusions: PNA-mediated arrest in RT-qPCR can be analyzed in a easy way; mutated PNA sequences are completely inactive; the effects of the employed PNAs are specific and no inhibitory effect occurs on other unrelated miRNA sequences. This activity is simple (cell culture, RNA extraction, RT-qPCR are all well-established technologies), fast (starting from isolated and characterized RNA, few hours are just necessary), highly reproducible (therefore easily employed by even untrained students). On the other hand, these laboratory lessons require some facilities, the most critical being the availability of instruments for PCR. While this might be a problem in the case these instruments are not available, we would like to underline that determination of the presence or of a lack of amplified product can be also obtained using standard analytical approaches based on agarose gel electrophoresis.
... Furthermore, they are able to generate triple helices with double-stranded DNA and to perform strand invasion (24)(25)(26). In virtue of these biological activities, PNAs have been demonstrated to be very efficient tools for pharmacologically-mediated alteration of gene expression, both in vitro and in vivo (27)(28)(29). In summary, PNAs and PNA-based analogues were employed as antisense molecules targeting mRNAs, triple-helix forming molecules targeting eukaryotic gene promoters, artificial promoters, and decoy molecules targeting transcription factors (26). ...
The present study investigated the effects of the combined treatment of two peptide nucleic acids (PNAs), directed against microRNAs involved in caspase‑3 mRNA regulation (miR‑155‑5p and miR‑221‑3p) in the temozolomide (TMZ)‑resistant T98G glioma cell line. These PNAs were conjugated with an octaarginine tail in order to obtain an efficient delivery to treated cells. The effects of singularly administered PNAs or a combined treatment with both PNAs were examined on apoptosis, with the aim to determine whether reversion of the drug‑resistance phenotype was obtained. Specificity of the PNA‑mediated effects was analyzed by reverse transcription‑quantitative polymerase‑chain reaction, which demonstrated that the effects of R8‑PNA‑a155 and R8-PNA-a221 anti‑miR PNAs were specific. Furthermore, the results obtained confirmed that both PNAs induced apoptosis when used on the temozolomide‑resistant T98G glioma cell line. Notably, co‑administration of both anti‑miR‑155 and anti‑miR‑221 PNAs was associated with an increased proapoptotic activity. In addition, TMZ further increased the induction of apoptosis in T98G cells co‑treated with anti‑miR‑155 and anti‑miR‑221 PNAs.
... PNA oligonucleotides are generally considered to have negligible toxicity for eukaryotic cells (Good et al., 2001;Evers et al., 2015). However, whether or not PNAs elicit toxic effects is highly dependent on their sequence composition and the presence of a transporter vehicle (Pandey et al., 2009;Rozners, 2012;Zeng et al., 2016). We demonstrated that treatment of HEK-293 cells with the designed PNAs at a concentration of 32 µM did not cause significant cytotoxicity. ...
The search for new, non-standard targets is currently a high priority in the design of new antibacterial compounds. Bacterial toxin–antitoxin systems (TAs) are genetic modules that encode a toxin protein that causes growth arrest by interfering with essential cellular processes, and a cognate antitoxin, which neutralizes the toxin activity. TAs have no human analogs, are highly abundant in bacterial genomes, and therefore represent attractive alternative targets for antimicrobial drugs. This study demonstrates how artificial activation of Escherichia coli mazEF and hipBA toxin–antitoxin systems using sequence-specific antisense peptide nucleic acid oligomers is an innovative antibacterial strategy. The growth arrest observed in E. coli resulted from the inhibition of translation of the antitoxins by the antisense oligomers. Furthermore, two other targets, related to the activities of mazEF and hipBA, were identified as promising sites of action for antibacterials. These results show that TAs are susceptible to sequence-specific antisense agents and provide a proof-of-concept for their further exploitation in antimicrobial strategies.
... Accordingly, they have been used as very efficient tools for pharmacologic alteration of gene expression, both in vitro and in vivo (1)(2)(3)(4)(5). PNAs and PNA-based analogues were proposed as antisense molecules targeting mRNAs, triple-helix forming molecules targeting eukaryotic gene promoters, artificial promoters, decoy molecules targeting transcription factors (4)(5)(6)(7)(8)(9)(10). Recently, PNAs have been shown to be able to alter biological functions of microRNAs, both in vitro and in vivo (11)(12)(13)(14)(15)(16)(17)(18). ...
The biological activity of a combined treatment of U251, U373 and T98G glioma cell lines with two anti-miR PNAs, directed against miR‑221 and miR‑222 and conjugated with an ocataarginine tail (R8-PNA-a221 and R8-PNA-a222) for efficient cellular delivery, was determined. Apoptosis was analyzed, and the effect of the combined treatment of glioma cells with either or both PNAs on the reversion of drug-resistance phenotype was assessed in the temozolomide-resistant T98G glioma cell line. Selectivity of PNA/miRNA interactions was studied by surface plasmon resonance (SPR)-based Biacore analysis. Specificity of the PNA effects at the cellular level was analyzed by RT-qPCR. These experiments support the concept that the effects of R8-PNA-a221 and R8-PNA-a222 are specific. The studies on apoptosis confirmed that the R8-PNA-a221 induces apoptosis and demonstrated the pro-apoptotic effects of R8-PNA-a222. Remarkably, increased pro-apoptotic effects were obtained with the co-administration of both anti-miR‑221 and anti-miR‑222 PNAs. In addition, co-administration of R8-PNA-a221 and R8-PNA-a222 induced apoptosis of TMZ-treated T98G cells at a level higher than that obtained following singular administration of R8-PNA-a221 or R8-PNA-a222.
