Design, synthesis and biological evaluation of TAR and cTAR binders as HIV-1 nucleocapsid inhibitors

Sosic, Alice; Frecentese, Francesco; Perissutti, Elisa; Sinigaglia, Laura; Santagada, Vincenzo; Caliendo, Giuseppe; Magli, Elisa; Ciano, Antonio; Zagotto, Giuseppe; Parolin, Cristina; Gatto, Barbara

doi:10.1039/c3md00212h

Cited by 18 publications

(86 citation statements)

References 40 publications

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“…Bell and colleagues hypothesized that the specific ability of Gag polyprotein, rather than mature NC, could effect structural changes in SL3 which may aid in genomic viral RNA packaging into the virus particle though avoiding binding by prematurely cleaved NC (Bell et al, 2013). This is in contrast with the significant chaperoning of NC alone on the TAR/cTAR oligonucleotides (Godet and Mely, 2010; Levin et al, 2010; Shvadchak et al, 2009; Sosic et al, 2013). Unlike SL3, the TAR/cTAR chaperoning is required during reverse transcription, which requires mature NC after cell infection.…”

Section: Rna Bindersmentioning

confidence: 80%

“…Compounds that bind to TAR can interfere with this important NC interaction, contributing to an antiviral effect. Indeed, a recent study characterizing the effect of positively charged anthraquinone derivatives, compound 13 (Fig 5), on TAR and cTAR melting and hybridization in presence of NC demonstrates that targeting these sequences inhibits the NC chaperoning function in vitro (Sosic et al, 2013). …”

Section: Rna Bindersmentioning

confidence: 99%

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Advances in targeting nucleocapsid–nucleic acid interactions in HIV-1 therapy

Garg

Torbett

2014

Virus Research

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The continuing challenge of HIV-1 treatment resistance in patients creates a need for the development of new antiretroviral inhibitors. The HIV nucleocapsid (NC) protein is a potential therapeutic target. NC is necessary for viral RNA packaging and in the early stages of viral infection. The high level of NC amino acid conservation among all HIV-1 clades suggests a low tolerance for mutations. Thus, NC mutations that could arise during inhibitor treatment to provide resistance may render the virus less fit. Disruption of NC function provides a unique opportunity to strongly dampen replication at multiple points during the viral life cycle with a single inhibitor. Although NC exhibits desirable features for a potential antiviral target, the structural flexibility, size, and the presence of two zinc fingers makes small molecule targeting of NC a challenging task. In this review, we discuss the recent advances in strategies to develop inhibitors of NC function and present a perspective on potential novel approaches that may help to overcome some of the current challenges in the field.

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Section: Rna Bindersmentioning

confidence: 80%

Section: Rna Bindersmentioning

confidence: 99%

Advances in targeting nucleocapsid–nucleic acid interactions in HIV-1 therapy

Garg

Torbett

2014

Virus Research

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“…14 Moreover, the increase in melting temperature correlates with the inhibition of the annealing catalyzed by HIV-1 NC, leading to the reduced formation of the heteroduplex TAR/cTAR and indicating that threading intercalators are an interesting class of binders for dynamic structures of RNA such as the TAR element. 14 The mechanism of action invoked for these compounds can be further validated by performing the NAME assay in different alternative formats, as shown in Figure 4: in the case of intercalators the inhibition of annealed heteroduplex is enhanced when the drug is incubated with the two folded oligos. Compounds acting with different mechanism of action, such as direct binders of the NC protein, would be less affected by the different preincubation mode.…”

Section: Discussionmentioning

confidence: 99%

“…Controls are: folded cTAR, folded TAR, and annealed hybrid (TAR/cTAR), obtained through thermal denaturation of the stably folded structures of TAR to cTAR followed by their slow annealing. 14 The ability of NC to denature two stable nucleic acid sequences into the extended heteroduplex helix is clearly evident: the full-length NC protein leads immediately to the formation of the TAR/cTAR hybrid: 0' indicates the minimal time passing between addition of NC to the samples and addition of gel loading buffer, which stops the reaction; complete formation is already achieved after 15' incubation time. The increase in the intensity of the annealed heteroduplex parallels the decrease in the intensity of the folded cTAR and TAR oligonucleotides.…”

