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Researchers Use Gene Editing to Improve Red Blood Cell Production From Stem Cells

NEW YORK (GenomeWeb) – A team led by researchers from Boston Children's Hospital and the Dana-Farber Cancer Institute has published new data showing that silencing a single gene can significantly improve the yield of stem cell-derived red blood cells (RBCs).

The findings suggest that this approach could one day be used to bring down the cost of lab-manufactured RBCs for transfusions and other clinical applications, as well as improve the creation of specific cell populations from stem and progenitor cells, according to Vijay Sankaran, a Boston Children's researcher and senior author of the study.

The technique may also prove valuable in the near term as a tool to those developing RBCs as drug-delivery vehicles, he said, adding that it has already been non-exclusively licensed to one company — startup Rubius Therapeutics — for such an application.

While it is possible to induce various types of stems cells to differentiate into clinical-grade RBCs, doing so is prohibitively expensive, costing as much as $15,000 per unit of blood, Sankaran told GenomeWeb this week. Given his lab's focus on the role of human genetic variation on normal and dysfunctional blood cell production, "we reasoned that maybe … we could improve upon this process." 

To do so, Sankaran and his colleagues focused on a gene called SH2B3. Notably, genome-wide association studies by a variety of research groups have shown that a common coding single-nucleotide polymorphism in this gene is significantly linked with hemoglobin levels and RBC counts in vivo.

With rare loss-of-function SH2B3 alleles having been recently associated both with elevated both RBC and elevated hemoglobin levels, Sankaran's team tested whether suppression of the gene can enhance erythropoiesis in vitro. The researchers designed SH2B3-targeting shRNAs to adult CD34+ hematopoietic stem and progenitor cells (HSPCs), then induced the cells to undergo erythroid differentiation.

They found that knocking down the gene with the RNAi molecules not only caused the cells to undergo differentiation more readily, but also increased RBC yield three- to five-fold over control HSPCs. Experiments in HSPCs derived from umbilical cord blood had even more pronouned results, with SH2B3 inhibition leading to a five- to seven-fold increase in RBC expansion and yield versus controls.

Recognizing that their RNAi approach would be difficult to implement on a large scale, the researchers turned to the genome-editing technology CRISPR/Cas9 to permanently inactivate SH2B3 in in human embryonic stem cell (hESC) lines, which can readily be renewed and differentiated toward the erythroid lineage using existing protocols. After treatment with factors that promote blood cell production, the edited hESCs produced three times more RBCs than control cells.

Importantly, there were no significant differences between the RBCs derived from either the RNAi- or CRISPR-modified cells and controls.

In their study, which appeared in Cell Stem Cell, Sankaran and his collaborators concluded that suppressing SH2B3 could enable the production of RBCs from HSPCs at one-fifth the current cost and with fewer starting stem cells. Still, it is unlikely that this approach can be implemented in the near future.

"The protocols we have for pluripotent stem cell differentiation and the availability of self-renewing cell lines is not quite optimized to allow for true mature red blood cell production," Sankaran said. "But certainly I think this allows us to say we can do this perturbation stably in pluripotent stem cells, and this potentially could be a great source for material in the future as we get better at … producing the right kind of cells."

Additionally, the data point to the possibility of someday using genome editing to produce cells other than RBCs for applications in regenerative medicine and cell therapeutics, he added. "You can imagine that, if we could figure out ways of expanding or [enhancing] hematopoietic stem cells, for example, that could change the way we're able to perform bone marrow transplants."

But Sankaran sees near-term applications for his approach, as well.

"Red blood cells are a great delivery vehicle," with a number of companies modifying them to carry therapeutic payloads, he said. But "one of the issues they are having is that it is very costly [as they can only] produce a limited number of cells. Getting more cells allows them to improve upon the process."

The intellectual property around Sankaran's work has already been licensed from Boston Children's by Rubius, he said, and it will "likely be used by other companies that are trying to use these red blood cells." 

Officials from Rubius declined to comment for this article.