CRISPR and stem cells to cure diabetes

In a molecular and cell transforming engineering feat researchers demonstrate for the first time a potential cure for genetic forms of diabetes. They take patient skin cells, turn them into stem cells, use CRISPR-Cas9 gene editing to correct a diabetes-causing mutation, and then transform the edited cells into the insulin-producing beta cells the patient lacks. Transplanting the beta cells into severe diabetic mice rapidly reverses their diabetes, providing a robust and functional cure.

»This technology now allows the potential use of gene therapy in combination with patients' own cells to treat their diabetes by transplantation of lab-grown beta cells,« says assistant professor Jeffrey Millman, Washington University School of Medicine, US.

He led the study in collaboration withphysician-scientist Dr. Fumihiko Urano, who treats patients with a disease called Wolframs syndrome.

»I have seen many patients of Wolframs syndrome, and I have been looking for a treatment for this devastating disorder. So this is great news,« says Urano.

The researchers acknowledge that more preclinical work is needed. Still, the therapeutic approach raises the possibility to cure Wolframs syndrome and other forms of diabetes caused by mutations in a single gene.

Diabetes is a loss of beta cells

Diabetes affects hundreds of millions of people globally, and the disease is caused by the loss or dysfunction of the insulin-producing beta cells in the pancreas. Functional loss of beta cells is generally treated with insulin injections, but long-term complications can arise.

Finally, they turned to new cocktails of growth factors and nutrients targetting several developmental pathways to make the iPSC stem cells differentiate into insulin-producing beta cells.

As a control they also turned patient-derived cells that had not been genetically modified into beta cells.

The unedited beta cells were also able to produce insulin, but at a much lower level. Measuring single-cell RNA sequences from several thousand edited and unedited beta cells, they observed major defects in the unedited beta cells, including very weak insulin secretion and many markers of stress and dysfunction.

»Amazingly this is all fixed in the gene-corrected beta cells, and they were indistinguishable from beta cells that we made from healthy donors,« says Jeffrey Millman.

Curing diabetic mice

For the final test, the researchers injected the CRISPR-corrected 'beta cells' into severely diabetic mice. These animals were rendered diabetic by a toxin injection (streptozotocin), which destroys the endogenous beta cells.

As controls, they injected unedited 'beta cells' and beta cells from human cadavers.

The edited 'beta cells' were superior to the unedited beta cells. They maintained stable insulin and blood glucose numbers for the remainder of the observation period up to six months.

»This is the first demonstration of the translational potential of patient-derived stem cells, let alone with CRISPR-correction of a diabetes-causing mutation. It now opens up the potential of using a patient’s own cells to treat their diabetes,« says Jeffrey Millman.

Off-targets not a big concern

One major concern of gene editing therapies is the risk of off-target mutations, which potentially can cause cancer.

»Fortunately, we did not observe any off-targets,« says Jeffrey Millman.

The researchers computationally predicted the five most likely genomic locations and sequenced those without finding any mutations.

Secondly, they saw no adverse effects in the mice after six months that would indicate off-targets, such as uncontrolled growth.

In translating the approach to humans, Jeffrey Millman doesn't believe off-target will be a significant safety risk, especially since the cells are edited ex vivo.

»I would imagine performing whole exome sequencing of gene-edited cells to confirm lack of off-targets in the protein-coding region of the genome. This would allow us to select only clones with the correct gene edits without off-targets, making off-targets in my opinion not a major concern in this context,« he says.