Applications of phages
Apart from their applications in the field of medicine and therapy, phages are being employed in a variety of other areas.
As drug delivery vehicles
Delivering therapeutic materials or imaging reagents into specific tissues or target organs is vitally important for the development of novel therapeutic methods or the early diagnosis of various critical diseases. The primary goal of drug delivery in medicine is to develop a carrier of specific therapeutic genes, proteins or chemicals to desired targets of interest with high specificity. The structural features of phages, such as their possession of genomic and proteomic information in the same body and their ability to be genetically engineered to accommodate various target genes and proteins, make them very useful in the design of novel drug delivery carriers. The main point here is that, engineering phages will require only the genome to be modified as per the requirement, and these genomic alterations will be directly reflected in the phenotype of the virus. Phages are used in gene therapy, drug targeting and target imaging. In gene therapy, it is essential that a gene is delivered efficiently to a specific tissue or cell type. (Poul M. A. and Marks J. D., 1999) Phages can be used to achieve this specificity by engineering targeting peptide sequences on their outer coats. Phages are also good gene delivery vectors because they protect DNA from degradation. (Clark J. R. and March J. B., 2006) One way in which phages have been used for gene therapy is as a vehicle for DNA vaccinations. DNA vaccinations are more advantageous than protein vaccinations as they are easier to make and have fewer side effects. However, naked DNA is quickly degraded once administered. In one study, the lambda phage was used as a vehicle for DNA and the results showed that antibody responses were achieved when the phage vehicle was used, as compared to the naked DNA injection.
The tissue- and cell-type specificity of phages is also beneficial for drug targeting; therapeutics can be targeted specifically to diseased tissue, thus reducing toxic effects on healthy tissue. Another application is to use tagged phages (labels or nanoparticles) to target the disease tissue for imaging. (Li K. et al., 2010)
But there are several challenges of this application of phages. Although several human studies have been successful at identifying target-homing peptides, most phage display studies in humans can only utilize a single round of phage-library selection per patient due to the immune responses associated with phage delivery. Repeated rounds of selection must be done in new patients, for whom target peptides may differ. Because of this limitation, it is often difficult to correctly characterize a single target peptide sequence. But in spite of all the challenges, phages are being considered seriously for delivery vehicles.
In tissue engineering
The field of tissue engineering is focused on the creation of desired artificial tissue or organ scaffold structures for biomedical applications, such as tissue or organ replacement, disease modelling and drug testing. Consequently, tissue engineering research requires combining biomaterials and cells in an intricate matrix to closely mimic in vivo tissue environments. Due to the vast number of factors controlling cell behaviour, engineering a suitable biomimetic matrix is a complicated task. Chemical, physical and mechanical properties have to be finely tuned. The vast number of signalling peptides in biological systems makes choosing the optimal chemical cues very difficult. Similarly, there are many variables influencing the physical features of a matrix, including pore size, scaffold alignment and other general microscopic features. Finally, mechanical features, such as substrate stiffness, can influence cell behaviour.
Natural tissue microenvironments are composed of networks of various nanofibrous proteins, such as collagen or fibronectin. Therefore, the predominant viral types used in tissue engineering applications are filament-shape viruses, such as the M13 phage, the turnip yellow mosaic virus (TYMV) and the tobacco mosaic virus (TMV). M13 phages have several characteristics that make them an ideal engineering tool: their ability to display peptides on their pIII, pVIII and pIX outer coats, their ability to replicate in large quantities, and their ability to self-assemble into nanofilaments. Hence, phages can be used to make scaffolds for tissue engineering, without having to worry much about biocompatibility and assembly.
In one study, when the neuronal progenitor cells were cultured on the phage tissue engineering substrates, enhanced neural progenitor cell proliferation and differentiation, with good viability, was observed. The phage scaffolds in fact guided the growth of neural cells. This makes it a potential cure for spinal injury repair. (Merzlyak A., Indrakanti S. and Lee S. W., 2009) Phage scaffolds are being explored for bone tissue culturing also, despite the fact that these type of cultures require sturdy scaffolds.
For waste water treatment
Pathogen removal from sewage water can be done by using phage cocktails. This approach does not harm the environment, does not require expensive equipment or sophisticated techniques. In one study, hospital wastewater was collected from different locations of Tamil Nadu and it was found that it had high concentrations of antibiotic resistant E. coli, Pseudomonas sp. Streptococcus sp and Bacillus spp. which clearly indicated the extent of pollution. Inoculation of the samples with specific phages resulted in complete removal of pathogen from the water within 14 hours. (Periasamy, D. and Sundaram A., 2013)
For food preservation
The food industry already employs phages to limit and control bacterial growth that eventually leads to spoilage of food. Several phage products are, in fact, commercially available for use in the food industry. One company, Intralytix, markets several products, including a six-phage cocktail, ListShield™, against Listeria monocytogenes. This phage cocktail is sprayed onto foods before packaging, and has been proven safe and effective against this harmful bacterium. In another study, When P100 was applied to contaminated cheese, there was significant reduction or complete eradication of L. monocytogenes. Toxicity tests were also conducted and no negative side effects were seen in the rat models. (Carlton, R. et al., 2005) Finally, of notable significance, in 2005, OmniLytics Inc. registered its phage product with the US Environmental Protection Agency. Its product is used to treat bacterial infections on tomato and pepper plants. Due to the positive impact phage therapy has thus far had on the food industry, research and development efforts are being continued.
