Anti-virulence therapeutics

 

Microbes actively interact with the human immune system in very sophisticated ways to cause disease.  The ability of a pathogen to cause disease in a person is dependent on several interrelated activities and functions of each, including the virulence of the microbe (its capability to cause disease) and the relative degree of resistance or susceptibility of the individual to the particular pathogen.  Often these interactions strongly determine the success or failure of a pathogen to inhabit humans.  Interestingly, the human body contains more microbial cells than human cells and living in peaceful coexistence with our microbial environments is essential to good health.  Improved knowledge about the regulation of these pathogen-host interactions will help determine exactly how the microbial world interacts in both favorable and unfavorable ways with human cells and particularly, the immune system.  Read More

The MvfR system and infochemicals

Arabidopsis thaliana is a widely used model organism in plant biology

Scanning electron microscopy of PA interacting with surface structures of Arabidopsis thaliana

The overall success of a pathogen ultimately to cause disease in humans depends on the efficacy of its virulence factors, anti-immune weapons, and the immune status of the human host.  Pseudomonas aeruginosa or PA is a common bacterium that causes serious diseases in both animals and humans.  Because of its ability to readily develop resistance to multiple antibiotics and its ability to produce acute and chronic infections, MGH investigators have studied PA extensively and these studies have provided novel insights into the regulation and signaling of molecules or chemicals that likely contribute to treatment failure and mortality in PA infections.

The MGH investigators have identified key bacterial genes and regulatory networks that control many virulence functions in PA and other Gram-negative bacterial pathogens.  Of key interest to MGH investigators are the “quorum sensing” (QS) networks that bacteria use to sense and respond to both external and internal bacterial cell signals and environmental cues.  Bacteria use QA networks to adapt and alter gene expression for their survival and persistence in patients.  Investigators have identified at least three QA systems in PA that are required to cause full virulence and one of these is the “multiple virulence factor regulator” or MvfR.

The MvfR system controls bacterial cell-to-cell communication and cooperative behavior via the production of specific small-excreted, low molecular weight molecules that act as “infochemicals”.  Some infochemicals are important mediators of inter-species, intra-species and inter-kingdom communications and disease development, while others promote host tolerance, or cancer in mammals.

&copy 2011 Que et al. Originally published in Journal of Pathogens. 2011:549302. doi: 10.4061/2011/549302

Muscle sample with myonecrosis with proliferating rods. Inset shows intramuscular bacteria

MGH investigators studying infochemical communication have uncovered the unique role of the MvfR system in QA and its importance in PA pathogenesis.  The investigators identified for the first time the existence of a small QS-regulated molecule called 2-AA (2-amino acetophenone) produced by PA that reduces bacterial virulence in vivo in two model organisms of acute infection.  Evidence shows that 2-AA produces immune-modulatory effects that enhance the host’s ability to tolerate a pathogen’s presence while silencing acute bacterial virulence functions.  The innovative models of infection used to interrogate the immuno-inflammatory and metabolic effects promoted by virulence factors enhance the MGH investigators’ abilities to discover how these infochemicals work in humans.  In fact, MGH investigators looking for the presence of infochemicals produced by PA in human wound infections, for the first time, were able to detect and quantify one of the QS signaling molecules from Pseudomonas acute wound infections in burn patients.  The HAQ molecule not only is measurable directly from an infected patient, but the ratio between the various HAQ molecule types resembles that found in the mouse models of burn injury and infection.  These results strongly suggest the importance of these animal models in the evaluation of the efficacy of new drugs.  The ability to follow bacterial pathogenesis is an important first step for the development of anti-virulence agents aimed at controlling the course of infection.

Antibiotic tolerant cells or “persisters”

Multidrug tolerance or antibiotic tolerance is a unique challenge worldwide to medical care.  Unlike the phenomena of multidrug resistance caused by mutations of the pathogen, antibiotic tolerance is created when a small subpopulation of microbial cells called “persisters” exist in a dormant, transient, and not-dividing state.  Persisters are not antibiotic resistant mutations, but do survive the antibiotic killing of their surrounding non-persister cells.

