Promoting Shorter HTT Protein Production Shows Promise in Mice

Approach may maintain full-length HTT function, prevent toxic fragments from forming

Patricia Inácio, PhD avatar

by Patricia Inácio, PhD |

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huntingtin protein | Huntington's Disease News | illustration of a mouse model

ProQR Therapeutics’ experimental therapy QRX-704 significantly reduced levels of the toxic huntingtin (HTT) protein clumps that fuel Huntington’s disease in a mouse model of the disease, a study showed.

The therapy, an antisense oligonucleotide (ASO) designed to promote the generation of a shorter, but working version of the HTT protein, also resulted in signs of improved nerve cell function. No significant differences were observed in the animals’ behavior, however. The researchers said these may require a longer follow-up.

These findings suggest approaches designed to boost producing the shorter HTT protein, which is resistant to the cleavage responsible for forming its toxic fragments, may have therapeutic potential for Huntington’s.

The study, “A pathogenic proteolysis–resistant huntingtin isoform induced by an antisense oligonucleotide maintains huntingtin function,” was published in JCI Insight.

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Huntington’s disease is caused by mutations in the HTT gene, which leads to a longer-than-normal HTT protein being produced. The mutated protein (mHTT) is cut by the caspase 6 enzyme into toxic fragments that stick together and accumulate inside nerve cells in the brain, eventually leading to their death.

Suppressing the HTT protein cleavage may be a way to prevent these toxic fragments from forming and slow Huntington’s progression. However, since the lack of a working HTT protein exacerbates the disease, such efforts should also work to keep the full-length HTT’s natural functions.

The HTT gene can generate alternative forms of HTT through a process called alternative splicing. Much like in a recipe, adding or removing certain key ingredients — in this case, pieces of genetic information — from the gene’s messenger RNA (mRNA) can change the resulting protein. Of note, mRNA is the molecule derived from DNA that guides protein production.

One protein, called HTT delta 12, lacks a region near the disease-associated cutting site, suggesting this shorter version may be resistant to caspase 6-induced cleavage.

Short HTT protein similar to full-length HTT

Researchers from Korea, Sweden, and France, in collaboration with ProQR Therapeutics, evaluated whether this shorter version maintained functions of the full-length HTT protein and if promoting its production eased signs of the disease in a mouse model.

The researchers first confirmed that HTT delta 12 was resistant to cleavage by caspase 6, in contrast to the full-length HTT protein. Then they discovered that the shorter version retained similar structural and biochemical properties to the full-length HTT, suggesting it could exert similar functions.

Because the HTT protein is key for brain embryonic development, the team tested whether mice genetically modified to produce an HTT delta 12 similar to humans developed normally.

No brain-related or behavioral differences were found between mice producing HTT delta 12 and those generating the full-length HTT. Lab-grown nerve cells from mice with the shorter HTT version showed no impairments in trafficking proteins across nerve cell fibers, including the brain-derived neurotrophic factor (BDNF).

HTT is known to be involved in this transport process and the progressive loss of BDNF, a factor that helps nerve cells grow and survive, is linked to Huntington’s progression in mouse models.

Further experiments were conducted in lab-grown connective tissue cells from these mice to assess changes in two other HTT functions: maintenance of the structure of the cell’s protein-sorting factory (called Golgi complex) and cells’ ability to form cilia, fingerlike projections that help move fluid across tissues, including that surrounding the brain and spinal cord.

No differences in these functions were seen between mice producing HTT delta 12 and those generating the full-length HTT. These findings suggest the cleavage-resistant HTT protein retained the full-length HTT protein’s neuroprotective functions.

Experimental therapy QRX-704’s effects

Finally, the team evaluated the therapeutic potential of promoting the production of HTT delta 12 in a mouse model of Huntington’s disease by administering QRX-704 directly into the animals’ brains.

ProQR’s QRX-704 is an experimental ASO designed to activate the specific splicing site in HTT’s mRNA that promotes HTT delta 12 production. Results showed QRX-704 was well tolerated, but resulted in different levels of HTT delta 12 mRNA across treated mice.

When looking at mice producing high levels of HTT delta 12 mRNA, the researchers found they showed significantly lower levels of HTT toxic fragments and clumps. In these mice, the number of dendritic spines — nerve cell structures involved in signal transmission and that are reduced in Huntington’s — were also significantly increased.

Despite this, QRX-704 was not associated with significant behavioral changes, “which may require long-term observation,” the researchers wrote.

Overall, “we have demonstrated a pharmacologically feasible ASO-based therapeutic approach, generating an alternative HTT [form], which is structurally, biochemically, and functionally intact in important neuroprotective HTT functions but is resistant to caspase-6 cleavage,” the researchers wrote, adding the approach might treat Huntington’s disease by maintaining full-length HTT functioning and preventing toxic HTT fragments from forming.