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April 8, 2016

Hitting HIV with CRISPR/Cas9 Can Arouse Resistance

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    This visual abstract depicts how HIV-1 can escape Cas9/sgRNA-mediated inhibition. The researchers reveal that the NHEJ repair machinery generates mutations in the HIV-1 Cas9 cleavage site that result in two outcomes: viral replication suppression and viral escape. [Wang et al./Cell Reports]

    Stretches of DNA altered by the human immunodeficiency virus (HIV) can be targeted by the CRISPR/Cas9 endonuclease system, resulting in strategically placed cuts, imperfect repairs to those cuts, and—finally—the end of viral replication. But in some cases, the battle-scarred DNA that CRISPR/Cas9 leaves behind won’t give up the fight. Worse, this DNA becomes harder to recognize, by dint of its scars, and becomes even more dangerous. It acquires a form of resistance, the ability to duck renewed attacks from CRISPR/Cas9.

    This finding emerged from a study carried out by an international team of scientists that represented McGill University, the University of Montreal, the Chinese Academy of Medical Sciences, and Peking Union Medical College. These scientists, led by McGill’s Chen Liang, Ph.D., found that when CRISPR/Cas9 is used to mutate HIV-1 within cellular DNA, two outcomes are possible: (1) inactivation of HIV-1 and (2) acceleration of viral escape. This finding, the researchers cautioned, potentially limits the use of CRISPR/Cas9 in HIV-1 therapy.

    The researchers also sounded an optimistic note. They pointed to strategies that could help overcome HIV’s tendency to escape CRISPR/Cas9’s antiviral action. For example, targeting multiple sites with CRISPR/Cas9 or using other enzymes aside from Cas9. Once a solution is identified, the next barrier will be identifying ways to deliver the treatment to patients.

    The research team’s work appeared April 7 in the journal Cell Reports, in an article entitled, “CRISPR/Cas9-Derived Mutations Both Inhibit HIV-1 Replication and Accelerate Viral Escape.” The article emphasized the importance of the CRISPR/Cas9 system’s reliance on single guide RNA (sgRNA), the programmable element of the system that allows DNA to be cleaved at specific sequences.

    “Using HIV-1, we have now demonstrated that many of [CRISPR/Cas9-derived mutations or indels] are indeed lethal for the virus, but that others lead to the emergence of replication competent viruses that are resistant to Cas9/sgRNA,” wrote the article’s authors. “This unexpected contribution of Cas9 to the development of viral resistance is facilitated by some indels that are not deleterious for viral replication, but that are refractory to recognition by the same sgRNA as a result of changing the target DNA sequences.”

    The authors added that indels that are compatible with viral viability should be taken into consideration if Cas9/sgRNA is used to treat virus infection and genetic diseases. They expect that such indels would contribute to virus escape not only when Cas9/sgRNA is utilized to control new infections, but also in the context of eliminating latent viral DNA of herpes viruses, hepatitis B virus (HBV), and HIV, among others.

    "CRISPR/Cas9 gives a new hope toward finding a cure, not just for HIV-1, but for many other viruses," said Dr. Liang. "We have a long road toward the goal, and there may be many barriers and limitations that we need to overcome, but we're confident that we will find success."

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