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Caribou Bio Nabs Patent, Lays Foundation for CRISPR-Based Dx


NEW YORK (GenomeWeb) – The US Patent and Trademark Office has awarded Berkeley, California-based Caribou Biosciences a new patent that points to potential diagnostic applications of CRISPR/Cas9 technology.

"It's one of a number of things we're investigating at the moment," Caribou CSO Andy May said, regarding diagnostic applications of CRISPR. "That's one of the areas we have an active effort in here."

May, who is listed as an inventor on US Patent No. 9,260,752, " Compositions and methods of nucleic acid-targeting nucleic acids," told GenomeWeb that the basic idea behind the patent is a method to detect proximal nucleotide sequences.

While the claim put forth in the patent is broadly worded, it is perhaps easiest to imagine the system described as two Cas9 nucleases, each fused to half of a split protein system, complexed with two single-guide RNAs targeting separate sequences. If the target sequences are adjacent, the fragments of the split protein will themselves join. The patent covers any method using site-directed polypeptides, but contains a specific claim where Cas9 is one or both of the polypeptides.

If the target sequences are chosen to detect a BCR-Abl translocation (aka the Philadelphia chromosome) ­— one claim set forth in the patent — and if the protein fragments comprise a split green fluorescent protein system — also a claim set forth — the possibility of a test for the Philadelphia chromosome emerges.  Fluorescent in situ hybridization- and qPCR-based tests detecting that particular translocation are used to diagnose and monitor chronic myeloid leukemia (CML).

Though the company had previously acknowledged its potential to develop CRISPR-based diagnostics, it was an area that had received considerably less attention than genome editing.

"We and everyone else are really excited of the prospect that you can make [genomic] manipulations in cells," May said, but he noted that Caribou was originally set up to look broadly at uses of CRISPR systems and how they might be commercialized. "You see all sorts of applications outside the cell, diagnostics being one of them," he said.

Given that Caribou filed the newly awarded patent in March 2014, it appears that diagnostics has long been a focus for the company, which was founded in 2011. 

May declined to elaborate on Caribou's research and development related to the patent and any diagnostics or other applications based on it. But the method affords Caribou to target any nucleic acid using any nucleic acid-targeting, nucleic acid-guided polypeptide. Moreover, the patent provides several split protein systems that could be engaged in the event of proximal sequence detection, including the aforementioned split GFP, split ubiquitin, split transcription factors, and split affinity tags.

The patent offers another flavor of combining split protein systems with CRISPR. In February 2015, scientists led by Feng Zhang of the Massachusetts Institute of Technology and the Broad Institute published a study on a split Cas9 protein that can be reassembled into an active form using rapamycin-binding dimerization domains. Also that month, scientists from Duke University published a study describing a toothless Cas9 fused to a vp64 transactivation domain and light-induced dimerizing protein pairs that enabled endogenous gene activation in the presence of light.

May declined to comment on whether Caribou was developing a diagnostic for BCR-Abl translocations. However, he acknowledged that the patented approach could be used to detect that and other gene fusions.

"Ultimately, businesses are built around products," May said. "We're continuing to do a lot of development work around systems with the longer-term view that we will be advancing products ourselves. [The patent] is all part of the process."