Potential of Azadirachta indica as a Capping Agent for Antiviral Nanoparticles against SARS-CoV-2.

A rare type of pneumonia later on referred to as COVID-19 was reported in China in December 2019. Investigations revealed that this disease is caused by a coronavirus previously identified as SARS-CoV-2, and since then, it has become a global pandemic with new strains emerging rapidly as a result of genetic mutations. Various therapeutic options are being explored in order to eradicate this pandemic even though approved vaccine candidates are being currently rolled out globally. Most medicinal plant extracts have astonishing properties, and they can therefore be used in the biosynthesis of effective antiviral nanoparticles. In this systematic review, we aimed to highlight the specific attributes that make Azadirachta indica (neem plant) a suitable candidate for the biosynthesis of anti-SARS-CoV-2 nanoparticles. A systematic investigation was therefore carried out in PubMed, Scopus, Web of Science, and AJOL databases with the keywords "Nanoparticles," "Biosynthesis," "Antivirals," "SARS-CoV-2," and "Azadirachta indica." 1216 articles were retrieved by the 21st of February 2022, but we screened studies that reported data on biomedical and antimicrobial assessment of Azadirachta indica extracts. We also screened studies that were reporting nanoparticles possessing antiviral properties against SARS-C0V-2, narrowing our results to 98 reports. Herein, the SARS-CoV-2 viral structure is briefly discussed with nanoparticles of biomedical importance in the design of SARS-CoV-2 antivirals. Most importantly, we focused on the biomedical and antiviral properties of Azadirachta indica extracts that could be of importance in the design of potential anti-SARS-CoV-2 nanoformulations.

Applications of Nanoparticles for Herpes Simplex Virus (HSV) and Human Immunodeficiency Virus (HIV) Treatment.

Human Immunodeficiency Virus (HIV) is a global pandemic that has contributed to the burden of disease, and the synergistic interaction between Herpes Simplex Virus (HSV) and HIV has assisted further in the spread of the HIV disease. Moreover, several chemotherapeutic treatment options from antiviral monotherapy to highly active antiretroviral therapy (HAART) have been adopted to manage the infection; however, HIV has developed new mechanisms against these active pharmaceutical agents (APAs), limiting the effect of the drugs. In this article, we reviewed different nanoparticles and their antiviral potency against HSV and HIV infection as well as the effect of drug encapsulated nanoparticles using different drug delivery systems as they palliate to some flaws or deficiencies that the stand-alone drugs present. Drug encapsulated nanoparticles show better treatment outcomes of HSV and HIV infection. The nanoparticles can transverse the anatomic privilege sites to exert their therapeutic effect, and a prolonged and higher dose of the encapsulated therapeutic agent can ease the dosage frequency, thus palliating low drug compliance which the stand-alone drugs fail to perform. Therefore, it is clear that nanoparticles prevent antiviral drug resistance by maintaining sustained drug release over an extended period, improving the therapeutic effect of the entrapped drug

Expressed anti-HBV primary microRNA shuttles inhibit viral replication efficiently in vitro and in vivo.

The use of RNA interference (RNAi) to inhibit gene expression is potentially applicable in the treatment of viral infections such as hepatitis B virus (HBV) persistence. Although efficient HBV gene silencing by short hairpin RNA (shRNA) expressed from RNA polymerase (Pol) III promoters has been reported, constitutive high-level transcription may cause harmful side effects. Here, we report an approach that allows the use of a Pol II promoter to improve transcription regulation of expressed RNAi effecters. Pol II [cytomegalovirus (CMV)] or Pol III (U6) promoter cassettes that transcribe anti-HBV primary microRNA (pri-miR)-122 and pri-miR-31 shuttles were generated. In cultured cells both types of pri-miR-like sequences effected knockdown of markers of viral replication (>80%) and were processed to form intended 21-nucleotide guides. The concentration of CMV-expressed miRs was approximately 85-fold lower than the U6 shRNA-derived guide RNA. When cells were co-transfected with pri-miR expression cassettes, attenuation of independent RNAi-mediated gene silencing was not observed, which is in contrast to the action of U6 shRNA expression cassettes. The efficacy of the anti-HBV pri-miR shuttles in vivo was verified using the murine hydrodynamic injection model. Employing Pol II-expressed pri-miR mimics may be useful in the treatment of HBV infection, and potentially also for generic application in RNAi-based therapy.

