Biotechnology

Yale Unveils Cas12a-Based Breakthrough for Multi-Gene CRISPR Editing

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Expanding CRISPR’s Potential

Since its discovery, CRISPR (“Clustered Regularly Interspaced Short Palindromic Repeats”), which won the 2020 Nobel Prize in Chemistry, has revolutionized medicine and biotechnology.

Source: Nobel Prize

CRISPR enables targeted changes to specific genetic sequences, but it does not eliminate the risk of unwanted mutations. FDA guidance calls for assessment of off-target editing and other unintended genomic changes.

This is important as undirected gene insertion has been linked to major problems, notably cancer risks, making their therapeutic use difficult and controversial.

CRISPR can be used in multiple ways to interrupt a gene already present, delete a specific sequence, or edit/insert the right genetic sequence.

 

This turned into a medical breakthrough with the FDA approval for the first CRISPR-based therapy in 2023, developed by CRISPR Therapeutics for genetic blood diseases (follow the link for a dedicated report about CRISPR Therapeutics).

CRISPR is not limited to editing one gene at a time: multiplex Cas9 editing was demonstrated in 2013. Designing and delivering reliable multi-gene interventions nevertheless remains challenging.

The Yale study adds a Cas12a-based mouse toolkit for studying multiple genes together; it does not establish the first multiplex CRISPR method or demonstrate a treatment in humans.

It was a breakthrough study that brought together a massive team of researchers: no less than 37 persons are mentioned as authors of the study1 published in Nature Biomedical Engineering under the title “Cas12a-knock-in mice for multiplexed genome editing, disease modeling, and immune-cell engineering”.

It could help generate new disease and treatment models, such as genetic disease in the liver, lung cancer, and skin cancer, and accelerate further research in the field.

The Many CRISPR Systems

When CRISPR is discussed, either in its initial Nobel Prize-winning discovery or the already FDA-approved therapies, it is actually CRISPR-Cas9 that is discussed.

The “Cas” part of the system is the protein responsible for cutting the DNA strand and performing the gene editing itself, guided by a strand RNA. There are many other Cas proteins being studied, with probably many more to be discovered.

Source: YourGenome

It might even, in the long run, become commonplace for researchers to just design custom Cas systems, notably with “OpenCRISPR-1″, an open-source AI made to design a custom CRISPR system for specific situations. A computer simulation called CREME (Cis-Regulatory Element Model Explanations) might also help in understanding better how to optimize CRISPR systems.

Cas12a is another extensively investigated CRISPR system, with useful properties for multiplex editing.

CRISPR-Cas12a

Since we first discussed it in 2023, much progress has been made on Cas12a technology.

CRISPR-Cas12a is a different system from Cas9 in a few aspects:

  • Provides alternative cutting sites to what Cas9 can do.
    • hard-to-solve problems with Cas9 could be workable with Cas12.
  • It cuts DNA in a way that leaves a “sticky” section of DNA instead of the “blunt” cut of Cas9 and can be cut multiple times.
    • This results in a higher chance of gene editing.
  • There is no need for a transactivating crRNA (tracrRNA)for Cas12a, contrary to Cas9. Due to the smaller size, it would allow for easier multiplex genome editing.
    • More than one gene can be modified at once with Cas12a.

Source: Wikipedia

It is this latest characteristic of Cas12a that has sparked the most interest for biologists, as many diseases or genetic engineering require multiple gene editing, insertion, and/or deletion.

Both Cas9 and Cas12a can support multiplex editing. Suitability for a particular disease depends on delivery, editing efficiency and safety, rather than a categorical inability of Cas9 to modify multiple genes.

Meanwhile, it is becoming clear that CRISPR has potential way beyond curing rare genetic diseases, with neurology, cancer (oncology), and metabolic issues the most likely to improve or save the lives of tens of millions of people.

Source: ARK Invest

Some proposed therapies may benefit from modifying several genes, but the required approach depends on the disease and treatment design.

Yale’s Cas12a Study

Diverse Targets

Before performing multi-gene editing on humans, we first need to have a better understanding of the effect of such an aggressive intervention on the genome.

The Yale researchers used lab mice to genetically engineer immune cells, including  CD4+ and CD8+ T-cells, B-cells, and bonemarrow-derived dendritic cells. They also modified cancer-related genes and in-vivo liver tissues.

The CRISPR gene editing material was delivered to each target with retroviruses, adeno-associated viruses, and lipid nanoparticles, respectively.

In order to expand the possibility of the method, they also used 2 variants of Cas12a.

Checking Efficiency

First, the researchers associated the mutation with a fluorescent protein to check if it was being performed properly.

They then checked how the gene editing efficiency varied depending on the targeted organs. It appeared that the liver, brain, kidney, and lungs were among the easiest for achieving a high level of in-vivo gene editing.

