As the pipeline of CRISPR therapies grows, developers in this rapidly evolving field strive to improve gene editing accuracy and safety with new processes for removing faulty DNA sequences, known as “knock-out,” and inserting functional sequences, or “knock-in.”   

Laboratories developing CRISPR innovations can work with CDMOs from the earliest stages of discovery to accelerate these innovations. CDMOs can provide the tools needed to accelerate workflows and offer next-generation technologies that can advance gene-editing applications. CDMOs can help researchers establish efficient workflows and select the tools they need to smoothly transition from the benchtop to the clinic.

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Ways that CDMOs can benefit CRISPR developers

One priority in the CRISPR field, for example, is developing alternatives to the long-preferred method for DNA knock-in, homology-directed repair (HDR), which is precise but often inefficient. One alternative is microhomology-mediated end joining (MMEJ), a method of inserting the desired knock-in at the sites of double-stranded DNA breaks created by programmable nucleases. This method is up to three times more efficient than HDR, and allows for editing in many more phases of cell development than are possible with HDR. This could broaden the universe of diseases that can be corrected with gene editing.

Minimizing the risk of off-target editing is another priority for CRISPR developers. CDMO partners can help identify novel enzymes designed to reduce the CRISPR error rates. Alternatives to Cas9, the traditional enzyme used to cut DNA, include dCas9 fusions that can improve targeting and non-Cas9 nucleases that can expand the options for targeted cutting of DNA sequences. CDMOs can help select alternative enzymes, complete cGMP stability studies, and draft sections pertaining to the selected enzymes for regulatory filings.

CDMOs have also introduced innovations in CRISPR delivery, such as reduced-size plasmids used to transport CRISPR into target cells. These small plasmid DNA backbones, which do not include antibiotic markers, can lower the risk of toxicity and improve efficiency by reducing rates of transgene silencing. Genentech demonstrated these attributes in a 2022 study comparing the efficiency of three DNA donor templates for generating CRISPR HDR knock-in primary CD8 T cells. They reported that the smallest of the plasmid vectors generated twice the number of edited cells as did a traditional pUC plasmid and three times as much as a linear double-stranded DNA donor template.

Improving the methods used to deliver CRISPR therapeutics into patients is also a priority in the field. CDMOs are working with CRISPR developers on alternatives to the commonly used adeno-associated virus (AAV), which is limited in the size of the cargo it can carry and its ability to target tissues beyond the liver. Emerging alternatives include delivering CRISPR enzymes in mRNA form, which could improve efficiency and reduce off-target editing. CDMOs are also developing extracellular vesicles and virus-like particles, both of which could improve the ability to carry large CRISPR cargos into a variety of target cells beyond the liver.

Securing a CDMO partner

Partnering with a CDMO early in the development of a novel CRISPR therapeutic is crucial because the CDMO can tailor and test various options for editing technologies and delivery methods. Once a process has been selected, the CDMO will establish efficient laboratory workflows that will ensure a smooth transition from the benchtop to clinical trials to commercialization. While it may be tempting to use an off-the-shelf plasmid DNA extraction kit, for example, there may be better strategies. These kits often result in workflows that are not easily scalable and plasmids that are low-quality or not well-suited to the therapeutic.

Partnering with a CDMO to set up the ideal process from the earliest stages of development can prevent later issues with manufacturing and scale-up, which will save costs over the long run. And when those processes work well, the CDMO can help establish CRISPR platforms that developers can easily apply to new diseases—an idea that the FDA solidified earlier this year with a draft guidance for the Platform Technology Designation Program.

All these CRISPR innovations will ultimately be a boon for patients, too. After all, improving the accuracy, safety, and efficiency of gene editing with technologies that can be applied across a wide range of genetic diseases will bring these lifesaving therapies to patients faster.