As drug delivery vehicles
Delivering therapeutic materials or imaging reagents into specific tissues or target organs is vitally important for the development of novel therapeutic methods or the early diagnosis of various critical diseases. The primary goal of drug delivery in medicine is to develop a carrier of specific therapeutic genes, proteins or chemicals to desired targets of interest with high specificity. The structural features of phages, such as their possession of genomic and proteomic information in the same body and their ability to be genetically engineered to accommodate various target genes and proteins, make them very useful in the design of novel drug delivery carriers. The main point here is that, engineering phages will require only the genome to be modified as per the requirement, and these genomic alterations will be directly reflected in the phenotype of the virus. Phages are used in gene therapy, drug targeting and target imaging. In gene therapy, it is essential that a gene is delivered efficiently to a specific tissue or cell type. (Poul M. A. and Marks J. D., 1999) Phages can be used to achieve this specificity by engineering targeting peptide sequences on their outer coats. Phages are also good gene delivery vectors because they protect DNA from degradation. (Clark J. R. and March J. B., 2006) One way in which phages have been used for gene therapy is as a vehicle for DNA vaccinations. DNA vaccinations are more advantageous than protein vaccinations as they are easier to make and have fewer side effects. However, naked DNA is quickly degraded once administered. In one study, the lambda phage was used as a vehicle for DNA and the results showed that antibody responses were achieved when the phage vehicle was used, as compared to the naked DNA injection.
The tissue- and cell-type specificity of phages is also beneficial for drug targeting; therapeutics can be targeted specifically to diseased tissue, thus reducing toxic effects on healthy tissue. Another application is to use tagged phages (labels or nanoparticles) to target the disease tissue for imaging. (Li K. et al., 2010)
But there are several challenges of this application of phages. Although several human studies have been successful at identifying target-homing peptides, most phage display studies in humans can only utilize a single round of phage-library selection per patient due to the immune responses associated with phage delivery. Repeated rounds of selection must be done in new patients, for whom target peptides may differ. Because of this limitation, it is often difficult to correctly characterize a single target peptide sequence. But in spite of all the challenges, phages are being considered seriously for delivery vehicles.
In tissue engineering
The field of tissue engineering is focused on the creation of desired artificial tissue or organ scaffold structures for biomedical applications, such as tissue or organ replacement, disease modelling and drug testing. Consequently, tissue engineering research requires combining biomaterials and cells in an intricate matrix to closely mimic in vivo tissue environments. Due to the vast number of factors controlling cell behaviour, engineering a suitable biomimetic matrix is a complicated task. Chemical, physical and mechanical properties have to be finely tuned. The vast number of signalling peptides in biological systems makes choosing the optimal chemical cues very difficult. Similarly, there are many variables influencing the physical features of a matrix, including pore size, scaffold alignment and other general microscopic features. Finally, mechanical features, such as substrate stiffness, can influence cell behaviour.
Natural tissue microenvironments are composed of networks of various nanofibrous proteins, such as collagen or fibronectin. Therefore, the predominant viral types used in tissue engineering applications are filament-shape viruses, such as the M13 phage, the turnip yellow mosaic virus (TYMV) and the tobacco mosaic virus (TMV). M13 phages have several characteristics that make them an ideal engineering tool: their ability to display peptides on their pIII, pVIII and pIX outer coats, their ability to replicate in large quantities, and their ability to self-assemble into nanofilaments. Hence, phages can be used to make scaffolds for tissue engineering, without having to worry much about biocompatibility and assembly.
In one study, when the neuronal progenitor cells were cultured on the phage tissue engineering substrates, enhanced neural progenitor cell proliferation and differentiation, with good viability, was observed. The phage scaffolds in fact guided the growth of neural cells. This makes it a potential cure for spinal injury repair. (Merzlyak A., Indrakanti S. and Lee S. W., 2009) Phage scaffolds are being explored for bone tissue culturing also, despite the fact that these type of cultures require sturdy scaffolds.
For waste water treatment
Pathogen removal from sewage water can be done by using phage cocktails. This approach does not harm the environment, does not require expensive equipment or sophisticated techniques. In one study, hospital wastewater was collected from different locations of Tamil Nadu and it was found that it had high concentrations of antibiotic resistant E. coli, Pseudomonas sp. Streptococcus sp and Bacillus spp. which clearly indicated the extent of pollution. Inoculation of the samples with specific phages resulted in complete removal of pathogen from the water within 14 hours. (Periasamy, D. and Sundaram A., 2013)
For food preservation
The food industry already employs phages to limit and control bacterial growth that eventually leads to spoilage of food. Several phage products are, in fact, commercially available for use in the food industry. One company, Intralytix, markets several products, including a six-phage cocktail, ListShield™, against Listeria monocytogenes. This phage cocktail is sprayed onto foods before packaging, and has been proven safe and effective against this harmful bacterium. In another study, When P100 was applied to contaminated cheese, there was significant reduction or complete eradication of L. monocytogenes. Toxicity tests were also conducted and no negative side effects were seen in the rat models. (Carlton, R. et al., 2005) Finally, of notable significance, in 2005, OmniLytics Inc. registered its phage product with the US Environmental Protection Agency. Its product is used to treat bacterial infections on tomato and pepper plants. Due to the positive impact phage therapy has thus far had on the food industry, research and development efforts are being continued.