Researchers say they’re one step closer to finding a potential cure for HIV after successfully eliminating the virus in living mice for the first time, reports CNBC. Using a combination of CRISPR gene-editing technology and a therapeutic treatment called LASER ART, scientists at Temple University and the University of Nebraska Medical Centre (UNMC) said they erased HIV DNA from the genomes of animals in what they call an unprecedented study.
“We think this study is a major breakthrough because it for the first time demonstrates after 40 years of the Aids epidemic that the HIV disease is a curable disease,” said study co-author Dr Kamel Khalili, chair of the department of neuroscience and director of the Centre for Neurovirology and the Comprehensive NeuroAIDS Centre at Temple University.
The virus is currently treated with antiretroviral therapy (ART), which suppresses it from replicating and prevents many patients in the US from developing Aids. ART does not rid the body of HIV, though, and if a patient stops treatment the virus will continue to replicate. But now researchers say they’re able to destroy the virus in “humanised” mice, which were injected with human bone marrow to imitate the human immune system.
The report says the study authors used two different tools to combat the virus: CRISPR technology and LASER ART.
CRISPR-Cas9 is a gene editing tool that’s been boasted as breakthrough technology that can help researchers treat or potentially cure genetic diseases. It gives scientists the ability to change an organism’s DNA, so they can add, remove or change certain genetic material.
LASER ART is a “super” form of ART that keeps replication of the virus at low levels for longer time periods, according to co-author Dr Howard Gendelman, chair of UNMC’s pharmacology and experimental neuroscience department and director of the Centre for Neurodegenerative Diseases. The antiretroviral drug is then stored in nanocrystals, which slowly release the drug where the virus is located.
“We’re going at the root cause,” Gendelman said. “We’re going after the virus that’s already integrated in the genome of the host cell.”
The researchers said they used the two treatments concurrently. First, they administered the LASER ART to reduce HIV growth, then they used the CRISPR treatment as “chemical scissors” to “eliminate the residual integrated HIV DNA still present,” Gendelman said. The virus did not return in nine of the 21 mice in which the method was tested, according to Khalili.
According to the report, the study authors say the findings are promising and they are now testing the CRISPR-LASER ART combination on primates. However, there is still much work that needs to be done before the method can be tested on humans.
“Things that work in mice, may not work in men,” Gendelman said. “The limitations of any mouse work have to do with the species, how the drug is administered, the distribution, which is a lot easier than a man or a woman.”
Gendelman noted humans are much larger than mice, so there are a lot more HIV DNA scientists will have to destroy if they test in humans. Khalili also said steps still need to be taken to increase efficiency and safety of the method, but the data from this study has researchers “hopeful” it will work in primates and that they are “cautious, but very aggressive” moving forward with testing.
The researchers aim to be granted approval by the US Food and Drug Administration to conduct a phase 1 clinical trial in humans by mid-2020, Khalili said.
Though the study was conducted on mice, Gendelman said it’s still important because it shows that sterilisation of HIV in living animals is possible. “We are at the cusp of a scientific revolution in human genomes that can change the course, quality and longevity of life,” Gendelman said.
A New Scientist report comments on the research:
What have researchers achieved?: The claim is they have eliminated HIV from living animals for the first time, by cutting it out of its hiding places in the body – in other words, that they have cured the animals.
How did they do it?: They used a gene-editing system called CRISPR, which is seen as a highly promising – yet still experimental – medical strategy. Unlike early forms of gene therapy, CRISPR allows precise targeting of specific genes. In this case, mice infected with HIV were injected with a different harmless virus that made a version of the CRISPR enzyme programmed to destroy HIV genes hiding in the mouse cells’ DNA.
Why would HIV be in the mouse cells’ DNA?: That’s where the virus hides out. While we have very effective medicines, called antiretroviral therapy (ART) which can get rid of most HIV from the body, it cannot affect cells where HIV has inserted itself into the DNA and gone dormant. If people stop taking their ART medicines, some of this dormant viral DNA wakes up, and blood virus levels surge back up again. So any hope of a permanent cure requires getting rid of this reservoir of virus-infected cells, found all over the body, including in the bone marrow, brain, and lymph nodes.
