The Dark Side of CRISPR

Could powerful molecular methodologies be used to engineer new bioweapons, or will it bring new hope for cures to devastating diseases?

Biological weapons (BW) have long represented an “existential” threat when governments consider the menace posed by their adversaries. In modern times, BW were employed by the Japanese during World War II. The Soviet Union had a large weaponizing program for anthrax, which was presumably dismantled in the 1990s, however, suspicions remain that the Russian facility has continued operation, developing strains of Anthrax with increased virulence and antibiotic resistance using genetic engineering technologies. In the 1990s, the psychopathic Japanese terrorist organization, Aum Shinrikyo, attempted to employ botulinum neurotoxin and Anthrax bacteria in a biowarfare plot, fortunately without success. In 2001, letters containing anthrax powder were sent to U.S. politicians and members of the media. These latter events caused several deaths, cleanup costs running into the billions of dollars, and tremendous panic and fear among the public.

These efforts were pursued with conventional microbial procedures that were available at the time. But in recent years there have been dramatic advances that may substantially change our level of concern over the risks of bio-warfare.

While technology for the manipulation of DNA and other biological molecules has been around for years, and is constantly improving, it has proven time-consuming, difficult, and expensive – with frequent unwanted side effects that severely limited their application. Until CRISPR.

A new method of modifying DNA appears poised to have a big impact on bio medicine. This revolutionary new lab technique, known as “CRISPR/Cas9”, is a means of inserting genes into bacteria and other organisms.

Scientists around the world are experimenting with gene editing.

Pronounced “crisper cas 9”, it was derived from a naturally occurring protective mechanism employed by bacteria to ward off viral invasion. As adopted to gene insertion protocols, it consists of two components. The first is an enzyme (Cas9), which acts as molecular scissors, cutting the host DNA (the molecule that stores and carries genetic information to the next generation), allowing bits of DNA to be added or removed. The second is a strand of predesigned RNA (Ribonucleic acid) called a “guide”. It recognizes a particular segment of DNA in the proper matching region of the host genetic material. The whole complex of molecules is referred to in the trade as a “cassette,” an analogy to information-containing components of electronic devices. The simplest way to understand this process is to think of the cassette as a tiny submarine that glides into the cell, recognizes its target DNA with its guide, hones in, snips it out and inserts its own DNA, which is now part of the host genetic material.  

The guide RNA can be designed to match any particular region in the genetic makeup of the target organism. Since the guide RNA contains a sequence of bases that are complementary to the target DNA, it will only bind to that specific region. When the cell recognizes that its DNA is clipped, it will try to repair the damage and zip the DNA back together. At this point, the system allows a modified gene to be carried into that position, thus programming the cell to carry out the wishes of the designer.

In the early days following its discovery, there were numerous glitches that compromised the CRISPR/Cas9 performance. However, the technology has been greatly improved since its initial discovery and is now available in kits from many molecular supply companies. The prices are quite modest (a perusal of the internet turns up several in the $100 to $200 range), and the kits can be employed by individuals trained in the art of molecular biology. This conjures up a nightmare scenario in which a rogue nation or terrorist group could produce a lethal bacterial or viral agent that would be impossible to counteract.

Today, large scale production, storage, protection and field testing of weaponized bacteria or viruses are beyond the abilities of a small group or a terrorist cell. However, a number of countries in the world have demonstrated the ability – and the will – to unleash horrific attacks upon their perceived enemies. They undoubtedly are following the current advances in gene manipulation technology with great interest. For now though, such advances in gene manipulation, while making the process faster, simpler and more accessible, are still quite a challenge to carry out.

Dr. John Seavitt, an Assistant Professor in the Department of Molecular and Human Genetics at Baylor College of Medicine in Houston, Texas, has long experience with the CRISPR technology. “Its value to us is that it is a rapid, robust technology. There are plenty of methods of introducing CRISPR reagents into cells, including the use of glass microneedles to inject CRISPR cassettes directly into cells. But following the initial entry, there are many variations of the desired outcome, meaning that a collection of cells doesn’t all get the same alteration, typically. So you need to sort through a number of changes to get the mutation that you want,” he notes.

“Another problem is that RNA is an extremely fragile molecule that is easily degraded in the experimental process. So the bottom line is that nobody is doing this in their basement. The effort that would be required to make the specific changes would be quite substantial. CRISPR requires the efficient delivery of several labile reagents into the target cells. Moreover, we don’t know how the more pathogenic varieties would behave. This is an important new genome editing tool, but it will only be useful if you know what you want to make.”

There are other obstacles to the execution of a credible biological warfare program, perhaps the greatest is the uncertainty of the behavior of these agents once released into the environment. In the commercial realm of engineered agricultural products (herbicides, pesticides, fertilizers), all manner of living and inert substances undergo arduous evaluation (usually for years) before they can be released to the environment; yet these new inventions still have phenomenal failure rates.

Given that engineered bacteria and viruses are lethal materials, their handling and use in battle would be extremely risky, and loading them with a burden of genetic modifications could affect their behavior outside of the laboratory in unpredictable ways. In order to be confident that the bioweapon would have its desired effect, it would be essential to have field data, which could require years of testing. Would a terrorist be content to keep deploying flawed product until hitting the motherlode?

CRISPR/Cas9 is the best of a new generation of tools for manipulating genes, and is being used to develop cures for diseases, improve agricultural products and engineer organisms that can carry out a variety of industrial processes. It is undergoing constant improvement, making it faster and easier to employ.

Last year, Chinese scientists alarmed the scientific community when they used CRISPR to alter human embryos, prompting an unprecedented international scientific conference to debate the ethics of using CRISPR to alter future generations.

In February of 2016, James Clapper, U.S. Director of National Intelligence, in the unclassified annual worldwide threat assessment report, added new recombinant DNA technologies to a litany of threats posed by weapons of mass destruction and proliferation. Robin Lovell-Badge, a scientist at Great Britain’s Francis Crick Institute, has also expressed apprehension that some malevolent researchers might be doing gene editing work outside the internationally acceptable rules of scientific behavior.

Although it represents a great step forward, the Lab science of CRISPR is far from trivial. It can only be successfully carried out in a modern, well equipped setting by experts with a good deal of training.

In the final analysis, CRISPR’s greatest threat may be its mere potential, driving fear and uncertainty throughout the public sphere. With a long and disturbing history of panic generated by relatively crude and ineffective agents, just the possibility of a new generation of more effective bioweaponry could drive governments to take radical and arbitrary steps to protect their terrified citizenry. The next few years will see many new advances.

___
John Morrow Jr. is a consultant with Newport Biotech in Kentucky.