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Another Milestone for CRISPR-Cas9 Technology: First Trial Data for Treatment Delivered Intravenously

Unlike most other CRISPR/Cas-9 therapies that are ex vivo treatments in which cells are modified outside the body, this study was successful with an in vivo treatment

Use of CRISPR-Cas9 gene editing technology for therapeutic purposes can be a boon for clinical laboratories. Not only is this application a step forward in the march toward precision medicine, but it can give clinical labs the essential role of sequencing a patient’s DNA to help the referring physician identify how CRISPR-Cas9 can be used to edit the patient’s DNA to treat specific health conditions.

Most pathologists and medical lab managers know that CRISPR-Cas9 gene editing technology has been touted as one of the most significant advances in the development of therapies for inherited genetic diseases and other conditions. Now, a pair of biotech companies have announced a milestone for CRISPR-Cas9 with early clinical data involving a treatment delivered intravenously (in vivo).

The therapy, NTLA-2001, was developed by Intellia Therapeutics (NASDAQ:NTLA) and Regeneron Pharmaceuticals (NASDAQ:REGN) for treatment of hereditary ATTR (transthyretin) amyloidosis, a rare and sometimes fatal liver disease.  

As with other therapies, determining which patients are suitable candidates for specific treatments is key to the therapy’s success. Therefore, clinical laboratories will play a critical role in identifying those patients who would most likely benefit from a CRISPR-delivered therapy.

Such is the goal of precision medicine. As methods are refined that can correct unwelcome genetic mutations in a patient, the need to do genetic testing to identify and diagnose whether a patient has a specific gene mutation associated with a specific disease will increase.

The researchers published data from a Phase 1 clinical trial of NTLA-2001 in the New England Journal of Medicine (NEJM), titled, “CRISPR-Cas9 In Vivo Gene Editing for Transthyretin Amyloidosis.” They also presented their findings at the Peripheral Nerve Society (PNS) Annual Meeting.

What is NTLA-2001 and Why Is It Important?

Cleveland Clinic describes ATTR amyloidosis as a “protein misfolding disorder” involving transthyretin (TTR), a protein made in the liver. The disease leads to deposits of the protein in the heart, nerves, or other organs.

According to Intellia and Regeneron, NTLA-2001 is designed to inactivate the gene that produces the protein.

The interim clinical trial data indicated that one 0.3 mg per kilogram dose of the therapy reduced serum TTR by an average of 87% at day 28. A smaller dose of 0.1 mg per kilogram reduced TTR by an average of 52%. The researchers reported “few adverse events” in the six study patients, “and those that did occur were mild in grade.”

Current treatments, the companies stated, must be administered regularly and typically reduce TTR by about 80%.

“These are the first ever clinical data suggesting that we can precisely edit target cells within the body to treat genetic disease with a single intravenous infusion of CRISPR,” said Intellia President and CEO John Leonard, MD, in a press release. “The interim results support our belief that NTLA-2001 has the potential to halt and reverse the devastating complications of ATTR amyloidosis with a single dose.”

He added that “solving the challenge of targeted delivery of CRISPR-Cas9 to the liver, as we have with NTLA-2001, also unlocks the door to treating a wide array of other genetic diseases with our modular platform, and we intend to move quickly to advance and expand our pipeline.”

Daniel Anderson, PhD

“It’s an important moment for the field,” MIT biomedical engineer Daniel Anderson, PhD (above), told Nature. Anderson is Professor, Chemical Engineering and Institute for Medical Engineering and Science at the Koch Institute for Integrative Cancer Research at MIT. “It’s a whole new era of medicine,” he added. Advances in the use of CRISPR-Cas9 for therapeutic purposes will create the need for clinical laboratories to sequence patients’ DNA to help physicians determine the best uses for a CRISPR-Cas9 treatment protocol. (Photo copyright: Massachusetts Institute of Technology.)

In Part 2 of the Phase 1 trial, Intellia plans to evaluate the new therapy at higher doses. After the trial is complete, “the company plans to move to pivotal studies for both polyneuropathy and cardiomyopathy manifestations of ATTR amyloidosis,” the press release states.

Previous clinical trials reported results for ex vivo treatments in which cells were removed from the body, modified with CRISPR-Cas9 techniques, and then reinfused. “But to be able to edit genes directly in the body would open the door to treating a wider range of diseases,” Nature reported.

How CRISPR-Cas9 Works

On its website, CRISPR Therapeutics, a company co-founded by Emmanuelle Charpentier, PhD, a director at the Max Planck Institute for Infection Biology in Berlin, and inventor of CRISPR-Cas9 gene editing, explained that the technology “edits genes by precisely cutting DNA and then letting natural DNA repair processes take over.” It can remove fragments of DNA responsible for causing diseases, as well as repairing damaged genes or inserting new ones.