... A modified NA analog is a non-RNase-H inducing 2 -O-methyl oligonucleotide (2 -OMe), useful to conserve the target RNA intact in splicing redirection [51]. PNA (peptide nucleic acid) molecules, which are neutral oligonucleotide analogs, are considered potential antisense drugs, and could be excellent candidates for gene therapy in AIDS patients [85] or in redirecting splicing [25]. Morpholino oligomers or phosphorodiamidate morpholino oligos (PMOs) are antisense nucleic acid analogs frequently used to modulate pre-mRNA splicing by originating a steric antisense blockage without activating RNase-H. ...
... Recent approaches have focused on development of chimeric molecules (peptide nucleic acid, PNA) involving back bone of repeating N-(2-aminoethyl)-glycine units linked by peptide bonds. The nucleic acid bases (purines and pyrimidines complimentary to the conserved base sequences of LTR, U5PBS,TAR, stem or loop regions of HIV-1 genome) are linked to the backbone by a methylene bridge (-CH2-) and a carbonyl group (-(C=O)-) which may specifically mimic viral replication without emergence of drug resistant mutants [26,27]. Apart from suitable retroviral drug development approaches, other strategies such as increasing drug adherence, conducting drug resistance genotyping, the frequent monitoring of patients and drug resistance surveillance, switching the classes of retrovirals in response to resistance and selection of best drugs combination may be employed to combat this issue effectively. ...
Copyright: © 2014 Sharma B, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Editorial According to an estimate of WHO, 35.3 million people were living with HIV/AIDS (PLWH) globally at the end of 2012 which included about 0.8% of adults aged 15-49 years. According to (NACO) of India, the prevalence of AIDS in India in 2013 was 0.27 million. In low and middle income countries, more than 8 million PLWH are receiving antiretroviral therapy (ART) at the end of 2011. Application of highly active antiretro viral therapy (HAART) worldwide has been able to significantly reduce the death rate of human immunodeficiency virus type 1 (HIV-1) infected individuals. However, the appearance of clinical drug resistance in AIDS patients due to nonadherance to medication (intake of antiretroviral) has been one of the primary reasons associated to chemotherapeutic and virologic failure. In addition, high rate of viral replication, appearance of heterogenous circulating viral quasispecies, infidelity in proviral cDNA synthesis as well as immunological and pharmacological facors are also associated to drug resistance [1]. The virus develops resistance when it evades the effects of the treatment of AIDS patients by antiHIV-1 drugs resulting into no effect on viral replication [2,3]. Antiviral drug resistance is defined by the presence of viral mutations that reduce drug susceptibility compared with the susceptibility of wild-type viruses. It happens because as a retrovirus, HIV employs reverse transcriptase (HIV-1RT) to synthesize a double stranded proviral DNA from its RNA genome. HIV-1RT lacks a proof reading mechanism for correcting errors made during replication of its genome and therefore HIV introduces several mutations in its newly synthesized genomic cDNA [4]. Some of these viral mutants naturally select as drug resistant variants with higher fitness thereby posing serious threat to chemotherapy of AIDS patients [3,5]. WHO has licensed six antiretroviral classes of antiHIV drugs so far which have been found to induce more than 200 mutations in the viral genome. These mutations have been reported to be associated with drug resistance with enhanced fitness: The number of total RT mutations associated with nucleoside/nucleoside reverse transcriptase inhibitor (nRTIs/NRTIs) resistance is more than 50; total number of RT mutations associated with nonnucleoside reverse transcriptase inhibitor (NNRTIs) resistance is more than 40; total mutations associated with protease inhibitor (PIs) resistance are more than 60; total integrase mutations associated with the licensed integrase strand transfer inhibitors are more than 30 and the number of gp41 mutations associated with the fusion / entry inhibitor resistance is more than 15. The mutations in the inhibitor binding sites of co-receptors such as CCR5 or CXCR4 have been reported to cause CCR5 / CXCR4 inhibitor resistance [5,6].
... Peptide nucleic acids (PNAs) have been largely used as efficient tools for alteration of gene expression [1][2][3][4][5]. PNA and PNA-based analogues have been proposed as antisense molecules targeting mRNAs, triple-helix forming molecules targeting eukaryotic gene promoters, artificial promoters, decoy molecules targeting transcription factors [4][5][6][7][8][9][10]. Recently, PNAs have been shown to be able to alter biological functions of microRNAs, both in vitro and in vivo [11][12][13][14][15][16][17]. ...
MicroRNAs are a family of small noncoding RNAs regulating gene expression by sequence-selective mRNA targeting, leading to a translational repression or mRNA degradation. The oncomiR miR-221 is highly expressed in human gliomas, as confirmed in this study in samples of low and high grade gliomas, as well in the cell lines U251, U373 and T98G. In order to alter the biological functions of miR-221, a peptide nucleic acid targeting miR-221 (R8-PNA-a221) was produced, bearing a oligoarginine peptide (R8) to facilitate uptake by glioma cells. The effects of R8-PNA-a221 were analyzed in U251, U373 and T98G glioma cells and found to strongly inhibit miR-221. In addition, the effects of R8-PNA-a221 on p27(Kip1) (a target of miR-221) were analyzed in U251 and T98G cells by RT-qPCR and by Western blotting. No change of p27(Kip1) mRNA content occurs in U251 cells in the presence of PNA-a221 (lacking the R8 peptide), whereas significant increase of p27(Kip1) mRNA was observed with the R8-PNA-a221. These data were confirmed by Western blot assay. A clear increment of p27(Kip1) protein expression in the samples treated with R8-PNA-a221 was detected. In addition, R8-PNA-a221 was found able to increase TIMP3 expression (another target of miR-221) in T98G cells. These results suggest that PNAs against oncomiRNA miR-221 might be proposed for experimental treatment of human gliomas.
