Phage-Based Therapy - A Primer

May 11, 2018

               

Antibiotic resistance is a serious concern. In the United States, methicillin-resistant S. aureus (MRSA) infections alone account for more deaths than HIV/AIDS and tuberculosis combined. Since the discovery of antibiotics, there has been a steady stream of novel antibacterial pharmaceuticals in what has been dubbed the “antibiotic pipeline.” However, due to the rate at which bacteria evolve resistance to antibiotics, there has been less commercial interest in the research and development of novel compounds.

With the growing concerns about antibiotic resistance it is common to hear phage therapy mentioned as an alternative. Phages (short for bacteriophages) are viruses that infect and, in most instances, kill bacteria. It has been said that every two days, phages kill half the bacteria on the planet.

Phage Basics

Phages make up the most abundant biological entity on Earth (there are an estimated 1031-1032 phages in the world at any given time) and play a crucial role in regulating bacterial populations. They are simple, yet incredibly diverse, non-living biological entities consisting of DNA or RNA enclosed within a protein capsid. There are two basic types of phages, virulent (lytic) and temperate (lysogenic). Lytic phages break open (lyse) and kill their bacterial hosts after replicating inside the host. This releases copies of the phages to infect more bacteria. Lysogenic phages, known as temperate phages, incorporate their DNA into the host bacteria. They do not necessarily harm the bacteria and may even impart antibiotic resistance. Temperate phages may become active under certain conditions and lyse the host cell. Some phages are capable of carrying out both lytic and lysogenic cycles. Virulent phages are the type used to treat bacterial infections.

Lytic phages work by attaching to specific receptors on a bacterial cell surface. Then they inject their genetic material into the host cell. Once inside the cell, they hijack the bacterial replication machinery to produce the next generation of phage progeny and lyse the cell. Upon reaching a critical mass of phage progeny, which can be anywhere from a few to over 1000 viral particles, depending on environmental factors, the lytic proteins become active and hydrolyze the peptidoglycan cell wall, releasing novel phage to reinitiate the lytic cycle.

Therapeutic Use

Phages do have a number of qualities that appear to make them ideal candidates for treating bacterial infections. Since phages typically bind to specific receptors on the bacterial cell surface, most phages are infectious only to the bacteria that carry their complementary receptor. This effectively determines lytic phage host range and can allow for phage therapy that targets specific pathogenic bacteria and spares large scale effect on the the microbiome of “good” bacteria.

Since lytic phages are self-replicating, a relatively small number of phages can be effective against bacterial infections (if the phages come into contact with the targeted bacteria). Theoretically, a single phage could be enough. Phages will replicate and infect their bacterial hosts until the bacteria are eliminated. At that point the phages are quickly degraded and eliminated. This quality can help eliminate concerns about using too low or too high a dose of phages to treat infections.

Other advantages include phages' ability to penetrate and disperse bacterial biofilms in some cases and low toxicity in humans. This low potential for toxicity and target specificity could allow phages to be used prophetically against specific infections.

While bacteria do develop resistance to phages, phages rapidly evolve to overcome this resistance. Though it may take many years to develop and test an antibiotic to treat resistant bacteria, phages can likely be discovered and prepared for use against resistant bacteria in weeks. As we discuss later, however, getting new products approved for use in humans would currently still require a long testing/trial period.

There are studies and evidence that phage therapy can be effective in treating bacterial infections. In one 1931 trial of phage therapy as a treatment for cholera in the Punjab region of India, a cohort of 118 control subjects and 73 experimental subjects who received phage treatment; Early phage proponent, Felix d’Herelle, observed a 90% reduction in mortality with 74 lethal outcomes in the control group and only 5 in the experimental group.

Animal models exploring the use of phages to treat certain types of infections have produced promising results. When challenged with gut-derived sepsis due to P. aeruginosa, oral administration of phage saved 66.7% of mice from mortality compared to 0% in the control group. In a hamster model of Clostridium difficile (C. difficile)-induced ileocecitis, a single dose of phage concurrent with C. difficile administration was sufficient prophylaxis against infection; phage treatments post-infection saved 11 of 12 mice whereas control animals receiving C. difficile and clindamycin died within 96 hours.

