Nature Journal: Clinical trials of antisense oligonucleotide drugs for the treatment of ALS

ALS, also known as amyotrophic lateral sclerosis, is one of the five terminal illnesses recognized by the World Health Organization (WHO). It is a chronic, progressive degenerative disease that affects upper/lower motor neurons and the muscles that innervate the trunk, limbs, and head and face. It is often characterized by progressively worsening muscle weakness, atrophy, and bundle tremors caused by combined damage to upper/lower motor neurons.

ALS is a common type of motor neuron disease, with approximately 200000 ALS patients in China alone. The age of onset is mostly between 30-60 years old, with more males than females. At present, there are no specific therapeutic drugs for ALS, and the treatment mainly aims to improve the patient's quality of life and delay the progression of the disease. Physicist Stephen Hawking, "People's Hero" Zhang Dingyu, and Vice President of JD.com Cai Lei are all ALS patients.

Previous studies have shown that abnormal amplification of the six nucleotide repeat sequence (GGGGCC) in the C9orf72 gene is the most common genetic cause of ALS and FTD (accounting for 40% of familial ALS, 10% of sporadic ALS, and 25% of FTD). After the abnormal amplification occurs, it will interfere with the growth and expression of the C9orf72 gene, and also produce a neurotoxin protein called Dipeptide Repair Proteins (DRP).

On December 23, 2021, researchers from the University of Massachusetts Medical School published a research paper titled "Suppression of mutant C9orf72 expression by a potent mixed backbone antisense oligonucleotide" in the international medical journal Nature Medicine.

Antisense oligonucleotides (ASO) are artificially synthesized short single stranded oligonucleotides, which are a type of small nucleic acid drug. After entering cells, they bind to the complementary target mRNA sequence through the principle of base pairing under the action of ribonuclease H1, thereby inhibiting the expression of the gene and reducing protein production.

The working principle of ASO

So far, four ASO drugs have been approved by the US FDA, including three for the treatment of Duchenne muscular dystrophy (DMD) and one for the treatment of spinal muscular atrophy (SMA).

For a long time, scientists have confirmed that knocking out single gene mutations that cause neurodegenerative diseases may have therapeutic benefits, but safely delivering nucleic acid drugs to neurons is not an easy task. Moreover, while eliminating mutant proteins, it is also necessary to retain sufficient functional proteins, which is a major obstacle faced in treatment. Therefore, when treating ALS patients with C9orf72 gene mutations, it is not enough to simply knock out all C9ORF72 proteins from the patient's neurons.

The research team first tested the C9orf72 gene mutation in cells from ALS patients and transgenic mice, and constructed sequence optimized antisense oligonucleotides (ASOs) that can selectively inhibit the expression of transcripts containing GGGGCC repeat sequences and effectively suppress the levels of Dipeptide Repair Proteins (DRPs).

Then, the research team chemically modified ASO to improve its safety, distribution, and stability in the brain and spinal cord. Naked ASO can be directly injected through the spinal cord without the need for delivery carriers.

Next, the research team conducted a human clinical trial of the ASO drug on a patient with ALS carrying abnormal amplification of the C9orf72 gene GGGGCC.

The experimental results showed that ASO drugs significantly reduced the toxic protein dipeptide repeat protein (DRP) in the spinal fluid of the patient. During the trial, the patient's ALS functional score and other influencing indicators remained stable or slightly improved, and there were no neurological or medical adverse reactions after treatment.

This human trial demonstrates that ASO drugs can effectively and safely inhibit the abnormal amplification of the C9orf72 gene transcript, and significantly reduce the levels of toxic proteins expressed in the abnormal transcript. It is reported that this is also the first time that the level of dipeptide repeat protein (DRP) caused by abnormal amplification of the C9orf72 gene has been reduced in the human body. This study strongly supports the clinical benefits of inhibiting mutations in the C9orf72 gene for patients with ALS, and further exploration is warranted.

In May of this year, the journal Nature Medicine published a clinical report that found a rare mutation in the SPTLC1 gene of early-onset ALS patients, which leads to uncontrolled production of phospholipids and accumulation in human motor neurons.

The research team also conducted experiments using siRNA, and experiments on patients' skin cells showed that siRNA not only reduced the expression level of SPLTC1 gene, but also restored the level of phospholipids to normal.

These research results indicate that the use of precise gene silencing strategies such as ASO or siRNA is a highly promising approach for treating patients with ALS caused by gene mutations.