Can CRISPR do that?: Not by itself, no. “The viruses are replicating so fast that CRISPR can’t catch up,” says Khalili. So his team used a two-pronged strategy of hitting the virus with both CRISPR and a highly potent form of the normal ART meds. The drug molecules were chemically tweaked to make them fat-soluble yet encased in a water-soluble molecular cage. This means just one injection into a mouse’s blood sends the drugs into immune cells where the active molecule is slowly released over time.
Could that be useful by itself?: Yes, Gendelman is separately trying to turn this strategy into a long-lasting form of ART, where people would need just one injection a year, instead of daily pills.
How effective are the two approaches together?: It’s not yet a useful cure. In the latest study, which tested different doses and formulations, about a third of 13 mice seemed to have HIV completely eliminated from their bodies; when the mice stopped being dosed with ART, there was no virus surge in their blood for up to five weeks. And when the animals were killed, highly sensitive genetic tests could detect no virus in the usual hiding places. The team are still tweaking their protocols to try to get higher success rates.
Was it safe?: This is the big question. In other groups’ work, the CRISPR enzyme hasn’t always been completely accurate, sometimes binding to similar genes as the one targeted. The main fear is it could introduce DNA mutations that could turn a cell cancerous. The team found no such “off-target” effects when they looked at the spleens of four treated animals. Still, that doesn’t prove it could never happen. And Allan Bradley of the University of Cambridge showed last year that even when the CRISPR enzyme hit the right target, it can sometimes lead to deletions of large flanking sequences of DNA, which could also cause cancerous mutations. “There could be damage of an adjacent gene,” says Bradley.
When could we start testing this approach in people?: As early as next year, says Khalili. The team have already started testing the approach in monkeys, a prerequisite before human trials, they announced at a conference earlier this year. But a big obstacle will be demonstrating to regulators that the technique is safe enough, given that in most people, anti-HIV meds reduce the virus to undetectable levels and give people a near-normal lifespan, as long as they keep taking their pills.
Elimination of HIV-1 requires clearance and removal of integrated proviral DNA from infected cells and tissues. Here, sequential long-acting slow-effective release antiviral therapy (LASER ART) and CRISPR-Cas9 demonstrate viral clearance in latent infectious reservoirs in HIV-1 infected humanized mice. HIV-1 subgenomic DNA fragments, spanning the long terminal repeats and the Gag gene, are excised in vivo, resulting in elimination of integrated proviral DNA; virus is not detected in blood, lymphoid tissue, bone marrow and brain by nested and digital-droplet PCR as well as RNAscope tests. No CRISPR-Cas9 mediated off-target effects are detected. Adoptive transfer of human immunocytes from dual treated, virus-free animals to uninfected humanized mice fails to produce infectious progeny virus. In contrast, HIV-1 is readily detected following sole LASER ART or CRISPR-Cas9 treatment. These data provide proof-of-concept that permanent viral elimination is possible.
Prasanta K Dash, Rafal Kaminski, Ramona Bella, Hang Su, Saumi Mathews, Taha M Ahooyi, Chen Chen, Pietro Mancuso, Rahsan Sariyer, Pasquale Ferrante, Martina Donadoni, Jake A Robinson, Brady Sillman, Zhiyi Lin, James R Hilaire, Mary Banoub, Monalisha Elango, Nagsen Gautam, R Lee Mosley, Larisa Y Poluektova, JoEllyn McMillan, Aditya N Bade, Santhi Gorantla, Ilker K Sariyer, Tricia H Burdo, Won-Bin Young, Shohreh Amini, Jennifer Gordon, Jeffrey M Jacobson, Benson Edagwa, Kamel Khalili,Howard E Gendelman