Phage therapy

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Phage injecting its genome into bacterial cell
An electron micrograph of bacteriophages attached to a bacterial cell. These viruses are the size and shape of coliphage T1

Phage therapy or viral phage therapy is the therapeutic use of bacteriophages to treat pathogenic bacterial infections.[1] Although extensively used and developed mainly in former Soviet Union countries circa 1920, the treatment is not approved in countries other than Russia and Georgia. Phage therapy has many potential applications in human medicine as well as dentistry, veterinary science, and agriculture.[2] If the target host of a phage therapy treatment is not an animal the term "biocontrol" (as in phage-mediated biocontrol of bacteria) is usually employed, rather than "phage therapy".

Bacteriophages are much more specific than antibiotics, so they can hypothetically be chosen to be indirectly harmless not only to the host organism (human, animal, or plant), but also to other beneficial bacteria, such as gut flora, reducing the chances of opportunistic infections.[3] They would have a high therapeutic index, that is, phage therapy would be expected to give rise to few side effects. Because phages replicate in vivo, a smaller effective dose can be used. On the other hand, this specificity is also a disadvantage: a phage will only kill a bacterium if it is a match to the specific strain. Consequently phage mixtures are often applied to improve the chances of success, or samples can be taken and an appropriate phage identified and grown.

Phages are currently being used therapeutically to treat bacterial infections that do not respond to conventional antibiotics, particularly in Russia[4] and Georgia.[5][6][7] They tend to be more successful than antibiotics where there is a biofilm covered by a polysaccharide layer, which antibiotics typically cannot penetrate.[8] In the West, no therapies are currently authorized for use on humans, although phages for killing food poisoning bacteria (Listeria) are now in use.[9]

History[edit]

Phage in action on Bacillus anthracis

Following the discovery of bacteriophages by Frederick Twort and Felix d'Hérelle[10] in 1915 and 1917, phage therapy was immediately recognized by many to be a key way forward for the eradication of bacterial infections. A Georgian, George Eliava, was making similar discoveries. He travelled to the Pasteur Institute in Paris where he met d'Hérelle, and in 1923 he founded the Eliava Institute in Tbilisi, Georgia, devoted to the development of phage therapy.

In neighbouring countries including Russia, extensive research and development soon began in this field. In the United States during the 1940s commercialization of phage therapy was undertaken by the large pharmaceutical company, Eli Lilly.

While knowledge was being accumulated regarding the biology of phages and how to use phage cocktails correctly, early uses of phage therapy were often unreliable.[11] When antibiotics were discovered in 1941 and marketed widely in the U.S. and Europe, Western scientists mostly lost interest in further use and study of phage therapy for some time.[12]

Isolated from Western advances in antibiotic production in the 1940s, Russian scientists continued to develop already successful phage therapy to treat the wounds of soldiers in field hospitals. During World War II, the Soviet Union used bacteriophages to treat many soldiers infected with various bacterial diseases e.g. dysentery and gangrene. Russian researchers continued to develop and to refine their treatments and to publish their research and results. However, due to the scientific barriers of the Cold War, this knowledge was not translated and did not proliferate across the world.[13][14] A summary of these publications was published in English in 2009 in "A Literature Review of the Practical Application of Bacteriophage Research" [15]

There is an extensive library and research center at the George Eliava Institute in Tbilisi, Georgia. Phage therapy is today a widespread form of treatment in that region.[citation needed]

As a result of the development of antibiotic resistance since the 1950s and an advancement of scientific knowledge, there has been renewed interest worldwide in the ability of phage therapy to eradicate bacterial infections and chronic polymicrobial biofilm (including in industrial situations[16]).

Phages have been investigated as a potential means to eliminate pathogens like Campylobacter in raw food[17] and Listeria in fresh food or to reduce food spoilage bacteria.[18] In agricultural practice phages were used to fight pathogens like Campylobacter, Escherichia and Salmonella in farm animals, Lactococcus and Vibrio pathogens in fish from aquaculture and Erwinia and Xanthomonas in plants of agricultural importance. The oldest use was, however, in human medicine. Phages have been used against diarrheal diseases caused by E. coli, Shigella or Vibrio and against wound infections caused by facultative pathogens of the skin like staphylococci and streptococci. Recently the phage therapy approach has been applied to systemic and even intracellular infections and the addition of non-replicating phage and isolated phage enzymes like lysins to the antimicrobial arsenal. However, actual proof for the efficacy of these phage approaches in the field or the hospital is not available.[18]

Some of the interest in the West can be traced back to 1994, when Soothill demonstrated (in an animal model) that the use of phages could improve the success of skin grafts by reducing the underlying Pseudomonas aeruginosa infection.[19] Recent studies have provided additional support for these findings in the model system.[20]