MGH investigators studying the mechanism of persister cell formation in PA have developed techniques for the isolation and quantification of these transient and infrequent cells.  Their results have shown that the molecule 2-AA affects the transcription of translation-related genes, thereby affecting the cell’s translational capacity, resulting in the formation of persisters.  Transcription and translation are part of the process of gene expression for translating a gene into a protein.  Knowing this, the investigators believe that QS inhibition may be an excellent target to stop the progression of or prevent multi-drug resistant PA infections.  Investigators are working to identify and test anti-virulence compounds for the treatment of multi-drug resistance and antibiotic tolerance while potentially preserving beneficial microbes.

Identifying susceptibility to infection after burn injury

MGH investigators are also examining the many factors known to predispose patients with burn injury to infection, including the impairment of immune responses.  It is well know that patients with severe burns are highly susceptible to bacterial infection, primarily from the disruption of the physical skin barrier.  The investigators are studying gene expression in the muscle, blood, fat, and skin of patients and how expression changes in the presence of a burn wound and infection.  Investigators hope to identify how these gene changes affect a patient’s susceptibility or resistance to infection.  Such clues could help identify which patients have a particularly high risk of infection after suffering a burn injury. This knowledge will help expanding this approach to also other types of trauma. Knowing this predisposition would allow clinicians to tailor individual treatment plans to promote the patient’s natural local and systemic defensive responses to injury and ensuing infection.

Relevant publications

Starkey M, Lepine F, Maura D, Bandyopadhaya A, Lesic B, He J, et al. Identification of anti-virulence compounds that disrupt quorum-sensing regulated acute and persistent pathogenicity. PLoS Pathog. 2014 Aug; 10(8):e1004321. PubMed PMID: 25144274; PubMed Central PMCID: PMC4140854

Bandyopadhaya A, Kesarwani M, Que YA, He J, Padfield K, Tompkins R, et al.  The quorum sensing volatile molecule 2-amino acetophenon modulates host immune responses in a manner that promotes life with unwanted guests. PLoS Pathog. 2012; 8(11):e1003024. PubMed PMID: 23166496; PubMed Central PMCID: PMC3499575

Tzika AA, Constantinou C, Bandyopadhaya A, Psychogios N, Lee S, Mindrinos M, et al. A small volatile bacterial molecule triggers mitochondrial dysfunction in murine skeletal muscle. PLoS One. 2013; 8(9):e74528. PubMed PMID: 24098655; PubMed Central PMCID: PMC3787027

Dulcey CE, Dekimpe V, Fauvelle DA, Milot S, Groleau MC, Doucet N, et al. The end of an old hypothesis: the pseudomonas signaling molecules 4-hydroxy-2-alkylquinolines derive from Fatty acids, not 3-ketofatty acids. Chem Biol. 2013 Dec 19; 20(12):1481-91.  PubMed PMID: 24239007; PubMed Central PMCID: PMC3877684

Bangi E, Pitsouli C, Rahme LG, Cagan R, Apidianakis Y. Immune response to bacteria induces dissemination of Ras-activated Drosophila hindgut cells. EMBO Rep. 2012 Jun; 13(6):569-76. PubMed PMID: 22498775; PubMed Central PMCID: PMC3367237

Kesarwani M, Hazan R, He J, Que YA, Apidianakis Y, Lesic B,  et al. A quorum sensing regulated small volatile molecule reduces acute virulence and promotes chronic infection phenotypes. PLoS Pathog. 2011 Aug;7(8):e1002192. PubMed PMID: 21829370; PubMed Central PMCID: PMC3150319

Hazan R, Maura D, Que YA, Rahme LG. Assessing pseudomonas aeruginosa persister/antibiotic tolerant cells. Methods Mol Biol. 2014; 1149:699-707. PubMed PMID: 24818944

Que YA, Hazan R, Strobel B, Maura D, He J, Kesarwani M, et al. A quorum sensing small volatile molecule promotes antibiotic tolerance in bacteria. PLoS One. 2013; 8(12):e80140. PubMed PMID: 24367477; PubMed PMCID: PMC3868577

