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Mission Bio Aims to Move Into CRISPR Applications; Customers Publish Single-Cell AML Data


SAN FRANCISCO (GenomeWeb) – Mission Bio, the South San Francisco, California startup developing droplet microfluidic single-cell analysis technology, is looking to move into the CRISPR space, harnessing its technology for quality control of CRISPR-based and other DNA editing technologies.

Last year, the firm launched its droplet microfluidic single-cell DNA analysis platform, Tapestri, which can process up to 10,000 single cells. It has also launched a research-use only panel targeting genes related to acute myeloid leukemia, as well as a kit that enables customers to design their own gene panels.

Today, Mission Bio said raised $30 million in a Series B financing round. Aside from continuing to scale up its presence in the blood cancer space, it is also looking to expand into CRISPR applications, according to CEO Charlie Silver.

Silver said that although the firm launched with a clear focus on cancer, over the last year, several of its customers, in particular pharmaceutical companies, have been interested in using the platform not only to detect cancer mutations but to identify edits that they're trying to insert into the genome. This includes companies developing CRISPR-based therapeutics and those working on other gene editing technologies. There has also been a bit of interest from academic researchers in studying how specific edits propagate and evolve through multiple generations of cells. 

"It's important to know as you're knocking in or out edits, how you're doing that, and the on-target and off-target effects," he said. "Even if you hit the right site, you need to make sure you've put the right mutation in." With Tapestri, customers can design a panel to analyze a set of specific mutations or genes to analyze edits at the single-cell level.

In addition, Silver said, the company joined the National Institute of Standards and Technology's gene editing consortium. This group aims to evaluate genome editing pipelines and develop benchmark materials and data. Silver said that the consortium will also work with the US Food and Drug Administration around developing regulations and guidelines for gene editing. He said that the Tapestri technology could be used to validate gene edits at the single-cell level. "We think there will be a large role for Tapestri to play in validating gene edits, specifically on the regulatory side," he said.

Currently, however, Mission Bio's main market is in oncology. Koichi Takahashi, previously an early-access user, who now has a Tapestri system installed in his lab at MD Anderson Cancer Center, said he has been using the system to study clonal evolution and assess minimal residual disease in acute myeloid leukemia.

Earlier this year, Takahashi, in collaboration with company researchers, described in a Genome Research study how they used Tapestri in two patients with AML to sequence 62 disease-related genomic loci across 16,000 single cells. Also, at the American Society of Hematology meeting earlier this month, Takahashi described using Tapestri to analyze 19 recurrently mutated AML genes in more than 300,000 cells from 76 AML samples taken from 68 patients.

Takahashi said that because the system is so high throughput, they were able to get around problems that plagued previous single-cell DNA sequencing experiments, such as allele dropout, that can "really affect the accuracy of interpretation." However, with data on 10,000 cells, "that dilutes those uncertainties," he said.

His group performed both single-cell analysis and bulk sequencing to first see whether the Tapestri platform picked up the same mutations as bulk sequencing, which he said it did. The benefit of single-cell sequencing, he added, is that unlike bulk sequencing, it is able to distinguish whether mutations with minor allele frequencies co-exist in the same cell or reside in different cells.

For example, Takahashi said that there were several patients who harbored both an NRAS and a KRAS mutation. There's been a theory that only one RAS family mutation is needed to give an oncogenic signal and that cells may in fact not even be able to survive with two mutations in genes from the RAS family. However, that's not been validated at the single-cell level. "Using this technology, we were able to clearly visualize that these two functionally redundant mutations were in fact affecting different cells," he said.

The finding raises other scientific questions, such as why certain sub-clones of cells acquire NRAS mutations while others gain KRAS mutations, or whether it is a random effect. Studying this process further could help shed light on the underlying biology and mechanism of the cancer, Takahashi said.

A second finding from the study was that the single-cell data helped to reconstruct the development and evolution of the disease. Using the single-cell data, Takahashi said, the researchers were able to piece together the evolutionary history.

Being able to reconstruct the phylogenetic tree from the first founder mutations enabled the researchers to identify two main branches that developed into leukemia from two different founder mutations through convergent evolution. Reconstructing the evolutionary history of cancer is another feature of single-cell sequencing that is more difficult with bulk sequencing, Takahashi said. Since bulk sequencing is less precise, it usually results in several different potential models. "There's a lot of controversy over how to pick the best model," he said, "but single-cell sequencing can really solve that problem."

Takahashi noted that while his work is focused on answering research questions about AML, it would eventually have clinical implications. For instance, with regards to understanding whether a single cell can carry two RAS mutations or only one, "clinically, we don't have data to act on that information, but the clinical arsenal always lags behind the science," he said.

Similarly, he noted that being able to reconstruct a more precise phylogenetic tree would have important implications for designing mouse models of disease and understanding disease progression, "how each mutation contributes to leukemia, in what order, and in what combination," he said.

The lab's next steps are to see whether Tapestri can be used to assess minimal residual disease. "We're actively working on that right now," he sai. In particular, he is hopeful that Tapestri could help in detecting FLT3 mutations, an important mutation for analyzing MRD which bulk sequencing struggles to identify. The FLT3 gene often has an internal tandem duplication involving a large insertion, Takahashi said, and "when the insertion is very large, it's challenging to call with NGS." The team has just started to design the experiments to test whether Tapestri will better be able to identify those mutations at the single-cell level. "We're gathering the MRD samples and trying to characterize the sensitivity and limit of detection of Tapestri," he said.