Human trials for phage therapy have taken place for almost a century at several institutes in Eastern Europe, including the Eliava Institute of Bacteriophage and the Institute of Immunology and Experimental Therapy in Wroclaw, Poland. The Eliava Institute has used phage therapy in preclinical and clinical treatment of a number of common bacterial pathogens. During a 1974 typhus outbreak 18577 children was enrolled in a prophylactic intervention trial using typhoid phages. The phage treated group had a five fold decrease in typhus incidence compared to placebo.

There are currently no FDA approved phage therapy products available in the US. However there are several FDA approved phage related products for control of bacteria in the food industry.

One route to developing phage therapies is the use of phage derived lytic proteins to kill bacteria. Two major protein classes are employed by the majority of phage species during the lysis of the bacterial host. One of which is the transmembrane protein holin and the other is a peptidoglycan cell wall hydrolase called endolysin (lysin). These two proteins work together in triggering the lysis of the bacterial cell. Since lysins alone can kill bacteria, they have received the most attention as a possible therapy.

Phage lysins have been used to treat mice with bacteremia caused by multidrug-resistant A. baumannii, Streptococcus pneumoniae, and MRSA, among others. A recent study exploring the isolation and application of phage proteins has revealed that lysins are even capable of crossing epithelial cell membranes to eliminate difficult to treat intracellular infections of S. pyogenes.

One advantage of using phage lysins as a therapy is that bacteria are unlikely to develop resistance to lysins. This is due to the fact that target sites on the peptidoglycan cell wall are critical for bacterial viability. Additionally engineered recombinant phage lytic proteins would be far easier to mass produce and administer than preparations of actual phage preparations.

Possible Problems Associated With Phage Therapy

While phages specificity is an advantage of the therapy (can kill pathogens and leave “good bacteria” unharmed), it can also be a drawback. By narrowly targeting a single pathogen, phage therapy might not be as useful in certain situations. One example might be burn wounds which are often colonized by more than one strain of bacteria. Also, there are situations where the strain of bacteria causing an infection might not be immediately known. This can sometimes be addressed using phage “cocktails” of different phages to target multiple targets.

Large scale production and distribution of phages in some situations might not be effective. This is due to the nature of bacteria which evolved significant differences in difference geographic regions. In one study a Russian phage cocktail used against E.Coli was unsuccessful in treating 180 Bangladeshi children with confirmed enterotoxogenic E. coli diarrhea.

Another issue is that bacteria can rapidly develop resistance to phages the same way they develop resistance to antibiotics. Fortunately, phages evolve rapidly to overcome resistance. The problem is that developing approved therapies is very time consuming and expensive. This can be a difficult barrier to overcome given phages' specificity.

Phages are recognized by the immune system as foreign. They are fairly rapidly removed from the system. This can make delivery of the phage to the infection challenging.

Conclusions

There is a very real need to develop new treatments for bacterial infections. Phages offer an important route to developing new treatments. However there are significant challenges to developing phages for large scale use. The development of phage derived lysins for treating bacterial infections is one of the most promising routes to developing a useful therapy.

Some Companies Developing Phage-Based Products

  • Omnilytics They have offerings in the Agriculture, Food and Water, Industrial and Contract Manufacturing markets.

  • AmpliPhi Biosciences (APHB) Developing phage products for human treatment. They have two products in development.   

  • Epibiome Developing phage cocktails for human treatment.        

  • Intralytix Intralytix is developing phage based products in the industrial, food safety, environmental sanitation, veterinary,and human therapeutic markets.    

  • Bluephage Developing phage detection solutions that use phage detection as indicators of     the likelihood of other viruses being present.    

  • Phage International     Associated with the well known Eliava Institute, in Tbilisi, Republic of Georgia. They create phage therapies to treat human infections.

  • Enbiotix Developing engineered phages to treat infections in humans.    

  • GangaGen Developing phage derived lysin therapy for infections in humans.    

  • C3J Therapeutics Developing antimicrobials using synthetic phages and peptides.
  • Phagelux Utilizes phages, lysins and other biologics and related delivery technologies to create antibacterial products and solutions.
  • Biomx Develops phages to target microbiome bacteria







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