Jonathan O. Wright and Dr. Paul Savage, Chemistry and Biochemistry

The projects that I have worked on this past year have been focused on a novel group of antibiotics developed Dr. Savage’s lab. The antibiotics, called ceragenins or cationic steroid antibiotics (CSA’s), are molecules synthesized from cholic acid and mimic the structure and functionality of antimicrobial peptides found in nature. Although the CSA’s are much smaller than the peptides, they have similar structural aspects, a hydrophobic side as well as a cationic side, and are believed to have a similar mechanism of action. They, like the antimicrobial peptides, have been found to be effective against a broad spectrum of bacteria, Gram-positive as well as Gram-negative, but at much lower concentrations than the peptides.1 They have been found to be effective against many strains of antibiotic-resistant bacteria,2 and some of the compounds have also been found to be active against membrane-enclosed viruses, including HIV and herpes.

Recently, there has been much interest in antimicrobial peptides and similarly functioning molecules. Even though many antimicrobial peptides are found in the innate immune systems of most organisms and bacteria have been exposed to them for thousands of years, bacteria generally remain susceptible to these compounds.3 This is of much interest as strains of bacteria are becoming more and more resistant to current antibiotics. Therefore, several cationic peptide antibiotics (CPAs) have been developed in recent years. These CPAs, however, are very expensive to synthesize, and some bacteria have been able to overcome their effects by secreting proteases. The CSAs developed in our lab overcome both the expense and the protease issues, making medical applications much more realistic.

A large project I worked on throughout the past year focused the use of one of these, CSA-13, to eradicate bacterial biofilms. A current problem facing patients in many hospitals is infection relating to long-term use of central venous catheters. One study reported 4 cases of bacteremia per every 1000 catheter-days in hemodialysis patients, caused by both gram-positive and gram-negative strains-4. This bacteremia is caused by bacteria forming biofilms in the lumen of the catheters-5, which thereafter provide reservoirs of bacteria which are continually pumped into the patients’ blood streams, causing severe infection. Additionally, these biofilms provide defense for the bacteria contained within them, rendering many traditional antibiotics ineffective against the biofilm-6.

In the past, once these catheter-related infections were detected, the patients were treated with systemic antibiotics and the catheter had to be removed. However, researchers in the past years have tested the efficacy of using an antibiotic lock solution to clear the biofilm from the catheter lumen, thereby salvaging the catheter and allowing continued use. This technique involves locking a highly concentrated solution of antibiotics into the lumen of the catheter and has shown promising results-7.

Looking at these results, we felt that CSA-13 would be a good candidate for use in this technique, as it disrupts bacterial membranes and is effective regardless of the growth stage of the bacteria, and we tested this hypothesis. Running head to head with Ciprofloxacin, currently the most effective antibiotic available, we demonstrated that CSA-13 was comparable to Ciprofloxacin in eradicating gram-negative biofilms, and it performed significantly better against gram-positive biofilms, including biofilms of highly antibiotic-resistant strains of Staphylococcus aureus. These results were published in the article “Activities of Ceragenin CSA-13 Against Established Biofilms in an In Vitro Model of Catheter Decolonization” in the journal Anti-Infective Agents in Medicinal Chemistry8.

Throughout this past semester I continued work on this same project. CSA-13 on its own is somewhat toxic, causing hemolysis (destruction of red blood cells) at concentrations fairly close to the MBCs (minimum bactericidal concentrations) for several strains of bacteria. Therefore, throughout the semester we tested the possible use of Pluronic F127 to regulate the hemolytic effects of CSA-13 in solution. Our studies have shown that we can increase the MHC (minimum hemolytic concentration) through the use of the Pluronic without changing the MBC significantly. These studies are still ongoing.

In conclusion, we showed that CSA-13 is a very good candidate for use in the catheter lock solutions and for other biofilm eradication applications. Additionally, research which is still in the process of being completed has shown promising results in optimizing the use of CSA-13 for human applications.

References

1. Schmidt, E, Boswell, J, Walsh, J, Schellenberg, M, Winter, T, Chunhong, L, Allman, G, Savage, P. (2001). Activities of cholic acid-derived antimicrobial agents against multidrug-resistant bacteria. Journal of Antimicrobial Chemotherapy 47, 671-674.
2. Chin, J, Rybak, M, Cheung, C, Savage, P. (2007). Antimicrobial Activities of Ceragenins against Clinical isolates of resistant Staphylococcus aureus. Antimicrobial Agents and Chemotherapy, Apr. 2007, 1268-1273.
3. Bambeke, F, Mingeot-Leclercq, M, Stuelens, M, Tulkens, P. (2007). The bacterial envelope as a target for novel anti-MRSA antibiotics. Trends in Pharmacological Sciences 29, 124-134.
4. Marr KA, Sexton DJ & Conlon PJ et al. (1997). Catheter-related bacteremia and outcome of attempted catheter salvage in patients undergoing hemodialysis. Ann Intern Med 127, 275–280.
5. Passerini L, Lam K, Costerton JW & King EG. (1992). Biofilms on indwelling vascular catheters. Crit Care Med 20, 665–673.
6. Stewart, P, Costerton, W. (2001). Antibiotic resistance of bacteria in biofilms. The Lancet 358, 135-138.
7. Krishnasami, Z, Carlton, D, Bimbo, L, Taylor, M, Balkovetz, D, Barker, J, Allon, M. (2002). Management of hemodialysis catheter-related bacteremia with an adjunctive antibiotic lock solution. Kidney International 61, 1136–1142.
8. Pollard, J, Wright, J, Feng, Y, Geng, D, Genberg, C, Savage, P. (2009). Activities of Ceragenin CSA-13 Against Established Biofilms in an In Vitro Model of Catheter Decolonization. Anti-Infective Agents in Medicinal Chemistry 8, 290-294.