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Inscripta Confirms CRISPR-MAD7 Editing Activity in Mammalian Cells, Releases New MAD2 Enzyme

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NEW YORK (GenomeWeb) – When gene-editing technology company Inscripta announced last December that it was releasing a new CRISPR enzyme it had created free of charge to the research community, the response was "overwhelming," according to CEO Kevin Ness.

The enzyme, called MAD7, is part of the so-called Madagascar family of novel RNA-guided nucleases that Inscripta is developing. MAD7 was initially characterized in Saccharomyces cerevisiae and Escherichia coli, and Inscripta was working with several partners in the research and business communities to characterize the full breadth of its editing abilities.

"We're an emerging company, and we have this really important asset, and we had a choice — are we going to hoard this internally and spend a while fully characterizing it across the literature for three or four years? Are we going to build that capability in-house and then get it out? Or can we work with the research community to get this out in this unrestricted manner and get people [to work] with it immediately, use it, test it, improve it, and feed the features back to us that they want improved?" Ness told GenomeWeb in March.

Inscripta chose option two, deciding to not only build a customer base, but to also use the expertise of that customer base to improve its own product and give the customers what they want.

Now, that choice seems to have paid off — the company announced late last week that the enzyme can indeed edit mammalian cells. MAD7 "is able to be expressed as an active protein in human HEK293T cells, and when combined with chemically synthesized guide RNAs targeting multiple genes, can edit several genes at multiple loci," the company said in a technical note.

When targeting the PPIB or DNMT3B genes, Inscripta showed that MAD7 editing resulted in high-efficiency cleavage in an in vitro cutting assay. The company also showed that MAD7 editing in the same cell line resulted in the formation of indels in an in vivo DNA mismatch detection assay.

The firm said it collaborated with multiple partners for its characterization experiments, but particularly pointed to its partnership with Horizon Discovery, whose transfection reagents and synthetic guide RNAs were used in the editing assays. In a statement, Horizon CSO Jon Moore noted that the potential commercial applications for MAD7 are diverse.

"What's beautiful about [the in vivo experiment in particular] is that it worked right out of the box with zero optimization," Ness said. "We're ecstatic about the potential of where it could go with even little bits of optimization, let alone with a real [R&D] program surrounding it."

This development also answers a question that researchers have been asking Inscripta since the company released MAD7 last December — does it work in mammalian cells?

"Eight months ago, when we started on this odyssey, the response was overwhelming. But we kept hearing two questions. Do you have IP protection, and does it work in mammalian cells?" Ness said. "And so we're excited about this latest release because it's addressing those two major questions."

He further noted that if the data had shown that the enzyme didn't work in mammalian cells, Inscripta would have reengineered it. "But luckily the MAD7 performed really well, and we had multiple groups give us feedback in a broader microbe set, in plants, and in mammalian cells," Ness added.

Regarding IP protection, the company said that the US Patent and Trademark Office has granted it a patent covering systems using MAD7. It's one thing for Inscripta to say this is a novel system, Ness noted, but for the USPTO to say it gives researchers confidence that this is indeed a new and novel enzyme that they can work with without fear of stepping on the intellectual property of another company or university.

The company also announced that it has released a new enzyme in the Madagascar family called MAD2, and that this enzyme has also received a patent from the USPTO. Inscripta is staying tight-lipped about how MAD2 works, but Ness said customers can reach out to the firm if they want access to it. The company has also chosen to not make MAD2 available for free at this point, as it did with MAD7.

But Ness also emphasized that Inscripta is offering far more than just two enzymes. Although he declined to disclose the exact number of MADzymes the firm has available at this point, he said it's "not a small number," adding, "We have a whole family [of enzymes], and if we don't have what people need, they can use our bespoke [enzyme engineering] program."

He also noted that the suite of MADzymes that are currently available have been fully characterized and tested, "and we know fully what they do," but said the firm's enzyme engineering pipeline also contains a whole host of candidate enzymes that are at various stages of characterization. Further, Ness added, the company's partners in the research and business communities are continuing to experiment with MAD2 and MAD7 in order to explore their potential — some doing head-to-head comparisons of the two enzymes and some working with each individual enzyme on their own.

Importantly, Ness noted, these developments are another step in getting Inscripta to its ultimate goal: creating a full suite of gene editing tools, including instruments, software, and reagents that all work together "to do the next-generation of gene editing experiments that can't even be done today."

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