Although not "phage therapy" in the original sense, the use of phages as delivery mechanisms for traditional antibiotics constitutes another possible therapeutic use.[21][22] The use of phages to deliver antitumor agents has also been described in preliminary in vitro experiments for cells in tissue culture.[23]

Potential benefits[edit]

Bacteriophage treatment offers a possible alternative to conventional antibiotic treatments for bacterial infection.[24] It is conceivable that, although bacteria can develop resistance to phage, the resistance might be easier to overcome than resistance to antibiotics.[25] Just as bacteria can evolve resistance, viruses can evolve to overcome resistance;[26] however, the ability to evolve raises serious safety questions.[16]

Bacteriophages are very specific, targeting only one or a few strains of bacteria.[27] Traditional antibiotics have more wide-ranging effect, killing both harmful bacteria and useful bacteria such as those facilitating food digestion. The specificity of bacteriophages might reduce the chance that useful bacteria are killed when fighting an infection.

Some evidence shows the ability of phages to travel to a required site—including the brain, where the blood brain barrier can be crossed—and multiply in the presence of an appropriate bacterial host, to combat infections such as meningitis. However the patient's immune system can, in some cases, mount an immune response to the phage (2 out of 44 patients in a Polish trial[28]).

A few research groups in the West are engineering a broader spectrum phage, and also a variety of forms of MRSA treatments, including impregnated wound dressings, preventative treatment for burn victims, phage-impregnated sutures.[29] Enzybiotics are a new development at Rockefeller University that create enzymes from phage. These show potential for preventing secondary bacterial infections, e.g. pneumonia developing in patients suffering from flu and otitis.[citation needed] Purified recombinant phage enzymes can be used as separate antibacterial agents in their own right.[30]

For some bacteria, such as multiple-resistant Klebsiella pneumoniae, there are no effective non-toxic antibiotics, but killing of this bacteria via intraperitoneal, intravenous, or intranasal route of phages in vivo has been shown to work in laboratory tests.[31]

Application[edit]

Collection[edit]

The simplest method of phage treatment involves collecting local samples of water likely to contain high quantities of bacteria and bacteriophages, for example effluent outlets, sewage and other sources.[5] They can also be extracted from corpses.[citation needed] The samples are taken and applied to the bacteria that are to be destroyed which have been cultured on growth medium.

If the bacteria die, as usually happens, the mixture is centrifuged; the phages collect on the top of the mixture and can be drawn off.

The phage solutions are then tested to see which ones show growth suppression effects (lysogeny) or destruction (lysis) of the target bacteria. The phage showing lysis are then amplified on cultures of the target bacteria, passed through a filter to remove all but the phages, then distributed.

Treatment[edit]

Phages are "bacterium-specific" and it is therefore necessary in many cases to take a swab from the patient and culture it prior to treatment. Occasionally, isolation of therapeutic phages can require a few months to complete, but clinics generally keep supplies of phage cocktails for the most common bacterial strains in a geographical area.

Phages in practice are applied orally, topically on infected wounds or spread onto surfaces, or used during surgical procedures. Injection is rarely used, avoiding any risks of trace chemical contaminants that may be present from the bacteria amplification stage, and recognizing that the immune system naturally fights against viruses introduced into the bloodstream or lymphatic system.

The direct human use of phages is likely to be very safe; in August and October 2006, the United States Food and Drug Administration approved spraying meat and cheese with phages. The approval was for ListShield and Listex (phage preparations targeting Listeria monocytogenes). These were the first approvals granted by the FDA and UDSA for phage-based applications. This confirmation of safety within the worldwide scientific community opened the way for other phages applications for example against Salmonella and E-coli (http://www.fsis.usda.gov/wps/portal/fsis/topics/regulations/directives/fsis-directives)

Phage therapy has been attempted for the treatment of a variety of bacterial infections including: laryngitis, skin infections, dysentery, conjunctivitis, periodontitis, gingivitis, sinusitis, urinary tract infections and intestinal infections, burns, boils,[5] poly-microbial biofilms on chronic wounds, ulcers and infected surgical sites.[citation needed]

In 2007 a Phase 1/2 clinical trial was completed at the Royal National Throat, Nose and Ear Hospital, London for Pseudomonas aeruginosa infections (otitis).[32][33][34] Documentation of the Phase-1/Phase-2 study was published in August 2009 in the journal Clinical Otolaryngology.[35]

Phase 1 clinical trials have now been completed in the Southwest Regional Wound Care Center, Lubbock, Texas for an approved cocktail of phages against bacteria, including P. aeruginosa, Staphylococcus aureus and Escherichia coli (better known as E. coli).[36] The cocktail of phages for the clinical trials was developed and supplied by Intralytix.