This makes for very promising results, as these organs are the most important for the key medical targets of future CRISPR therapies.

Practical Potential

Once the concept had been proven to have potential, the researchers went on to test use cases of modifying immune cells and cancer cells.

When it came to immune cells, transformation efficiency, the metric measuring how much of the total exposed cells are genetically modified, was remarkably high, reaching up to 90-100%.

Another remarkable result, and an absolute requirement for any potential application of in-vivo therapies, is a very high rate of on-target mutation. This means that only the intended gene is being modified, and very few off-target gene editions occur.

DAKO

Pushing even further the possibility of CIRSPR-Cas12a technology, the researchers developed a novel dual-gene activation and knockout (DAKO).

This means that a single treatment can later on split and produce both the activation of one gene and suppression of another at once.

This is an important step for modifying cells with complex biochemical mechanisms, for example, one protein activating a mechanism and another one suppressing it. By performing DAKO gene editing, it becomes possible to both boost the desired activity beyond normal levels and remove the system that would inhibit this desired effect.

As unwanted or poorly managed retrocontrols are often a key reason for gene editing not performing as hoped, this DAKO system could prove to be a revolution, especially in oncology, neurology, and metabolic treatments.

Animal Models

This new gene-editing system using Cas12a can be as precise as Cas9 but can perform multiple edits at once, and those edits can be both the activation and suppression of genes.

This makes it a perfect system to create a new biological model on which biologists can test their hypotheses.

“These mice can facilitate rapid and seamless workflows for in vivo therapeutic gene targeting, disease/tumor modeling, and primary immune-cell engineering.

Combined with pooled oligo library synthesizing and cloning, these mice could be used for high-throughput CRISPR screening to study complex genetic questions, such as effector and memory phenotype transition in immune cells.

Such animal models are very important to test potential treatment, or better understand complex diseases. So even if no therapy using Cas12a ever reaches the market, this alone would mean that this CRISPR variant will have a large impact on future biotech research.

Going Further

As it was proven in the study that the intensity of the gene editing can vary greatly depending on the organ, it is likely that further improvements can be achieved.

A key element will likely be testing new vectors for the gene editing system, either other viruses or other lipid nanocapsules. In particular, better vectors for the organs the least affected will be required for specific studies or diseases.

In the long run, it is most likely that gene editing therapies for complex diseases like cancer and neurological issues will use a system more akin to this cas12a system than the “traditional” mono-gene Cas9.

Cas12a Company

Editas

EDIT Price Chart

Editas was founded by CRISPR-Cas9 co-discoverer Jennifer Doudna. Editas started working with Cas9 but is now focused on a proprietary version of Cas12a that they engineered: AsCas12a.

You can read more about Cas12a’s unique properties in our dedicated article “What Is CRISPR-Cas12a2? & Why Does It Matter?”.

Source: Editas

You can also read an overview of all of Jennifer Doudna’s companies in the corresponding article “Top Jennifer Doudna Companies to Watch.”

Editas is focused on Sickle Cell Disease (SCD) and beta-thalassemia, 2 diseases where it lost the race for first treatment approval to competitors CRISPR Therapeutics and BlueBirdBio.

Overall, the SCD program (recently renamed reni-cel) has been delayed several times, sparking concern among investors, and has since been refocused on in-vivo therapy to distinguish it from already approved SCD therapies.

Nevertheless, Editas owns significant patents on CRISPR-Cas12, which has been used by researchers at the University of New South Wales, Australia, to develop a COVID-19 strip test, illustrating the technology potential beyond gene editing.

Editas also signed in 2023 a $50M deal with Vertex for the company to use Editas’ Cas9 IP.

Editas focuses on other CRISPR versions than the “classical” CRISPR-Cas9 and its research IP might come in handy in establishing partnerships and generating revenues without an FDA-approved product, on top of a cash runway going into 2026.

As Cas12a seems to become increasingly proven as a best-in-class method for multi-gene editing, Editas’ expertise and pipeline focus on this CRISPR variant might prove a winning bet in the long run.

(You can read more about CRISPR companies in our corresponding article “Top 5 CRISPR Companies To Invest In”.)

Latest on Editas

Studies Referenced:

1. Tang, K., Zhou, L., Tian, X. et al. Cas12a-knock-in mice for multiplexed genome editing, disease modelling and immune-cell engineering. Nat. Biomed. Eng (2025). https://doi.org/10.1038/s41551-025-01371-2

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Jonathan is a former biochemist researcher who worked in genetic analysis and clinical trials. He is now a stock analyst and finance writer with a focus on innovation, market cycles and geopolitics in his publication 'The Eurasian Century".