Tag Archives: Han Zhao

Off-target CRISPR-Cas9 gene-editing changes closer to home than originally believed according to three studies

Heidi Ledford’s June 25, 2020 article (Note: Links have been removed) for Nature focuses on three studies (not yet peer-reviewed) that viewed together suggest CRISPR (clustered regularly interspaced short palindromic repeats) gene-editing is less like using a pair of scissors to cut out unwanted mutations and more like using a catalyst (a chemical agent which increases chemical reactions) and getting unanticipated and unwatned reactions. Except, it’s an unpredictable catalyst.

A suite of experiments that use the gene-editing tool CRISPR–Cas9 to modify human embryos have revealed how the process can make large, unwanted changes to the genome at or near the target site. [emphasis mine]

The studies were published this month on the preprint server bioRxiv, and have not yet been peer-reviewed1,2,3. But taken together, they give scientists a good look at what some say is an underappreciated risk of CRISPR–Cas9 editing. Previous experiments have revealed that the tool can make ‘off target’ gene mutations far from the target site, but the nearby changes identified in the latest studies can be missed by standard assessment methods.

These safety concerns are likely to inform the ongoing debate over whether scientists should edit human embryos to prevent genetic diseases — a process that is controversial because it creates a permanent change to the genome that can be passed down for generations. “If human embryo editing for reproductive purposes or germline editing were space flight, the new data are the equivalent of having the rocket explode at the launch pad before take-off,” says Fyodor Urnov, who studies genome editing at the University of California, Berkeley, but was not involved in any of the latest research.

These studies,if borne out, offer new concerns (from Ledford’s June 25, 2020 article),

The changes are the result of DNA-repair processes harnessed by genome-editing tools. CRISPR–Cas9 uses a small strand of RNA to direct the Cas9 enzyme to a site in the genome with a similar sequence. The enzyme then cuts both strands of DNA at that site, and the cell’s repair systems heal the gap.

The edits occur during that repair: most often, the cell seals up the cut using an error-prone mechanism that can insert or delete a small number of DNA letters. If researchers provide a DNA template, the cell might sometimes use that sequence to mend the cut, resulting in a true rewrite. But broken DNA can also cause shuffling or loss of a large region of the chromosome.

Previous work using CRISPR in mouse embryos and other kinds of human cell had already demonstrated that editing chromosomes can cause large, unwanted effects4,5. But it was important to demonstrate the work in human embryos as well, says Urnov, because different cell types might respond to genome editing differently.

Such rearrangements could be missed in many experiments, which typically look for other unwanted edits, such as single DNA-letter changes or small insertions or deletions of only a few letters. The latest studies, however, looked specifically for large deletions and chromosomal rearrangements near the target site. [emphasis mine] “This is something that all of us in the scientific community will, starting immediately, take more seriously than we already have,” says Urnov. “This is not a one-time fluke.”

Ledford’s article offers some description and analysis of each of the three papers.Note: All of the research was done with nonviable embryos. For anyone who wants to read the papers for themselves here are links and citations for each of the three,

Frequent loss-of-heterozygosity in CRISPR-Cas9-edited early human embryos by Gregorio Alanis-Lobato, Jasmin Zohren, Afshan McCarthy, Norah M.E. Fogarty, Nada Kubikova, Emily Hardman, Maria Greco, Dagan Wells, James M.A. Turner, Kathy K. Niakan. bioRxiv DOI: https://doi.org/10.1101/2020.06.05.135913 Posted: June 5, 2020

Reading frame restoration at the EYS locus, and allele-specific chromosome removal after Cas9 cleavage in human embryos by Michael V. Zuccaro, Jia Xu, Carl Mitchell, Diego Marin, Raymond Zimmerman, Bhavini Rana, Everett Weinstein, Rebeca T. King, Morgan Smith, Stephen H. Tsang, Robin Goland, Maria Jasin, Rogerio Lobo, Nathan Treff, Dieter Egli. bioRxiv DOI: https://doi.org/10.1101/2020.06.17.149237 Posted June 18, 2020

FREQUENT GENE CONVERSION IN HUMAN EMBRYOS INDUCED BY DOUBLE STRAND BREAKS by Dan Liang, Nuria Marti Gutierrez, Tailai Chen, Yeonmi Lee, Sang-Wook Park, Hong Ma, Amy Koski, Riffat Ahmed, Hayley Darby, Ying Li, Crystal Van Dyken, Aleksei Mikhalchenko, Thanasup Gonmanee, Tomonari Hayama, Han Zhao, Keliang Wu, Jingye Zhang, Zhenzhen Hou, Jumi Park, Chong-Jai Kim, Jianhui Gong, Yilin Yuan, Ying Gu, Yue Shen, Susan B. Olson, Hui Yang, David Battaglia, Thomas O’Leary, Sacha A. Krieg, David M. Lee, Diana H. Wu, P. Barton Duell, Sanjiv Kaul, Jin-Soo Kim, Stephen B. Heitner, Eunju Kang, Zi-Jiang Chen, Paula Amato, Shoukhrat Mitalipov. bioRxiv DOI: https://doi.org/10.1101/2020.06.19.162214 Posted June 20, 2020

These papers are open access.