Reviews of phage therapy indicate that more clinical and microbiological research is needed to meet current standards.[37]

Administration[edit]

Phages can usually be freeze-dried and turned into pills without materially impacting efficiency.[5] Temperature stability up to 55 °C and shelf lives of 14 months have been shown for some types of phages in pill form.[citation needed]

Application in liquid form is possible, stored preferably in refrigerated vials.[citation needed]

Oral administration works better when an antacid is included, as this increases the number of phages surviving passage through the stomach.[citation needed]

Topical administration often involves application to gauzes that are laid on the area to be treated.[citation needed]

Obstacles[edit]

General[edit]

The high bacterial strain specificity of phage therapy may make it necessary for clinics to make different cocktails for treatment of the same infection or disease because the bacterial components of such diseases may differ from region to region or even person to person.

In addition, due to the specificity of individual phages, for a high chance of success, a mixture of phages is often applied. This means that 'banks' containing many different phages must be kept and regularly updated with new phages.[3]

Further, bacteria can evolve different receptors either before or during treatment; this can prevent the phages from completely eradicating the bacteria.

The need for banks of phages makes regulatory testing for safety harder and more expensive. Such a process would make it difficult for large-scale production of phage therapy. Additionally, patent issues (specifically on living organisms) may complicate distribution for pharmaceutical companies wishing to have exclusive rights over their "invention", making it unlikely that a for-profit corporation will invest capital in the widespread application of this technology.

As has been known for at least thirty years, mycobacteria such as Mycobacterium tuberculosis have specific bacteriophages.[38] No lytic phage has yet been discovered for Clostridium difficile, which is responsible for many nosocomial diseases, but some temperate phages (integrated in the genome) are known for this species, which opens encouraging avenues.

To work, the virus has to reach the site of the bacteria, and viruses do not necessarily reach the same places that antibiotics can reach.

Funding for phage therapy research and clinical trials is generally insufficient and difficult to obtain, since it is a lengthy and complex process to patent bacteriophage products. Scientists comment that 'the biggest hurdle is regulatory', whereas an official view is that individual phages would need proof individually because it would be too complicated to do as a combination, with many variables. Due to the specificity of phages, phage therapy would be most effective with a cocktail injection, which is generally rejected by the U.S. Food and Drug Administration (FDA). Researchers and observers predict that for phage therapy to be successful the FDA must change its regulatory stance on combination drug cocktails.[3] Public awareness and education about phage therapy are generally limited to scientific or independent research rather than mainstream media.[39]

The negative public perception of viruses may also play a role in the reluctance to embrace phage therapy.[40]

Safety[edit]

Approval of phage therapy for use in humans has not been given in Western countries. Much of the problem is how to prove safety when using a self-replicating entity which has the capability to evolve.[16]

As with antibiotic therapy and other methods of countering bacterial infections, endotoxins are released by the bacteria as they are destroyed within the patient (Herxheimer reaction). This can cause symptoms of fever, or in extreme cases toxic shock (a problem also seen with antibiotics) is possible.[41] Janakiraman Ramachandran[42] argues that this complication can be avoided in those types of infection where this reaction is likely to occur by using genetically engineered bacteriophages which have had their gene responsible for producing endolysin removed. Without this gene the host bacterium still dies but remains intact because the lysis is disabled. On the other hand this modification stops the exponential growth of phages, so one administered phage means one dead bacterial cell.[7] Eventually these dead cells are consumed by the normal house-cleaning duties of the phagocytes, which utilise enzymes to break the whole bacterium and its contents down into harmless proteins, polysaccharides and lipids.[43]

Lysogenic bacteriophages are not generally used therapeutically, as this group can act as a way for bacteria to exchange DNA; this can help spread antibiotic resistance or even, theoretically, make the bacteria pathogenic (see Cholera).

Effectiveness[edit]

In Russia, mixed phage preparations may have a therapeutic efficacy of 50%. This equates to the complete cure of 50 of 100 patients with terminal antibiotic-resistant infection. The rate of only 50% is likely to be due to individual choices in admixtures and ineffective diagnosis of the causative agent of infection.[44]

Other Animals[edit]

Brigham Young University is currently researching the use of phage therapy to treat American foulbrood in honeybees.[45]

Cultural references[edit]

See also[edit]

External links[edit]

References[edit]

