Scientists Engineer the First CRISPR-Based Drug Candidate to Directly Target E. Coli

Many people have experienced infections from E. coli, which are primarily seen as inconvenient and unpleasant. For some patients, like those with blood cancer, however, there is a risk that the bacteria will travel into the bloodstream. In those cases, an E. coli infection is too often fatal. The mortality rate is 15-20%.

The predominant cure for such infections is the use of antibiotics that have detrimental effects on the patient's microbiome, which play a key part in our physical and emotional well-being, and other side effects. Furthermore, growing problems with antibiotic resistance render such treatments less effective in treating infections.

An international team of scientists has now engineered the first published CRISPR-based candidate (see fact box) for a drug that targets E. coli directly and leaves the microbiome intact. A new paper in Nature Biotechnology titled 'Engineered phage with antibacterial CRISPR–Cas selectively reduce E. coli burden in mice' describes the development of the drug candidate to a stage where it is ready for tests on humans.

Through extensive use of synthetic biology, the team designed four bacterial viruses that use CRISPR technology to kill the unwanted bacteria precisely.

We believe that a narrow spectrum drug with these properties could be very useful to cancer patients, among others, who often get serious infections that are difficult to treat with current antibiotics."

Morten Otto Alexander Sommer, professor at DTU Biosustain, Co-founder of SNIPR Biome, and lead author of the paper

The work was carried out in collaboration with JAFRAL (Slovenia), JMI Laboratories (US), and Division of Infectuous Diseases at Weill Cornell Medicine (US).

Engineering Phages to Target E. Coli

The team, primarily based at SNIPR Biome, screened a library of 162 naturally occurring phages (viruses that kill specific bacteria; see fact box). They found that eight of these phages showed promise in targeting E. coli. They then engineered the phages through gene editing to improve their ability to target E. coli.

A cocktail of four of these phages, which they named SNIPR001, very effectively targeted bacteria in biofilms and reduced the number of E. coli in a manner that surpassed that of naturally occurring phages. Further, they showed that the cocktail of phages was tolerated well in the gut of mice and mini pigs while reducing the emergence of E. coli. SNIPR001 is now in clinical development and has been granted a Fast-Track designation (expedited review) by the US Food and Drug Administration.

FACT BOX: An Overview of the SNIPR001 Creation Process:

1. Naturally occurring phages are screened against a panel of E. coli strains.
2. Phages with broad activity against E. coli are tail fibre engineered and/or armed with CRISPR–Cas systems containing sequences specific to E. coli, creating CAPs (Cas-armed phages).
3. These CAPs are tested for host range, in vivo efficacy, and CMC specifications.

SNIPR001 comprises four complementary CAPs and is a new precision antibiotic that selectively targets E.coli to prevent bacteremia in hematological cancer patients at risk of neutropenia (low levels of white blood cells).

Blood Cancer Patients are First in Line

The reason this new development is exciting for blood cancer patients has to do with side effects stemming from their chemotherapy treatment. It causes the patient's bone marrow to produce fewer blood cells and inflammation of the intestines. The latter increases the intestines' permeability allowing bacteria from the gut to travel into the bloodstream. This combination of side effects leaves the patient vulnerable to infections from bacteria like E. coli. In such cases, the

Today, patients at risk (i.e., with low levels of white blood cells) receive antibiotic treatments ahead of their chemotherapy, but in some cases, E. coli shows very high resistance to commonly used antibiotics. Also, the antibiotics themselves have several side effects that in some cases reduce the effect of the cancer treatments.

"We need a wider variety of options available to treat these patients, preferably ones where we can specifically target the bacteria responsible to avoid side effects and that do not add to the problem of antibiotic resistance," says Morten Otto Alexander Sommer.

In recent years, researchers have been looking back towards using phages to treat infections because of the increase in antibiotic resistance. Before antibiotics were broadly available, phages were widely used and studied in countries that were then part of the Soviet Union. Still, there are few clinical trials, and the results haven't been convincing, according to the paper.

"Through emerging technologies like CRISPR, the use of phages in treating infections has become a viable pathway. As our results show, there is potential for enhancing naturally occurring phages through genetic engineering. It is my hope that this approach may also serve as a blueprint for new antimicrobials targeting resistant pathogens," says Morten Otto Alexander Sommer.

FACT BOX: CRISPR, Phages, and Phage Therapy

CRISPR technology is a way for scientists to edit DNA sequences in cells. It's based on a defence mechanism bacteria naturally use to protect themselves. CRISPR technology uses a molecule called Cas9, which works like a pair of scissors to cut DNA at a specific spot.

After the cut, the DNA can be fixed, or a new piece can be added. Scientists can use this tool to create genetically modified organisms, find new ways to treat genetic diseases, and learn more about how genes work.

Phages are tiny viruses that can kill specific bacteria. They're everywhere on Earth and help regulate bacterial populations and nutrient cycling. They infect and kill bacteria, and when the bacteria die, they release nutrients into the environment.

Scientists use phages to treat bacterial infections, which is called phage therapy. They identify and isolate phages that can kill a specific bacterial strain and use them to fight infections caused by that strain.

Phage therapy has some advantages to antibiotics, like targeting specific bacteria without side effects and potentially reducing antibiotic resistance.