CRISPR Used to Silence Crucial Hepatitis B Gene

Christina Bennett

November 12, 2019

The CRISPR gene-editing tool can be used to silence an important hepatitis B virus gene, a proof-of-concept in vitro study suggests.

"It's the first time we've seen CRISPR editing done in a hepatitis B model," said Douglas Dieterich, MD, director of the Institute of Liver Medicine and professor of medicine at the Icahn School of Medicine at Mount Sinai in New York City.

Hepatitis B can lead to liver disease and is the primary cause of hepatocellular carcinoma. In 2015, more than 250 million people around the world were infected with the virus, according to the World Health Organization.

For their study, investigator Hao Zhou, from The First Hospital of Jilin University in China and the Department of Medicine at the University of Minnesota in Minneapolis, and colleagues targeted the S gene. Zhou presented the findings at the Liver Meeting 2019 in Boston.

The S gene gives rise to the hepatitis B surface antigen, the presence of which indicates that a person is infected with the virus. "The question is whether it's the right target," Dieterich told Medscape Medical News.

Reducing the amount of the hepatitis B surface antigen is a "good idea" because that's what is believed to inhibit the immune system from clearing the virus. Doing so might help the immune system recover and clear the virus, "with a little help from some antivirals," explained Dieterich, who was not involved in the study.

However, "the surface is not the only DNA that's integrated into the host genome," he pointed out. "I think maybe a broader application might be necessary to actually get the hepatitis B genome out of the hepatocytes."

Zhou's team used a newer CRISPR approach, called CRISPR-STOP, for their gene-editing procedure.

CRISPR-STOP

"The idea is that CRISPR-STOP can be as efficient as standard CRISPR editing, but it's safer," said Kiran Musunuru, MD, PhD, associate professor of cardiovascular medicine and genetics at Penn Medicine in Philadelphia, who was not involved in the study. Musunuru is cofounder of and senior scientific advisor at Verve Therapeutics, a company using gene editing to prevent cardiovascular disease.

The standard CRISPR-Cas9 approach requires a double-strand break in the genome, and the problem with that is it introduces the possibility for "mischief," he explained. "If you have more than one double-strand break occurring in the human genome at the same time, you have the potential for different parts of different chromosomes coming together in the wrong ways and then causing problems."

Instead of creating a double-strand break, CRISPR-STOP uses a base editor to chemically modify the DNA base from one base to another and introduce a stop codon into the target gene sequence, effectively hamstringing the ability of the target gene to produce a functional protein.

This is a very nice, clean way to turn off a gene effectively.

"This is a very nice, clean way to turn off a gene effectively," Musunuru told Medscape Medical News.

For their CRISPR-STOP procedure, Zhou's team first transduced liver cells infected with the hepatitis B virus using a base editor called AncBE4max. Next, to activate the base editor so that gene editing could begin, they transduced the cells with one of two lentivectors: one encoded for single-guide RNA that targets the S gene; and an empty one, which served as the control.

With the gene-editing approach, 71% of the liver cells that expressed the base editor gained the desired stop codon in the target gene.

"That's a very robust number," said Musunuru.

In addition, hepatitis B surface antigen secretion was reduced by 92% with the gene-editing approach.

The investigators report a high degree of conservativity for hepatitis B genotypes B, C, F, and H. Specifically, 94% of the S gene sequence was conserved for genotype B, 92% for genotype C, 91% for genotype F, and 71% for genotype H.

The Liver Meeting 2019: American Association for the Study of Liver Diseases (AASLD): Abstract 86. Presented November 10, 2019.

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