  1. ^ Silent Killers: Fantastic Phages?, by David Kohn, CBS News: 48 Hours Mystery.
  2. ^ McAuliffe et al. "The New Phage Biology: From Genomics to Applications" (introduction) in Mc Grath, S. and van Sinderen, D. (eds.) Bacteriophage: Genetics and Molecular Biology Caister Academic Press ISBN 978-1-904455-14-1.reprint
  3. ^ a b c Keen, E. C. (2012). "Phage Therapy: Concept to Cure". Frontiers in Microbiology 3. doi:10.3389/fmicb.2012.00238. PMC 3400130. PMID 22833738. 
  4. ^ "Eaters of bacteria: Is phage therapy ready for the big time?". Discover Magazine. Retrieved 12 April 2013. 
  5. ^ a b c d BBC Horizon: Phage - The Virus that Cures 1997-10-09
  6. ^ Parfitt T (2005). "Georgia: an unlikely stronghold for bacteriophage therapy". Lancet 365 (9478): 2166–7. doi:10.1016/S0140-6736(05)66759-1. PMID 15986542. 
  7. ^ a b Thiel, Karl (January 2004). "Old dogma, new tricks—21st Century phage therapy". Nature Biotechnology (London UK: Nature Publishing Group) 22 (1): 31–36. doi:10.1038/nbt0104-31. PMID 14704699. Retrieved 2007-12-15. 
  8. ^ Aguita, Maria. "Combatting Bacterial Infection". LabNews.co.uk. Retrieved 2009-05-05. 
  9. ^ Pirisi A (2000). "Phage therapy—advantages over antibiotics?". Lancet 356 (9239): 1418. doi:10.1016/S0140-6736(05)74059-9. PMID 11052592. 
  10. ^ Shasha SM, Sharon N, Inbar M (2004). "[Bacteriophages as antibacterial agents]". Harefuah (in Hebrew) 143 (2): 121–5, 166. PMID 15143702. 
  11. ^ Kutter, E; De Vos, D, Gvasalia, G, Alavidze, Z, Gogokhia, L, Kuhl, S, Abedon, ST (January 2010). "Phage therapy in clinical practice: treatment of human infections". Current pharmaceutical biotechnology 11 (1): 69–86. doi:10.2174/138920110790725401. PMID 20214609. 
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  18. ^ a b Mc Grath S and van Sinderen D (editors). (2007). Bacteriophage: Genetics and Molecular Biology (1st ed.). Caister Academic Press. ISBN 978-1-904455-14-1. [1]. 
  19. ^ Soothill JS (1994). "Bacteriophage prevents destruction of skin grafts by Pseudomonas aeruginosa". Burns 20 (3): 209–11. doi:10.1016/0305-4179(94)90184-8. PMID 8054131. 
  20. ^ McVay CS, Velásquez M, Fralick JA (2007). "Phage therapy of Pseudomonas aeruginosa infection in a mouse burn wound model". Antimicrob. Agents Chemother. 51 (6): 1934–8. doi:10.1128/AAC.01028-06. PMC 1891379. PMID 17387151. 
  21. ^ Yacoby I, Bar H, Benhar I (2007). "Targeted drug-carrying bacteriophages as antibacterial nanomedicines". Antimicrob. Agents Chemother. 51 (6): 2156–63. doi:10.1128/AAC.00163-07. PMC 1891362. PMID 17404004. 
  22. ^ Yacoby I, Shamis M, Bar H, Shabat D, Benhar I (2006). "Targeting antibacterial agents by using drug-carrying filamentous bacteriophages". Antimicrob. Agents Chemother. 50 (6): 2087–97. doi:10.1128/AAC.00169-06. PMC 1479106. PMID 16723570. 
  23. ^ Bar H, Yacoby I, Benhar I (2008). "Killing cancer cells by targeted drug-carrying phage nanomedicines". BMC Biotechnol. 8: 37. doi:10.1186/1472-6750-8-37. PMC 2323368. PMID 18387177. 
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  29. ^ Scientists Engineer Viruses To Battle Bacteria : NPR
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  36. ^ Rhoads DD, Wolcott RD, Kuskowski MA, Wolcott BM, Ward LS, Sulakvelidze A (2009). Bacteriophage therapy of venous leg ulcers in humans: results of a phase I safety trial. J Wound Care. 2009 Jun;18(6):237-8, 240-3.
  37. ^ Brüssow, H (2005). "Phage therapy: The Escherichia coli experience". Microbiology (Reading, England) 151 (Pt 7): 2133–40. doi:10.1099/mic.0.27849-0. PMID 16000704. 
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  41. ^ Evergreen PHAGE THERAPY: BACTERIOPHAGES AS ANTIBIOTICS
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  44. ^ http://files.school-collection.edu.ru/dlrstore/378ba3de-4e8f-dadd-e84f-4cf4f0b632f1/11-15_03_2002.pdf
  45. ^ https://www.youtube.com/watch?v=rj9_QGBJN0w
  46. ^ Summers WC (1991). "On the origins of the science in Arrowsmith: Paul de Kruif, Felix d'Herelle, and phage". J Hist Med Allied Sci 46 (3): 315–32. doi:10.1093/jhmas/46.3.315. PMID 1918921. 
  47. ^ "Phage Findings - TIME". Time. 1938-01-03. Retrieved 2007-12-13. 
  48. ^ "SparkNotes: Arrowsmith: Chapters 31–33". Retrieved 2007-12-13.