Anti-infectives: Targeting SARS
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686 | SEPTEMBER 2004 | VOLUME 2 www.nature.com/reviews/micro So far, many different classes of antimicrobial peptides have been identified in mammals, including α-, β- and θ-defensins and catheli- cidins. For more information on defensins in primates, see the review by Robert Lehrer on page 727 of this issue. Hornef et al. were interested in a family of mouse-intestine-specific CRS (cryptdin-related sequence) peptides. CRS mRNAs have been characterized previously but the peptide products have never been characterized before. Using mRNA from intestinal tissue of healthy mice, Hornef et al. identified 15 new cDNAs encoding CRS peptides, bringing the total number of CRS peptide coding sequences to 23, comparable to the number of mouse α-defensin genes. Each CRS gene encodes a peptide containing a pro-region that is cleaved off during processing, leaving a mature cationic peptide containing nine conserved cysteines. Of the 15 new sequences identified, most of the sequence variation observed was in the pro-region, and so a total of 7 mature peptides were predicted by the coding sequences. To screen for mature CRS peptides in vivo, whole mouse intestinal tissue was fractionated by reverse-phase HPLC. Analysis of the mass of fractions in the presence and absence of a reducing agent indicated the presence of dimers. To exclude the possibility that dimer formation was an experimental artefact, the extraction process was repeated in the pres- ence of an alkylating agent that inhibits the formation of disulphide bonds. This con- firmed that CRS peptides form covalent dimers in vivo. Six mature peptides were iden- tified in a variety of homo- and heterodimers and, additionally, four peptides were also found in truncated form. Examination of the antibacterial activity of synthetic CRS dimers showed these agents had potent activity against bacterial species including Salmonella enterica serovar Typhimurium and Streptococcus pyogenes, with no cytotoxicity. The complement of antimicrobial peptides varies from species to species. This study has revealed that in mice, the complement of CRS peptides matches that of the α-defensins in number, and perhaps also in potency. The abil- ity to form hetero- and homodimers is a neat trick to expand the repertoire of intestinal pep- tide diversity and strengthen antimicrobial defence. Sheilagh Clarkson References and links ORIGINAL RESEARCH PAPER Hornef, M. W. et al. Increased diversity of intestinal antimicrobial peptides by covalent dimer formation. Nature Immunol. 5, 836–843 (2004) A new study published in the Proceedings of the National Academy of Sciences (USA) has identified 15 compounds that are potent against the severe acute respiratory syndrome virus (SARS-CoV) in cell-based assays, including two drugs that are currently in use and several that are already in the clinical development pipeline. The only available therapy for SARS-CoV is a combination of ribavirin and corticosteroids, which is effective — but only at concentrations that produce severe side effects. Since SARS was first isolated in 2002, it has caused more than 700 deaths worldwide, so finding effective antivirals for SARS is a priority. High-throughput screening requires a simple and reliable assay system. Wu et al. used a cell-based assay to screen more than 10,000 compounds, all administered at 10 µM, for inhibition of virus cytopathic effects. From this primary screen, 50 compounds were selected for further analysis, first to determine if the compounds were cytotoxic and second to measure the concentration of the compound required to inhibit virus replication by 50% (EC 50 ). The most potent inhibitor was valinomycin, a peptide insecticide with an EC 50 of 0.85 µM. Aescin and reserpine, already in use in Europe and the United States, respectively, were also active against SARS-CoV, but neither drug is currently prescribed as an antiviral. Compounds that are structurally similar to reserpine and aescin were also screened, resulting in 10 new anti-SARS active compounds. Finally, a potent inhibitor of the HIV protease, TL-3, which is being developed as a treatment for AIDS in cats, was also effective against the SARS protease. These studies offer new hope for rapidly available SARS therapeutics. Susan Jones References and links ORIGINAL RESEARCH PAPER Wu, C.-Y. et al. Small molecules targeting severe acute respiratory syndrome human coronavirus. Proc. Natl Acad. Sci. USA 101, 10012–10017 (2004) FURTHER READING Stadler, K. et al. SARS — beginning to understand a new virus. Nature Rev. Microbiol. 1, 209–218 (2003) WEB SITE Wong laboratory: http://wong.scripps.edu/ Targeting SARS ANTI-INFECTIVES Diverse defence INNATE IMMUNITY RESEARCH HIGHLIGHTS A report published in Nature Immunology presents the first detailed characterization of a new family of antimicrobial peptides in the mouse intestine. Antimicrobial peptides are small, endoge- nous peptides containing <100 residues and are an important component of innate immunity.
