Chapter 1: The Origins of CRISPR and Its Implications

In September 2016, Jennifer Doudna invited her colleague Kyle Watters to discuss a pressing concern. Doudna, renowned for co-developing the CRISPR technology, had gained significant recognition and wealth from this groundbreaking discovery. However, she had been troubled by a nightmare featuring Adolf Hitler requesting the CRISPR blueprint, leaving her to ponder the potential misuse of such a powerful tool.

Now, Doudna posed a crucial question: would Watters be interested in collaborating to find a way to neutralize CRISPR?

CRISPR, a natural defense mechanism in bacteria, has evolved over millions of years to combat viral invasions by cleaving their DNA. Doudna's contributions had transformed this biological process into a revolutionary gene-editing tool, sparking a surge of innovative research and potential medical breakthroughs. Yet, the prospect of delivering CRISPR into human bodies raises fears of malicious use by individuals or states, leading to targeted attacks or the creation of super-soldiers. Doudna herself had expressed worries in her book, "A Crack in Creation," about gene editing attracting attention akin to nuclear weapons. "Could I and other concerned scientists prevent CRISPR from spiraling out of control before a catastrophe occurs?" she questioned.

As luck would have it, 2016 marked a significant shift; U.S. intelligence agencies identified gene editing as a potential weapon of mass destruction. In response, the Defense Advanced Research Projects Agency (DARPA) initiated a program called Safe Genes, allocating over $65 million to explore methods to manage or reverse gene-editing technologies. Doudna and Watters's work was vital to this initiative, aiming to develop countermeasures against bioterrorism, including the potential misuse of CRISPR itself.

What could CRISPR weapons entail? While the specifics are speculative, DARPA has tasked Doudna and her team with investigating preventative treatments, perhaps akin to taking antibiotics in response to an anthrax threat. Researchers are preparing to conduct initial trials on mice to test the feasibility of rendering them immune to CRISPR's effects.

"Can we deactivate CRISPR?" inquired Joseph S. Schoeniger, leading one segment of the defense initiative at Sandia National Laboratories. "That's our goal. Given the rapid development of this technology, it would be beneficial to have a means to turn it off."

Section 1.1: Discovering Anti-CRISPR Mechanisms

By the time Doudna submitted her proposal to DARPA, research had already uncovered valuable insights into counteracting CRISPR. In the ongoing battle between bacteria and their phage viruses, phages have evolved mechanisms to counteract CRISPR's effects. Researchers found these phages possess proteins that can effectively neutralize CRISPR—dubbed "anti-CRISPRs."

The discovery of the first anti-CRISPRs emerged serendipitously in 2013, thanks to Joseph Bondy-Denomy, then a student at the University of Toronto. "We accidentally observed that some phages seemed resilient to CRISPR," he explained. "Once we introduced the phage into a bacterial cell, it could not defend itself." He quickly identified a specific gene within the virus responsible for this resistance. "This might be the key to disabling CRISPR."

While the number of laboratories investigating anti-CRISPRs is comparatively fewer than those studying CRISPR itself, interest in this area is rapidly growing. Over 40 anti-CRISPR proteins have been identified, many through Doudna's lab. Other groups are also making strides in discovering conventional chemicals capable of inhibiting CRISPR. Recently, Amit Choudhary from Harvard Medical School announced his findings of two drugs that can obstruct gene editing when combined with human cells. "Control is a hallmark of any potent technology," Choudhary noted succinctly.

Subsection 1.1.1: Potential Applications of Anti-CRISPR

Researchers like Bondy-Denomy believe that anti-CRISPRs could enhance future gene-editing treatments by providing more precise control. For instance, a German team demonstrated that combining CRISPR with anti-CRISPR could allow for targeted DNA modifications solely in liver cells, sparing other tissues.

Another avenue of research is the potential use of anti-CRISPRs as a safeguard against "gene drives." Supported by the Bill & Melinda Gates Foundation, one project aims to develop a CRISPR tool that would spread through wild mosquito populations, significantly reducing their numbers to combat malaria. Others aspire to create gene drives in mice to eliminate them from specific islands without resorting to poisons.

However, concerns arise regarding the unintended consequences of these experiments leading to extinction. Researchers are optimistic that they can genetically engineer organisms with anti-CRISPR capabilities, ensuring their immunity. In an initial proof-of-concept, scientists in Kansas successfully engineered yeast cells with anti-CRISPR to resist a gene drive. "If a laboratory in North Korea attempts to deploy a gene drive to eliminate a vital crop, a transgenic crop could be developed to withstand it," Erik Sontheimer from the University of Massachusetts Medical School explained.

Chapter 2: The Growing Urgency for Safety Measures

The emergence of CRISPR in 2012 caught scientists off guard, revolutionizing genetic engineering overnight. Traditional methods were swiftly replaced by a cost-effective and adaptable tool for altering DNA in any living organism. According to Renee Wegrzyn, a biodefense scientist overseeing DARPA's program, forecasters failed to anticipate the dangers posed by CRISPR, which soon became a "critical urgent issue for national security."

As researchers, medical professionals, and venture capitalists raced to implement CRISPR across various applications—plants, animals, and humans—concerns about the potential biothreats escalated. By 2015, Doudna began to question the safety of CRISPR's everyday laboratory applications, fearing the consequences of potential accidents. "We are pushing these technologies into the world without adequate safety measures," Wegrzyn expressed during a 2017 gathering of the Long Now Foundation in San Francisco. "There is a pressing need to address these issues."

Wegrzyn highlighted the risks posed by experiments that intentionally make mice ill by disrupting essential genes. "One doesn't need to be a biosecurity expert to recognize the necessity for scrutiny when dealing with a tool that can both cure and cause disease," she stated. "If we need to halt a gene editor instantly, we currently lack the means to do so."

The debate continues regarding the extent of CRISPR's dangers in malicious hands. "Red team" exercises conducted by the CIA in 2016, which tasked analysts with envisioning worst-case scenarios, did not reach a consensus. Subsequently, the National Academies of Sciences, Engineering and Medicine, at the Department of Defense's request, compiled a risk ranking for synthetic biology threats, placing CRISPR weapons in the middle tier. The military indicated that it did not perceive an immediate danger to its personnel.

Doudna believes that CRISPR's risks should not be exaggerated. "I often receive inquiries about the potential nefarious uses of CRISPR, and I feel equally concerned about it as I do about other technologies. It is entirely possible to synthesize the smallpox virus," she remarked. While her research may eventually yield a gene-editing antidote, she emphasizes that her lab's exploration of anti-CRISPRs primarily seeks to address fundamental biological questions. "I am still at step one," she asserts. "How do these mechanisms function?"

In contrast, some scientists worry that the risks are already evident and that antidotes are urgently needed. Certain researchers have even advocated for restricting public discourse on specific CRISPR studies or removing references to them online, presumably to buy time for developing countermeasures. "The prevailing attitude is to avoid alarming the public while we actively seek solutions. There is always apprehension about an early panic," noted Watters, Doudna's former collaborator, who authored a 2018 review on gene editing's implications for biosecurity.

A video still illustrating CRISPR editing DNA in real time. Screenshot: Osamu Nureki/Nature Communications