Tag Archives: Chen Liu

Effective safety strategies for CRISPR (clustered regularly interspaced short palindromic repeats) gene drive experiments

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It’s very peculiar being able to understand each word individually in clustered regularly interspaced short palindromic repeats (CRISPR) but not being able to puzzle out much meaning other than the widely known ‘it’s a gene editor’.

Regardless, CRISPR is a powerful gene editing tool and that can lead to trouble. Even before CRISPR, we’ve had some genetic accidents. Perhaps the best known is the ‘killer bee’ or Africanized bee (from its Wikepedia entry),

The Africanized bee, also known as the Africanised honey bee, and known colloquially as “killer bee”, is a hybrid of the western honey bee species (Apis mellifera), produced originally by cross-breeding [emphasis mine] of the East African lowland honey bee (A. m. scutellata) with various European honey bees such as the Italian honey bee A. m. ligustica and the Iberian honey bee A. m. iberiensis.

The Africanized honey bee was first introduced to Brazil in 1956 in an effort to increase honey production, but 26 swarms escaped quarantine in 1957 [emphasis mine]. Since then, the hybrid has spread throughout South America and arrived in North America in 1985. Hives were found in South Texas of the United States in 1990.

Africanized bees are typically much more defensive than other varieties of honey bee, and react to disturbances faster than European honey bees. They can chase a person a quarter of a mile (400 m); they have killed some 1,000 humans, with victims receiving ten times more stings than from European honey bees. They have also killed horses and other animals.

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Getting back to how powerful CRISPR is, a group of scientists has developed a set of strategies for safeguarding gene drive experiments (from a January 22, 2019 eLife press release also on EurekAlert),

Researchers have demonstrated for the first time how two molecular strategies can safeguard CRISPR gene drive experiments in the lab, according to a study published today in eLife.

Their findings, first reported on bioRxiv, suggest that scientists can effectively use synthetic target sites and split drives to conduct gene drive research, without the worry of causing an accidental spread throughout a natural population.

Gene drives, such as those trialled in malaria mosquitoes, are genetic packages designed to spread among populations. They do this via a process called ‘drive conversion’, where the Cas9 enzyme and a molecule called guide RNA (gRNA) cut at a certain site in the genome. The drive is then copied in when the DNA break is repaired.

“CRISPR-based gene drives have sparked both enthusiasm and deep concerns due to their potential for genetically altering entire species,” explains first author Jackson Champer, Postdoctoral Fellow in the Department of Biological Statistics and Computational Biology at Cornell University, New York. “This raises the question about our ability to prevent the unintended spread of such drives from the laboratory into the natural world.

“Current strategies for avoiding accidental spread involve physically confining drive-containing organisms. However, it is uncertain whether this sufficiently reduces the likelihood of any accidental escape into the wild, given the possibility of human error.”

Two molecular safeguarding strategies have recently been proposed that go beyond simply confining research organisms. The first is synthetic target site drive, which homes into engineered genomic sites that are absent in wild organisms. The second is split drive, where the drive construct lacks a type of enzyme called the endonuclease and relies instead on one engineered into a distant site.

“The nature of these strategies means that they should prevent an efficient spread outside of their respective laboratory lines,” Champer adds. “We wanted to see if they both had a similar performance to standard homing drives, and if they would therefore be suitable substitutes in early gene-drive research.”

To do this, the team designed and tested three synthetic target site drives in the fruit fly Drosophila melanogaster. Each drive targeted an enhanced green fluorescent protein (EGFP) gene introduced at one of three different sites in the genome. For split drives, they designed a drive construct that targeted the X-linked gene yellow and lacked Cas9.

Their analyses revealed that CRISPR gene drives with synthetic target sites such as EGFP show similar behaviour to standard drives, and can therefore be used for most testing in place of these drives. The split drives demonstrated similar performance, and also allow for natural sequences to be targeted in situations where the use of synthetic targets is difficult. These include population-suppression drives that require the targeting of naturally occurring genes

“Based on our findings, we suggest these safeguarding strategies should be adopted consistently in the development and testing of future gene drives,” says senior author Philipp Messer, Assistant Professor in the Department of Biological Statistics and Computational Biology at Cornell University. “This will be important for large-scale cage experiments aimed at improving our understanding of the expected population dynamics of candidate drives. Ultimately, this understanding will be crucial for discussing the feasibility and risks of releasing successful drives into the wild, for example to reduce malaria and other vector-borne diseases.”

Here’s a link to and a citation for the paper,

Molecular safeguarding of CRISPR gene drive experiments by Jackson Champer, Joan Chung, Yoo Lim Lee, Chen Liu, Emily Yang, Zhaoxin Wen, Andrew G Clark, Philipp W Messer. DOI: 10.7554/eLife.41439 Short Report Jan 22, 2019

This paper is open access. For anyone who doesn’t mind reading an earlier version of a paper you can find it at bioRxiv, at https://www.biorxiv.org/content/early/2018/09/08/411876.

elife, which i’ve mentioned here here before in a February 8, 2018 posting is a (from their About eLife webpage)

… non-profit organisation inspired by research funders and led by scientists. Our mission is to help scientists accelerate discovery by operating a platform for research communication that encourages and recognises the most responsible behaviours in science.

