Bacteria as a drug vector?

From the beginning of our kill switch design, we foresaw the application of the project being used within a drug delivery context. As intracellular AMP production builds, protegrin-1 would slowly integrate into the bacterial phospholipids bilayer, causing pores to form between the membrane and the extracellular matrix. We believe that the pressure difference between the inside and the outside of the cell would cause the intracellular contents of the cell to be exported upon pore formation. This means that along with the cytosol, the potential drug will also be released into the extracellular matrix.

Why use a bacterium as a drug vector? Isn’t using a pill a much safer and cost-effective way to absorb chemicals into the body? Capsules have been used since their creation in 1834, as they allow the medicine to be contained within functionally-important media, such as oil, and in pre-measured amounts and combinations to assist with accurate dosing. A capsule simply contains medicine within a stable shell coating, which will dissolve within the body and allow the uptake of the contained chemical into the system. Pills are the current norm within drug delivery, but they are far from the most effective way to deliver medicine to the body.

The advantage that bacterial drug vectors have over pills is the natural specificity that bacteria have towards it preferred ecological niche. Escherichia coli, for instance, move through the body after being ingested and colonize the lower intestines. They have evolved over millions of years, and trillions of replications, to survive within the environment of the gut, evading host defenses, gathering nutrients, and reproducing within the natural parameters of the lower intestine. If there was a drug that needed to be directly delivered to the gut, E. coli may offer a hyper-specific delivery vector that could outshine pill counterparts.

One of the obvious benefits of site-specific drug delivery would be the pronounced reduction in unwanted systemic effects of useful, functional drugs. For instance, catabolic steroids that would benefit a single organ must currently be taken via injection or oral means, resulting in the possibility of a whole range of detrimental side effects including hypertension and weight gain (caused by water retention), decreased muscle mass, cataracts, and a decrease in infection sensitivity and response associated with long term use. Rather than the site of absorption be into the bloodstream, which spreads the steroids to internal areas where they can cause these side effects, site-specific steroid release would greatly reduce excess steroid concentrations in the blood. Not only that, our cells would need to release a much lower concentration of drug than required in pill form. We would no longer have to flood the bloodstream with a high concentration of drug so the intended site in effected, and this would reduce the severity of, or perhaps even remove altogether, some of the drugs side effects. Another example would be the ingestion of antibiotics used to fight other infections within the body. High dose broad-spectrum antibiotics would normally damage gut flora before being absorbed into the bloodstream. Our E. coli vectors would be able to release this drug not only in small doses, but to smaller areas within the gut, resulting in far lower collateral damage to symbiotic bacteria. The bacterial ability to move to the site of drug delivery may result in a more effective drug absorbance by the area in need, and less damage to the rest of the body, resulting in a more effective treatment.

SPACE

It is also important to mention that the AMPs providing the functionality of our kill switch could themselves be the “drug” that must be delivered. If the drug needed were for attacking other invasive prokaryotic bacteria in the body, redesigning the promoter and ribosome-binding site of our kill switch would lead to a higher PoPS output, resulting in faster AMP production. In theory, if enough AMPs could be produced before complete membrane disruption, intracellular non-membrane bound AMPs (and, perhaps, membrane-bound AMPs as well) would also be exported from the cell upon pore formation. Using peptides instead of antibiotics to fight other bacterial infections reduces the odds of creating antibiotic resistance. AMPs attack the membrane, the structure of which is highly conserved within evolution; the number of mutations required in a single replication to completely re-structure the bacterial membrane makes the odds of developing AMP resistance near impossible. Not only that, but the AMPs would not attack host eukaryotic cells due to the structure of their membrane (as described in our Kill Switch page), instead binding to only prokaryotic cell membranes, nullifying any potential negative interaction between the drug and the host.

There are, however, dangerous factors involved in using live bacteria as a drug delivery system. The act of releasing the drug from the burst cell also involves releasing the rest of intracellular contents, including chemicals which may indirectly act as stimuli, and could illicit potentially detrimental responses from the body. One concern we’ve raised before is the release of endotoxins imbedded in the phospholipid membrane. While the word “endotoxin” in a general sense can describe any toxin generated within the bacteria, gram-negative endotoxins are more particular. These are found specifically within the outer layer of the phospholipid bilayer and are responsible for the disease-causing aspect of gram-negative toxicity. Release of these endotoxins induce an inflammatory response by the body, causing macrophages and endothelial cells to secrete pro-inflammatory cytokines and nitric oxide. Septic shock is dangerous, and the problem of endotoxin production would have to be addressed before any in vivo testing could occur. However, this is not beyond the reach of future scientific advances, as the endotoxin molecules are produced by DNA just as malleable as our kill switch. Synthetic biology would need to advance to a point where non-endotoxic bacteria could be engineered before bacterial vectors would be safe for in vivo testing.

Experimental Results

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