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ADVANCES IN CRISPR GENOME EDITING

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 WHAT IS CRISPR?

“CRISPR” (pronounced “crisper”) stands for Clustered Regularly Interspaced Short Palindromic Repeats, which are the hallmark of a bacterial defense system that forms the basis for CRISPR-Cas9 genome editing technology. In the field of genome engineering, the term “CRISPR” or “CRISPR-Cas9” is often used loosely to refer to the various CRISPR-Cas9 and -CPF1, (and other) systems that can be programmed to target specific stretches of genetic code and to edit DNA at precise locations, as well as for other purposes, such as for new diagnostic tools. With these systems, researchers can permanently modify genes in living cells and organisms and, in the future, may make it possible to correct mutations at precise locations in the human genome in order to treat genetic causes of disease. Other systems are now available, such as CRISPR-Cas13, that target RNA provide alternate avenues for use, and with unique characteristics that have been leveraged for sensitive diagnostic tools, such as SHERLOCK.

WHAT IS THE ORIGIN OF CRISPR?

 CRISPRs were first discovered in archaea (and later in bacteria) by Francisco Mojica, a scientist at the University of Alicante in Spain. He proposed that CRISPRs serve as part of the bacterial immune system, defending against invading viruses. They consist of repeating sequences of genetic code, interrupted by “spacer” sequences – remnants of genetic code from past invaders. The system serves as a genetic memory that helps the cell detect and destroy invaders (called “bacteriophages”) when they return. Mojica’s theory was experimentally demonstrated in 2007 by a team of scientists led by Philippe Horvath.

In January 2013, the Zhang lab published the first method to engineer CRISPR to edit the genome in mouse and human cells.


HOW DOES IT WORK?    

CRISPR “spacer” sequences are transcribed into short RNA sequences (“CRISPR RNAs” or “crRNAs”) capable of guiding the system to matching sequences of DNA. When the target DNA is found, Cas9 – one of the enzymes produced by the CRISPR system – binds to the DNA and cuts it, shutting the targeted gene off. Using modified versions of Cas9, researchers can activate gene expression instead of cutting the DNA. These techniques allow researchers to study the gene’s function.

Research also suggests that CRISPR-Cas9 can be used to target and modify “typos” in the three-billion-letter sequence of the human genome in an effort to treat genetic diseases



GENOME EDITING USING CRISPR TODAY

Trials of CRISPR-based treatment for hematological diseases such as SCD were particularly successful in 2022, says Guillermo Montoya, PhD, research director of the Protein Structure & Function Program at the University of Copenhagen.

Dr. Martin Steinberg, professor of medicine, Boston University Chobanian & Avedisian School of Medicine, Massachusetts, says exa-cel’s clinical success represents a certain milestone. “It is a landmark. Not a very long time ago, it was unbelievable that you could do such a thing around the expression of a gene that was mainly suppressed, and that will result in what appears to be a cure or a near cure,” he says.

According to results from the Phase II/III CLIMB SCD-121 trial in SCD, shared at last year’s ASH, all 31 treated patients remained free of vaso-occlusive crises. The median time from exa-cel infusion to last red blood cell transfusion was 19 days.

Exa-cel is not the only product in clinical development that captured the attention of experts in the field. The SCD therapy OTQ923, which is being developed by Intellia Therapeutics and Novartis is in Phase I/II study. Further study is warranted based on preliminary results from a study where two patients who were treated with OTQ923 saw improvements in their fetal haemoglobin levels, says Dr. Akshay Sharma, research associate at St. Jude Children’s Hospital in Memphis, Tennessee, an investigator on that trial.



REQUIREMENT OF MORE RESEARCH

An important achievement in the coming months will be a further strengthening of the number of CRISPR clinical trials, says Montoya. Testing CRISPR-based therapies in clinical trials for different diseases would show that the technology can be safe, he explains. This could then provide confidence in testing the treatment for more complex diseases, he notes.

Lignani’s research includes studying how CRISPR systems can be used to treat genetic conditions such as Dravet syndrome. The next milestone in this area will be conducting long-term toxicity studies, particularly in larger animals. According to Lignani, a big challenge for CRISPR-based treatments in the brain is the immune system’s response to Cas9 expression.

Developing CRISPR therapies for these complex indications remains a long-term goal. Montoya says the development of new approaches and technologies to target complex diseases where multiple genes are involved will be the next frontier. “That may be one of the remaining and long-standing challenges that we will have to address in the future,” Montoya says.





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