Magical nanobots at University of Florida kill (almost) 100% of Hepatitis C virus—in the lab

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I’ve always preferred the term nanobots but the folks at the University of Florida are calling them nanorobots, from the July 16, 2012 news item on phys.org,

University of Florida researchers have moved a step closer to treating diseases on a cellular level by creating a tiny particle that can be programmed to shut down the genetic production line that cranks out disease-related proteins.

In laboratory tests, these newly created “nanorobots” all but eradicated hepatitis C virus infection. The programmable nature of the particle makes it potentially useful against diseases such as cancer and other viral infections.

The research effort, led by Y. Charles Cao, a UF associate professor of chemistry, and Dr. Chen Liu, a professor of pathology and endowed chair in gastrointestinal and liver research in the UF College of Medicine, is described online this week in the Proceedings of the National Academy of Sciences.

The news item originated with a July 16, 2012 news release from the University of Florida which goes on to explain how the researchers succeeded,

The Holy Grail of nanotherapy is an agent so exquisitely selective that it enters only diseased cells, targets only the specified disease process within those cells and leaves healthy cells unharmed.

To demonstrate how this can work, Cao and colleagues, with funding from the National Institutes of Health, the Office of Naval Research and the UF [University of Florida] Research Opportunity Seed Fund, created and tested a particle that targets hepatitis C virus in the liver and prevents the virus from making copies of itself.

Hepatitis C infection causes liver inflammation, which can eventually lead to scarring and cirrhosis. The disease is transmitted via contact with infected blood, most commonly through injection drug use, needlestick injuries in medical settings, and birth to an infected mother. More than 3 million people in the United States are infected and about 17,000 new cases are diagnosed each year, according to the Centers for Disease Control and Prevention. Patients can go many years without symptoms, which can include nausea, fatigue and abdominal discomfort.

Current hepatitis C treatments involve the use of drugs that attack the replication machinery of the virus. But the therapies are only partially effective, on average helping less than 50 percent of patients, according to studies published in The New England Journal of Medicine and other journals. Side effects vary widely from one medication to another, and can include flu-like symptoms, anemia and anxiety.

Cao and colleagues, including graduate student Soon Hye Yang and postdoctoral associates Zhongliang Wang, Hongyan Liu and Tie Wang, wanted to improve on the concept of interfering with the viral genetic material in a way that boosted therapy effectiveness and reduced side effects.

The particle they created can be tailored to match the genetic material of the desired target of attack, and to sneak into cells unnoticed by the body’s innate defense mechanisms.

Recognition of genetic material from potentially harmful sources is the basis of important treatments for a number of diseases, including cancer, that are linked to the production of detrimental proteins. It also has potential for use in detecting and destroying viruses used as bioweapons.

The new virus-destroyer, called a nanozyme, has a backbone of tiny gold particles and a surface with two main biological components. The first biological portion is a type of protein called an enzyme that can destroy the genetic recipe-carrier, called mRNA, for making the disease-related protein in question. The other component is a large molecule called a DNA oligonucleotide that recognizes the genetic material of the target to be destroyed and instructs its neighbor, the enzyme, to carry out the deed. By itself, the enzyme does not selectively attack hepatitis C, but the combo does the trick.

“They completely change their properties,” Cao said.

In laboratory tests, the treatment led to almost a 100 percent decrease in hepatitis C virus levels. In addition, it did not trigger the body’s defense mechanism, and that reduced the chance of side effects. Still, additional testing is needed to determine the safety of the approach. [emphases mine]

This treatment builds on some previous research,

The UF nanoparticle design takes inspiration from the Nobel prize-winning discovery of a process in the body in which one part of a two-component complex destroys the genetic instructions for manufacturing protein, and the other part serves to hold off the body’s immune system attacks. This complex controls many naturally occurring processes in the body, so drugs that imitate it have the potential to hijack the production of proteins needed for normal function. The UF-developed therapy tricks the body into accepting it as part of the normal processes, but does not interfere with those processes.

Since there’s no mention of human clinical trials, I’m guessing that we are at least 10 years from seeing this therapeutic agent on the market.

After drafting this post yesterday (July 17, 2012) and while waiting to post it today, I found Dexter Johnson’s July 17 2012 posting where he makes some important points about this research (Note: I have removed a link),

Of course, this is a long way from becoming a treatment anytime soon. A major caveat is that the use of nanotreatments for the targeting and destroying of abnormal cells like cancer cells is always problematic since those cells are “still us” as George Whitesides noted some time back.  It’s always a bit of a tricky business to make sure that nanoparticles are targeting those biological processes within us that we want stopped and not the ones we want to keep.

Dexter goes on to comment about using the terms ‘nanobots’ or ‘nano robots’; he’s less sanguine about it than I am.