Efficient inhibition of hepatitis B virus replication in vivo, using polyethylene glycol-modified adenovirus vectors.

Achieving safe delivery of anti-hepatitis B virus (HBV) RNA interference (RNAi) effectors is an important objective of this gene-silencing technology. Adenoviruses (Ads) have a natural tropism for the liver after systemic administration, and are useful for delivery of expressed anti-HBV RNAi sequences. However, a drawback of Ad vectors is diminished efficacy and toxicity that results from stimulation of innate and adaptive immunity. To attenuate these effects we used monomethoxy polyethylene glycol-succinimidyl propionate (mPEG-SPA) to modify first-generation vectors that express an anti-HBV RNAi effector. Efficient hepatocyte transduction and knockdown of HBV replication were achieved after intravenous administration of 5 x 10(9) PEGylated or native recombinant Ads to HBV transgenic mice. After the first injection, circulating HBV viral particle equivalents (VPEs) remained low for 3 weeks and began to increase after 5 weeks. A second dose of PEGylated anti-HBV Ad caused a less sustained decrease in circulating VPEs, but no silencing after a second dose was observed in animals treated with unmodified vector. Release of inflammatory cytokines, including monocyte chemoattractant protein-1 (MCP-1), interferon-gamma, interleukin-6, and tumor necrosis factor-alpha, was elevated in animals receiving unmodified vectors. However, only a modest increase in MCP-1 was observed in mice that received a second dose of PEG Ads. Also, polymer-conjugated vectors induced a weaker adaptive immune response and were less hepatotoxic than their unmodified counterparts. Collectively, these observations show that PEG modification of Ads expressing RNAi effectors improves their potential for therapeutic application against HBV infection.

Effective anti-hepatitis B virus hammerhead ribozymes derived from multimeric precursors.

Endonucleolytic hammerhead ribozymes have advantages of inhibiting gene expression by acting specifically, independently of cellular pathways, and within all cell compartments. However, there are concerns about inefficient silencing because of reduced intracellular cleavage of target RNA by ribozymes. To enable production of defined single-unit ribozymes and thereby increase effectiveness, we developed self-cleaving multimeric cassettes that generate several trans-acting ribozyme units from a single transcript. cis and trans ribozyme cleavage, as assessed in vitro against three different sites within the X sequence of hepatitis B virus (HBV), occurred efficiently and precisely according to predictions deduced from the ribozyme designs. Significant knockdown of markers of viral replication in transfected cultured liver-derived cells was achieved by multiribozyme Pol II expression cassettes. To assess silencing efficacy of RNA prepared in vitro, transcription and cis cleavage reactions were carried out to prepare defined single-unit ribozymes. Transfection of ribozyme RNA was capable of inhibiting HBV surface antigen secretion from liver-derived cells without associated elevation of interferon-alpha or interferon-beta secretion into the culture upernatants. The approach described here is potentially useful for several applications, such as generation of RNA interference (RNAi) effectors, which require rapid and inexpensive generation of defined RNA sequences.

Specific inhibition of HBV replication in vitro and in vivo with expressed long hairpin RNA.

Activating RNA interference to achieve specific gene silencing has shown promise for the development of RNA-based treatment of chronic hepatitis B virus (HBV) infection. To further this approach, we assessed the efficacy of expressed long hairpin RNAs (lhRNAs) that target the conserved HBx open reading frame of HBV. As substrates for Dicer, lhRNAs have the potential to generate multiple short interfering RNAs (siRNAs) to enable simultaneous targeting of different sites. Two U6 Pol III vectors were constructed that encode anti-HBV lhRNAs with a 62 base pair stem sequence containing multiple G:U pairings. Assessment in transfected cultured cells and also in vivo using the murine hydrodynamic injection model showed that one of the lhRNA vectors (lhRNA 1) diminished markers of virus replication by 70-90% without evidence of interferon response induction. Greatest silencing efficacy was observed for targets that are complementary to sequences located at the base of the hairpin stem and this correlated with a higher concentration of siRNAs derived from this region of the lhRNA. Although lhRNA 1 has the advantage of targeting a greater viral sequence, incomplete cellular processing may result in unequal silencing across the span of the viral target RNA.