Transcript of Anti-infectives: Targeting SARS
686 | SEPTEMBER 2004 | VOLUME 2 www.nature.com/reviews/micro
So far, many different classes of antimicrobialpeptides have been identified in mammals,including α-, β- and θ-defensins and catheli-cidins. For more information on defensins inprimates, see the review by Robert Lehrer onpage 727 of this issue.
Hornef et al. were interested in a family ofmouse-intestine-specific CRS (cryptdin-relatedsequence) peptides. CRS mRNAs have beencharacterized previously but the peptideproducts have never been characterized before.Using mRNA from intestinal tissue of healthymice, Hornef et al. identified 15 new cDNAsencoding CRS peptides, bringing the totalnumber of CRS peptide coding sequences to23, comparable to the number of mouse α-defensin genes. Each CRS gene encodes apeptide containing a pro-region that is cleavedoff during processing, leaving a mature cationicpeptide containing nine conserved cysteines.Of the 15 new sequences identified, most ofthe sequence variation observed was in thepro-region, and so a total of 7 mature peptideswere predicted by the coding sequences.
To screen for mature CRS peptides in vivo,whole mouse intestinal tissue was fractionatedby reverse-phase HPLC. Analysis of the massof fractions in the presence and absence of areducing agent indicated the presence of
dimers. To exclude the possibility that dimerformation was an experimental artefact, theextraction process was repeated in the pres-ence of an alkylating agent that inhibits theformation of disulphide bonds. This con-firmed that CRS peptides form covalentdimers in vivo. Six mature peptides were iden-tified in a variety of homo- and heterodimersand, additionally, four peptides were alsofound in truncated form. Examination of theantibacterial activity of synthetic CRS dimersshowed these agents had potent activity againstbacterial species including Salmonella entericaserovar Typhimurium and Streptococcus pyogenes,with no cytotoxicity.
The complement of antimicrobial peptidesvaries from species to species. This study hasrevealed that in mice, the complement of CRSpeptides matches that of the α-defensins innumber, and perhaps also in potency. The abil-ity to form hetero- and homodimers is a neattrick to expand the repertoire of intestinal pep-tide diversity and strengthen antimicrobialdefence.
Sheilagh Clarkson
References and linksORIGINAL RESEARCH PAPER Hornef, M. W. et al. Increaseddiversity of intestinal antimicrobial peptides by covalent dimerformation. Nature Immunol. 5, 836–843 (2004)
A new study published in the Proceedingsof the National Academy of Sciences (USA)has identified 15 compounds that arepotent against the severe acute respiratorysyndrome virus (SARS-CoV) in cell-basedassays, including two drugs that arecurrently in use and several that arealready in the clinical developmentpipeline.
The only available therapy for SARS-CoVis a combination of ribavirin andcorticosteroids, which is effective — butonly at concentrations that produce severeside effects. Since SARS was first isolated in2002, it has caused more than 700 deathsworldwide, so finding effective antivirals forSARS is a priority.
High-throughput screening requires asimple and reliable assay system. Wu et al.used a cell-based assay to screen more than10,000 compounds, all administered at 10µM, for inhibition of virus cytopathiceffects. From this primary screen, 50compounds were selected for furtheranalysis, first to determine if the compoundswere cytotoxic and second to measure the
concentration of the compound required toinhibit virus replication by 50% (EC
50).
The most potent inhibitor wasvalinomycin, a peptide insecticide with anEC
50of 0.85 µM. Aescin and reserpine,
already in use in Europe and the UnitedStates, respectively, were also active againstSARS-CoV, but neither drug is currentlyprescribed as an antiviral. Compounds thatare structurally similar to reserpine andaescin were also screened, resulting in 10new anti-SARS active compounds. Finally, apotent inhibitor of the HIV protease, TL-3,which is being developed as a treatment forAIDS in cats, was also effective against theSARS protease. These studies offer new hopefor rapidly available SARS therapeutics.
Susan Jones
References and linksORIGINAL RESEARCH PAPER Wu, C.-Y. et al. Smallmolecules targeting severe acute respiratory syndromehuman coronavirus. Proc. Natl Acad. Sci. USA 101,10012–10017 (2004) FURTHER READING Stadler, K. et al. SARS — beginningto understand a new virus. Nature Rev. Microbiol. 1, 209–218(2003)WEB SITEWong laboratory: http://wong.scripps.edu/
Targeting SARS
A N T I - I N F E C T I V E S
Diverse defence
I N N AT E I M M U N I T Y
R E S E A R C H H I G H L I G H T S
A report published in Nature Immunologypresents the first detailed characterization of anew family of antimicrobial peptides in themouse intestine.
Antimicrobial peptides are small, endoge-nous peptides containing <100 residues and arean important component of innate immunity.