The therapies have two components: Cas9, an enzyme that cuts the DNA, and Guide RNA (gRNA), which specifies where the DNA should be cut.

Charpentier and biochemist Jennifer Doudna, PhD, Nobel Laureate, Professor of Chemistry, Professor of Biochemistry and Molecular Biology, and Li Ka Shing Chancellor’s Professor in Biomedical and Health at the University of California Berkeley, received the 2020 Nobel Prize in Chemistry for their work on CRISPR-Cas9, STAT reported.

It is important to pathologists and medical laboratory managers to understand that multiple technologies are being advanced and improved at a remarkable pace. That includes the technologies of next-generation sequencing, use of gene-editing tools like CRISPR-Cas9, and advances in artificial intelligence, machine learning, and neural networks.

At some future point, it can be expected that these technologies will be combined and integrated in a way that allows clinical laboratories to make very early and accurate diagnoses of many health conditions.

—Stephen Beale

Related Information

Intellia and Regeneron Announce Landmark Clinical Data Showing Deep Reduction in Disease-Causing Protein After Single Infusion of NTLA-2001, an Investigational CRISPR Therapy for Transthyretin (ATTR) Amyloidosis

CRISPR-Cas9 In Vivo Gene Editing for Transthyretin Amyloidosis

Landmark CRISPR Trial Shows Promise Against Deadly Disease

CRISPR Milestone Pushes Gene Editing Toward Its Promise

CRISPR Clinical Trials: A 2021 Update

CRISPR Gene Therapy: Applications, Limitations, and Implications for the Future

Diseases CRISPR Could Cure: Latest Updates on Research Studies and Human Trials

Faster, Better, Cheaper: The Rise of CRISPR in Disease Detection

The Potential of CRISPR-Based Diagnostic Assays and Treatment Approaches Against COVID-19

Two Female CRISPR Scientists Make History, Winning Nobel Prize in Chemistry for Genome-Editing Discovery

CRISPR-Cas9 DNA Editing Possibly Linked to Cancer, But CRISPR-Cas13d RNA Editing Could Offer New Avenues for Treatment

CRISPR-Cas9 connection to cancer prompts research to investigate different approaches to gene editing

Dark Daily has covered CRISPR-Cas9 many times in previous e-briefings. Since its discovery, CRISPR, or Clustered Regularly Interspaced Short Palindromic Repeats, has been at the root of astonishing breakthroughs in genetic research. It appears to fulfill precision medicine goals for patients with conditions caused by genetic mutations and has anatomic pathologists, along with the entire scientific world, abuzz with the possibilities such a tool could bring to diagnostic medicine.

All of this research has contributed to a deeper understanding of how cells function. However, as is often the case with new technologies, unforeseen and problematic questions also have arisen.

CRISPR-Cas9 Connection to Cancer

Research conducted at the Wellcome Sanger Institute in the United Kingdom (UK) and published in Nature Biotechnology, examined potential damage caused by CRISPR-Cas9 editing.

“Here we report significant on-target mutagenesis, such as large deletions and more complex genomic rearrangements at the targeted sites in mouse embryonic stem cells, mouse hematopoietic progenitors, and a human differentiated cell line,” wrote the authors in their introduction.

Another study, this one conducted by biomedical researches at Cambridge, Mass., and published in Nature, describes possible toxicity caused by Cas9.

“Our results indicate that Cas9 toxicity creates an obstacle to the high-throughput use of CRISPR-Cas9 for genome engineering and screening in hPSCs [human pluripotent stem cells]. Moreover, as hPSCs can acquire P53 mutations, cell replacement therapies using CRISPR-Cas9-enginereed hPSCs should proceed with caution, and such engineered hPSCs should be monitored for P53 function.”

Essentially what both groups of researchers found is that CRISPR-Cas9 cuts through the double helix of DNA, which the cell responds to as it would any injury. A gene called p53 then directs a cellular “first-aid kit” to the “injury” site that either initiates self-destruction of the cell or repairs the DNA.

Therefore, in some instances, CRISPR-Cas9 is inefficient because the repaired cells continue to function. And, the repair process involves the p53 gene. P53 mutations have been implicated in ovarian, colorectal, lung, pancreatic, stomach, liver, and breast cancers.

Though important, some experts are downplaying the significance of the findings.

Erik Sontheimer, PhD (above), Professor, RNA Therapeutics Institute, at the University of Massachusetts Medical School, told Scientific American that the two studies are important, but not show-stoppers. “This is something that bears paying attention to, but I don’t think it’s a deal-breaker,” he said. (Photo copyright: University of Massachusetts.)