A July 17, 2018 posting is probably the first time I featured work showing that CRISPR gene-editing can result in off-target effects; it was followed up by a September 20, 2019 posting on the topic.

3-D underwater acoustic carpet cloak

Who can resist a ‘Black Panther’ reference (Wikipedia Black Panther film entry)? Certainly not me. Scientists from the Chinese Academy of Sciences made this June 4, 2018 announcement (also on EurekAlert),

Cloaking is one of the most eye-catching technologies in sci-fi movies. In two 2018 Marvel films, Black Panther and Avengers: Infinity War, Black Panther conceals his country Wakanda, a technologically advanced African nation, from the outside world using the metal vibranium.

However, in the real world, if you want to hide something, you need to deceive not only the eyes, but also the ears, especially in the underwater environment.

Recently, a research team led by Prof. YANG Jun from the Institute of Acoustics (IOA) of the Chinese Academy of Sciences designed and fabricated a 3D underwater acoustic carpet cloak (UACC) using transformation acoustics.

The research was published online in Applied Physics Letters on June 1 [2018].

Like a shield, the carpet cloak is a material shell that can reflect waves as if the waves were reflecting off a planar surface. Hence, the cloaked target becomes undetectable to underwater detection instruments like sonar.

Using transformation acoustics, the research team first finished the 2D underwater acoustic carpet cloak with metamaterial last year (Scientific Reports, April 6, 2017). However, this structure works only in two dimensions, and becomes immediately detectable when a third dimension is introduced.

To solve this problem, YANG Jun and his IOA team combined transformation acoustics with a reasonable scaling factor, worked out the parameters, and redesigned the unit cell of the 2D cloak. They designed the 3D underwater acoustic carpet cloak and then proposed a fabrication and assembly method to manufacture it. The 3D cloak can hide an object from top to bottom and deal with complex situations, such as acoustic detection in all directions.

The 3D underwater acoustic carpet cloak is a pyramid comprising eight triangular pyramids; each triangular pyramid is composed of 92 steel strips using a rectangle lattice, similar to a wafer biscuit. More vividly, if we remove the core from a big solid pyramid, we can hide something safely in the hollow space left.

“To make a 3D underwater acoustic carpet cloak, researchers needed to construct the structure with 2D period, survey the influence of the unit cell’s resonance, examine the camouflage effect at the ridge of the sample, and other problems. In addition, the fabrication and assembly of the 3D device required more elaborate design. The extension of the UACC from 2D to 3D represents important progress in applied physics,” said YANG.

In experimental tests, a short Gaussian pulse propagated towards the target covered with the carpet cloak, and the waves backscattered toward their origin. The cloaked object successfully mimicked the reflecting surface and was undetectable by sound detection. Meanwhile, the measured acoustic pressure fields from the vertical view demonstrated the effectiveness of the designed 3D structure in every direction.

“As the next step, we will try to make the 3D underwater acoustic carpet cloak smaller and lighter,” said YANG.

Funding for this research came from the National Natural Science Foundation of China (Grant No.11304351, 1177021304), the Youth Innovation Promotion Association of CAS (Grant No. 2017029), and the IACAS Young Elite Researcher Project (Grant No. QNYC201719).

Prof. YANG Jun and Dr. JIA Han led the research team from the Institute of Acoustics (IOA) of the Chinese Academy of Sciences. Prof. YANG Jun engages in research on sound, vibration and signal processing, and especially sound field control and array signal processing. They also work on other devices based on metamaterial in order to manipulate the propagation of sound waves.

A model of the device,

Caption: This is a model and photograph of the 3D underwater acoustic carpet cloak composed of over 700 steel strips. Credit: IOA

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

Experimental demonstration of three-dimensional broadband underwater acoustic carpet cloak by Yafeng Bi, Han Jia, Zhaoyong Sun, Yuzhen Yang, Han Zhao, and Jun Yang.
Appl. Phys. Lett. 112, 223502 (2018); https://doi.org/10.1063/1.5026199 Published Online: June 2018

This paper is open access.