Ehrlich II –2nd World Conference on Magic Bullets

Antibacterial RNA Silencing

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Bu səhifədəki naviqasiya:

• Conclusions
• How did the MAGIC BULLETS drop the Knife out of the Surgeons Hand. GORENSTEIN A 1,3
• Background

Antibacterial RNA Silencing

GOOD L
Department of Pathoology and Infectious Diseases, Royal Veterinary College, University of London

Background: New strategies are needed to combat antibacterial resistance. Our stategy is to target bacterial genes at the RNA level as a flexible genetics tool and possibly a new strategy for drug development. RNA silencing (or antisense) technologies can effectively target essential gene transcripts to block bacterial growth and sensitise bacteria to drugs that target proteins.

Methods: We have developed short synthetic antisense peptide nucleic acids (PNA). Cell uptake is enhanced using attached cell penetrating peptides (CPPs). In a second strategy, we have developed stabilised, expressed antisense RNAs. Therefore, we are able to silence bacterial RNA using both synthetic and expressed RNA strategies. Both RNA silencing tools have been used to silence established and putative antimicrobial drug targets. The effects of RNA silencing were assessed using Northern analysis, RT-PCR and by monitoring bacterial growth and survival.

Results: Synthetic antisense PNAs kill bacteria when targeted to stringently-required essential genes in Esherichia coli, Staphylococcus aureus and Mycobacterium smegmatis. These synthetic RNA silencers accummulate in target cells and display a long post antibiotic effect in the absence of cell lysis. It is also possible to activate cell death pathways by silencing antitoxin RNAs.

Expressed antisense RNAs are effective when modified to improve stability against endogenouse RNAse. We improved stability by using a paired termini design, where the 5´and 3´transcript are complementary and form a stable dsRNA duplex in cells. This design increases the abundance of antisense transcripts by extending transcript half life. Similar to synthetic RNA silencers, expressed RNA silencers are sufficiently effective to prevent bacaterial growth when targeted against growth essential genes.

Conclusions: Synthetic and expressed RNA silencers provide complimentary new tools for antimicrobial drug development and drug mechanism of action studies. Interestingly, partial silencing showed that established and putative antimicrobial drug targets differ significantly in their stringent requirement for growth. In other words, certain target genes are particularly sensitive to RNA silencing and may also provide more sensitive drug targets.

How did the MAGIC BULLETS drop the Knife out of the Surgeon's Hand. GORENSTEIN A1,3, SOMEKH E2,3, SEROUR F1,3 1Department of Pediatric Surgery, Wolfson Medical Center, Holon, Israel; 2Pediatric Infectious/Immunology Unit, Wolfson Medical Center, Holon, Israel; 3Affiliated to Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel Background: Surgical drainage has been considered the treatment of choice in several abscess formations. We demonstrated that surgical drainage may be avoided by the use of antibiotics in certain pediatric surgical infections.. Methods: In two clinical studies among pediatric patients, we used needle aspiration combined with antibiotics as the treatment of suppurative cervical lymphadenitis and of perianal abscess referred for incision and drainage. In both cases, a local anesthetic ointment was applied for 1 hour before the aspiration and aspiration procedure was performed using a 19-gauge needle. In some cases, a conscious sedation with midazolam was used. Results: Suppurative cervical lymphadenitis: This group included 35 children aged 4 months-13 years, mean 2.2 years referred for surgical drainage of node. Antibiotics used in most patients was cloxacillin. There were no complications. Patients were followed-up for 2-6 months. None required an open drainage of cervical abscess. A complete regression of the nodes was obtained in all patients within 21 days, with no relapse or scar formation. Perianal abscess: Forty-seven infants (< 24 months of age) were treated by needle aspiration and antibiotics (gentamycin and metranidazole). A primary cure was obtained in 29 (61.7%) patients, and 16 (34%) children had an evolution toward a fistula in ano. Two infants had recurrent abscess successfully treated by the same way. Conclusions: Needle aspiration combined with antibiotics seems to be an effective and safe treatment of suppurative cervical lymphadenitis and of perianal abscess. This modality which does not require general anesthesia may avoid the painful dressing change and drain removal, and does not lead to scar formation. We believe that our studies are additional steps for the continuous effort to replace invasive procedures by Magic Bullets.

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