“It’s something we need to pay attention to, especially as CRISPR expands to more diseases. We need to do the work and make sure edited cells returned to patients don’t become cancerous,” Sam Kulkarni, PhD, CEO of CRISPR Therapeutics, told Scientific American.

Both studies are preliminary. The implications, however, is in how genes that have become corrupted are used.

“It is unclear if the findings translate into cells actually used in clinical studies,” Bernhard Schmierer, PhD, co-author of a paper titled, “CRISPR-Cas9 Genome Editing Induces a p53-mediated DNA Damage Response,” told Scientific American.

Nevertheless, the cancer-cat is out of the bag.

Targeting RNA Instead of DNA with CRISPR-Cas13d

A team from the Salk Institute may have found a solution. They are investigating a different enzyme—Cas13d—which, in conjunction with CRISPR would target RNA rather than DNA. “DNA is constant, but what’s always changing are the RNA messages that are copied from the DNA. Being able to modulate those messages by directly controlling the RNA has important implications for influencing a cell’s fate,” Silvana Konermann, PhD, a Howard Hughes Medical Institute (HHMI) Hanna Gray Fellow and member of the research team at Salk, said in a news release.

The Salk team published their findings in the journal Cell. The paper describes how “scientists from the Salk Institute are reporting for the first time the detailed molecular structure of CRISPR-Cas13d, a promising enzyme for emerging RNA-editing technology. They were able to visualize the enzyme thanks to cryo-electron microscopy (cryo-EM), a cutting-edge technology that enables researchers to capture the structure of complex molecules in unprecedented detail.”

The researchers think that CRISPR-Cas13d may be a way to make the process of gene editing more effective and allow for new strategies to emerge. Much like how CRISPR-Cas9 led to research into recording a cell’s history and to tools like SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing), a new diagnostic tool that works with CRISPR and changed clinical laboratory diagnostics in a foundational way.

Dark Daily reported on this breakthrough last year. (See, “CRISPR-Related Tool Set to Fundamentally Change Clinical Laboratory Diagnostics, Especially in Rural and Remote Locations,” August 4, 2017.)

Each discovery will lead to more branches of inquiry and, hopefully, someday it will be possible to cure conditions like sickle cell anemia, dementia, and cystic fibrosis. Given the high expectations that CRISPR and related technologies can eventually be used to treat patients, pathologists and medical laboratory professionals will want to stay informed about future developments.

—Dava Stewart

Related Information:

Repair of Double-Strand Breaks Induced by CRISPR-Cas9 Leads to Large Deletions and Complex Rearrangements

P53 Inhibits CRISPR-Cas9 Engineering in Human Pluripotent Stem Cells

CRISPR-Edited Cells Linked to Cancer Risk in 2 Studies

CRISPR-Cas9 Genome Editing Induces a p53-Mediated DNA Damage Response

Decoding the Structure of an RNA-Based CRISPR System

Structural Basis for the RNA-Guided Ribonuclease Activity of CRISPR-Cas13d

CRISPR Timeline

What Are Genome Editing and CRISPR-Cas9?

Federal Court Sides with Broad in CRISPR Patent Dispute

Top Biologists Call for Moratorium on Use of CRISPR Gene Editing Tool for Clinical Purposes Because of Concerns about Unresolved Ethical Issues

CRISPR-Related Tool Set to Fundamentally Change Clinical Laboratory Diagnostics, Especially in Rural and Remote Locations

Researchers at Several Top Universities Unveil CRISPR-Based Diagnostics That Show Great Promise for Clinical Laboratories

Patent Dispute over CRISPR Gene-Editing Technology May Determine Who Will Be Paid Licensing Royalties by Medical Laboratories

U.S. Patent and Trademark Office will hold hearings to determine whether University of California Berkeley, or Broad Institute of Harvard and MIT, should receive patents for new genomic engineering technique

In the race to master gene-editing in ways that will advance genetic medicine and patient care, one of the hottest technologies is CRISPR, which stands for Clustered Regularly Interspaced Short Palindromic Repeats. But now a patent fight has the potential to complicate how pathologists and other scientists use this exciting technology.

This dispute over the CRISPR patent—a tool that has been hailed as one of the biggest biotech breakthroughs of the decade—will likely be settled in the coming months by the United States Patent and Trademark Office (USPTO).

The USPTO will be reviewing key patents awarded for what is called CRISPR/Cas9. The technology is already generating novel therapies for diseases, which should create new opportunities for pathologists and medical laboratories. (more…)

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