Preclinical research has revealed a potential treatment strategy for dyskeratosis congenita (DC).
Researchers found that DC is characterized by reductions in telomerase, telomere length, and telomere capping, which reduces Wnt pathway activity, resulting in intestinal stem cell failure.
However, treatment with Wnt agonists restored the Wnt-telomere feedback loop and reversed gastrointestinal DC phenotypes in vitro and in vivo.
Christopher J. Lengner, PhD, of the University of Pennsylvania in Philadelphia, and his colleagues reported these discoveries in Cell Stem Cell.
“Right now, the main therapy for [DC] patients is a bone marrow transplant,” Dr Lengner said. “That can address the bone marrow failure but doesn’t fix other problems associated with the disease, and especially not the risk of cancer. This work suggests a way to address the underlying cause of the disease.”
Earlier research with mouse models of DC suggested there might be a connection between the Wnt pathway and telomerase. And a recent study in DC patients’ cells revealed a decrease in activity in the Wnt pathway.
So Dr Lengner and his colleagues wanted to explore whether activating Wnt could reverse the effects of the disease. To do so, the team used induced pluripotent stem cells (iPSCs), the CRISPR/Cas9 gene-editing system, and directed differentiation.
The researchers generated iPSCs from DKC1-mutant fibroblasts and from wild-type cells. The team also used CRISPR to introduce a DKC1 mutation into healthy human iPSCs and to correct the disease-causing mutation in iPSCs generated from DC patient samples.
The researchers then grew organoids through directed differentiation. iPSCs were coaxed to form a human intestinal organoid, which naturally forms a tube-like structure, recapitulating the tubes of the human gastrointestinal system.
When the researchers observed the development of intestinal organoids, they found that, initially, the DC cells seemed to form normally.
The original DKC1-mutant cells and the cells that had the DKC1 mutation introduced by CRISPR appeared to follow a normal course of development for several days. But by 2 weeks, they lacked the tube-like structure seen in the healthy samples and the disease-corrected samples.
The DKC1-mutant cells also had shorter telomeres, with the intestinal organoids from DC patients having the shortest of any cell type.
“We could see, at the molecular level, that this is accompanied by a failure to activate specific intestinal stem cell gene programs—specifically, genes in the Wnt pathway,” Dr Lengner said.
The next logical step was to activate Wnt to see if these defects could be reversed. The researchers treated organoids derived from DC patient iPSCs with a compound called CHIR that stimulates the Wnt pathway.
This restored the formation of the tube-like structure as well as intestinal stem cell gene expression. The treatment also increased telomerase activity and telomere length in the cells with mutant DKC1.
To assess this treatment approach in a more clinically relevant model, the researchers transplanted the human intestinal organoids into mice.
Mice that received a transplant containing the DKC1 mutation and received treatment with lithium, a stimulator of the Wnt pathway, maintained their intestinal tissue structure and had high expression of Wnt target genes.
In effect, these mice resembled the mice that received a transplant of an organoid derived from a healthy patient.
The researchers said this study offers proof of principle that activating the Wnt pathway can reverse at least the gastrointestinal phenotypes associated with DC. Looking ahead, the team would like to try accomplishing the same feat in other tissue types affected by the disease.