Apidianakis Y, Que YA, Xu W, Tegos GP, Zimniak P, Hamblin MR, et al. Down-regulation of glutatione S-transferase a 4 (hGSTA4) in the muscle of thermally injured patients is indicative of susceptibility to bacterial infection. FASEB J. 2012 Feb; 26(2):730-7. PubMed PMID: 22038048; PubMed Central PMCID: PMC3290433

Yan S, Tsurumi A, Que YA, Ryan CM, Bandyopadhaya A, Morgan AA,  et al. Prediction of multiple infections after severe burn trauma: a prospective cohort study. Ann Surg. 2014 Jun 19.  PubMed PMID: 24950278; PubMed Central PMCID: PMC4284150

Que YA, Hazan R, Ryan CM, Milot S, Lépine F, Lydon M, et al. Production of pseudomonas aeruginosa intercellular small signaling molecules in human burn wounds. J Pathog. 2011;2011:549302. PubMed PMID: 23533774

Hazan R, He J, Xiao G, Dekimpe V, Apidianakis Y, Lesic B, et al. Homeostatic interplay between bacterial cell-cell signaling and iron in virulence. PLoS Pathog. 2010 Mar; 6(3):e1000810. PubMed PMID: 20300606; PubMed Central PMCID:PMC2837411

Lépine F, Dekimpe V, Lesic B, Milot S, Lesimple A, Mamer OA, et al. PqsA is required for the biosynthesis of 2,4-dihydroxyquinoline (DHQ), a newly identified metabolite produced by Pseudomonas aeruginosa and Burkholderia thailandensis. Biol Chem. 2007 Aug; 388(8):839-45. PubMed PMID:17655503

Déziel E, Lépine F, Milot S, He J, Mindrinos MN, Tompkins RG, et al.   Analysis of Pseudomonas aeruginosa 4-hydroxy-2-alkylquinolines (HAQs) reveals a role for 4-hydroxy-2-heptylquinoline in cell-to-cell communication. Proc Natl Acad Sci U S A. 2004 Feb 3; 101(5):1339-44. PubMed PMID: 14739337; PubMed Central PMCID: PMC337054

He J, Baldini RL, Déziel E, Saucier M, Zhang Q, Liberati NT, et al. The broad host range pathogen Pseudomonas aeruginosa strain PA14 carries two pathogenicity islands harboring plant and animal virulence genes. Proc Natl Acad Sci U S A. 2004 Feb 24; 101(8):2530-5.  PubMed PMID: 14983043; PubMed Central PMCID: PMC356984

Rahme LG, Ausubel FM, Cao H, Drenkard E, Goumnerov BC, Lau GW, et al. Plants and animals share functionally common bacterial virulence factors. Proc Natl Acad Sci U S A. 2000 Aug 1; 97(16):8815-21. PubMed PMID: 10922040; PubMed Central PMCID: PMC34017

Rahme LG, Tan MW, Le L, Wong SM, Tompkins RG, Calderwood SB, et al. Use of model plant hosts to identify Pseudomonas aeruginosa virulence factors. Proc Natl Acad Sci U S A. 1997 Nov 25; 94(24):13245-50.  PubMed PMID: 9371831; PubMed Central PMCID: PMC24294

Mahajan-Miklos S, Tan MW, Rahme LG, Ausubel FM. Molecular mechanisms of bacterial virulence elucidated using a Pseudomonas aeruginosa-Caenorhabditis elegans pathogenesis model. Cell. 1999 Jan 8; 96(1):47-56. PubMed PMID: 9989496

Rahme LG, Stevens EJ, Wolfort SF, Shao J, Tompkins RG, Ausubel FM. Common virulence factors for bacterial pathogenicity in plants and animals. Science. 1995 Jun 30; 268(5219):1899-902. PubMed PMID: 7604262

Contact

Laurence Rahme, Ph.D.
rahme@molbio.mgh.harvard.edu
617-724-5003

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