Section: Nucleocapsid Annealing-mediated Electrophoresis (Name) Assaymentioning

confidence: 99%

Nucleocapsid Annealing-Mediated Electrophoresis (NAME) Assay Allows the Rapid Identification of HIV-1 Nucleocapsid Inhibitors

Sosic

Cappellini

Scalabrin³

et al. 2015

JoVE

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RNA or DNA folded in stable tridimensional folding are interesting targets in the development of antitumor or antiviral drugs. In the case of HIV-1, viral proteins involved in the regulation of the virus activity recognize several nucleic acids. The nucleocapsid protein NCp7 (NC) is a key protein regulating several processes during virus replication. NC is in fact a chaperone destabilizing the secondary structures of RNA and DNA and facilitating their annealing. The inactivation of NC is a new approach and an interesting target for anti-HIV therapy. The Nucleocapsid Annealing-Mediated Electrophoresis (NAME) assay was developed to identify molecules able to inhibit the melting and annealing of RNA and DNA folded in thermodynamically stable tridimensional conformations, such as hairpin structures of TAR and cTAR elements of HIV, by the nucleocapsid protein of HIV-1. The new assay employs either the recombinant or the synthetic protein, and oligonucleotides without the need of their previous labeling. The analysis of the results is achieved by standard polyacrylamide gel electrophoresis (PAGE) followed by conventional nucleic acid staining. The protocol reported in this work describes how to perform the NAME assay with the full-length protein or its truncated version lacking the basic N-terminal domain, both competent as nucleic acids chaperones, and how to assess the inhibition of NC chaperone activity by a threading intercalator. Moreover, NAME can be performed in two different modes, useful to obtain indications on the putative mechanism of action of the identified NC inhibitors.

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“…The 5′ UTR includes the transactivation responsive element (TAR), the poly A tract, the primer binding site (PBS), and four stem-loops (SL1-SL4) that make up the packaging domain. TAR has been targeted using anthraquinone compounds in in vitro studies [142]. A variety of peptides that disrupt viral binding to NC have been reported [143–146].…”

Section: Inhibitors Targeting Gag Assembly Processes and Maturationmentioning

confidence: 99%

HIV-1 Gag as an Antiviral Target: Development of Assembly and Maturation Inhibitors

Spearman¹

2015

CTMC

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HIV-1 Gag is the master orchestrator of particle assembly. The central role of Gag at multiple stages of the HIV lifecycle has led to efforts to develop drugs that directly target Gag and prevent the formation and release of infectious particles. Until recently, however, only the catalytic site protease inhibitors have been available to inhibit late stages of HIV replication. This review summarizes the current state of development of antivirals that target Gag or disrupt late events in the retrovirus lifecycle such as maturation of the viral capsid. Maturation inhibitors represent an exciting new series of antiviral compounds, including those that specifically target CA-SP1 cleavage and the allosteric integrase inhibitors that inhibit maturation by a completely different mechanism. Numerous small molecules and peptides targeting CA have been studied in attempts to disrupt steps in assembly. Efforts to target CA have recently gained have considerable momentum from the development of small molecules that bind CA and alter capsid stability at the post-entry stage of the lifecycle. Efforts to develop antivirals that inhibit incorporation of genomic RNA or to inhibit late budding events remain in preliminary stages of development. Overall, the development of novel antivirals targeting Gag and the late stages in HIV replication appears much closer to success than ever, with the new maturation inhibitors leading the way.

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Design, synthesis and biological evaluation of TAR and cTAR binders as HIV-1 nucleocapsid inhibitors

Cited by 18 publications

References 40 publications

Advances in targeting nucleocapsid–nucleic acid interactions in HIV-1 therapy

Advances in targeting nucleocapsid–nucleic acid interactions in HIV-1 therapy

Nucleocapsid Annealing-Mediated Electrophoresis (NAME) Assay Allows the Rapid Identification of HIV-1 Nucleocapsid Inhibitors

HIV-1 Gag as an Antiviral Target: Development of Assembly and Maturation Inhibitors

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