Milasen: The Drug Designed for Only One Person
By Natalie Bratset
-Featured Image by Zöe Petroff
As the complexity of rare genetic diseases becomes more apparent with major scientific advancements in whole-genome sequencing, patient-customized drugs are becoming a reality to treat individuals who have never been treatable before. Personalized medicine is a broad term that encompasses the use of a patient’s genetic or biological information when making medical and treatment-related decisions.1 Patient-customized drugs take this a step further by using the patient’s genetic information to create a custom treatment that directly targets the cause of the disease. In 2019, a groundbreaking “N-of-1” study described the timeline of how a drug—made for only one person—was designed, produced, and administered in record time.
A SERENDIPITOUS REFERRAL
In November 2016, a 6-year-old girl named Mila was admitted to the hospital with symptoms of blindness, developmental regression, and seizures. Shortly after, she was diagnosed with Batten’s Disease, a rare neurodegenerative disorder. This stops the lysosomes—our cellular trash cans—in Mila’s brain cells from breaking down waste. Rapidly filling with cellular byproducts, the lysosomes then burst, ultimately killing her brain cells.2 Batten’s Disease is also classified as an orphan disease, which is typically a very rare disease that has not been adopted, so to speak, by the pharmaceutical industry. Because it provides little financial incentive, corporations scarcely aim to create new medications to treat or prevent orphan diseases.3
Mila’s form of Batten’s Disease was particularly hard to treat because her doctors could not find all of the mutations in the MFSD8 gene, which causes this disease. Jinkuk Kim and his team from the Division of Genetics and Genomics at Boston Children’s Hospital heard of Mila’s puzzling case and were officially referred in mid-January of 2017. Two months later, the mutation was discovered and an additional RNA splicing defect was identified.2 Less than one year later, Dr. Kim and his team developed and acquired FDA approval for the first splice-modulating antisense oligonucleotide drug designed for one person: milasen. Let’s break that down.
WHAT IS RNA SPLICING?
Your DNA has sections of both protein-coding and non-protein-coding DNA—exons and introns. These segments of exons and introns alternate in your genes and must be precisely cut and pasted back together to make a functional protein. That is where splicing comes in! When a blueprint of your DNA is made, this pre-messenger RNA (pre-mRNA) has all of the introns and exons from the gene. The spliceosome is a massive complex that performs and regulates splicing in eukaryotic cells. It recognizes which parts are introns, cuts them out of the pre-mRNA, and stitches the exons back together to make the mRNA used to create proteins. But how does the spliceosome know where introns start and end? Two segments of DNA in exons, called Exonic Splicing Enhancers (ESEs) and Exonic Splicing Silencers (ESSs), are the genetic markers used to indicate where to splice. ESEs mark the beginning of the exon, and ESSs mark the end of an exon. The spliceosome can recognize these two sequences and start splicing out the intron.
As you could probably guess, this is an incredibly complicated process that involves a lot of moving parts, which makes it easy for something to go wrong. When a different combination of exons and introns is spliced together than the standard combination, this is called Alternative Splicing.4 Alternative splicing is not necessarily always disease-causing and, in some cases, it can produce different versions of the same protein. However, when a splicing issue becomes disease-causing, this can have dramatic effects on the protein and the cells making the protein.
In Mila’s case, she had a mutation in MFSD8 that created an ESE in the middle of intron 6. This caused the rest of intron 6 to look like the beginning of an exon. The spliceosome recognizes this new ESE and leaves a portion of intron 6 in the mRNA, which is then translated into a non-functional protein. To complicate things, intron 6 also had a Sine Variable-Non-Tandem-Repeat Alu (SVA) insertion, which is a large region of repeating DNA that makes intron 6 significantly longer.2 All of this adds together to make a large, non-functional protein in Mila’s lysosomes, causing her Batten’s Disease. Dr. Kim and his team wanted to design something that would be able to change or inhibit the defective splicing of MFSD8 to stop the defective protein from being made, and that is where the antisense oligonucleotide comes in.
An oligonucleotide is a general name for a short chain of nucleic acids, like DNA or RNA, that usually can be from 8 to 50 nucleotides long. What makes an Antisense Oligonucleotide (ASO) special is that its job is to bind to the pre-mRNA in your cells and stop that portion of RNA from being turned into proteins, by covering a splice site, and to fix the splicing error.
This idea came from a novel drug that was FDA approved in 2016 called nusinersen (or Spinraza commercially) to treat Spinal Muscular Atrophy (SMA).5 SMA, like Batten’s Disease, is also a neurodegenerative disease that causes the degradation of motor neurons in the brain and spinal cord, resulting in the loss of many motor functions. With a median life expectancy of two years, it is extremely fatal in children. 90% of SMA cases are caused by a specific splicing error that skips exon 7, essentially killing the protein.5 If injected as soon as an infant is diagnosed with SMA, nusinersen can substantially increase the life expectancy and decrease the neuromuscular damage in these children.
Building off of this successful ASO, milasen uses an RNA modification called 2′-O-methoxyethyl (2′-MOE)—a modification that adds an extra carbon chain with one oxygen in it to the 2’-OH of the RNA molecule. The 2’-MOE modification increases the binding affinity of this ASO to RNA, essentially locking it to the pre-mRNA of MFSD8.2 The ASO is designed to pair with the mutated ESE in intron 6, making the spliceosome skip over this faulty ESE, splice out intron 6 with the SVA region, and form a correct mature mRNA.
Drug design is a complicated and lengthy process, so a ton of iterations of ASOs were tested in cells by using two programs called RESCUE-ESE and ESEfinder to predict exactly where this mutated ESE was in Mila’s DNA sequence.2 The different sequences were then injected into copies of Mila’s cells and the mRNA produced was extracted to be sequenced. By comparing the ratios of properly spliced (E6-E7) to improperly spliced (E6-i6) mRNA, the ASO that we now know as milasen more than tripled the amount of normal E6-E7 splicing.2 After a couple more in-depth experiments in human cells, milasen was ready for human trials.
THE CLINICAL TRIAL AND ITS RESULTS
So, what happened after milasen was FDA approved? In January 2018, Mila’s condition worsened substantially, so Dr. Kim and his team received permission to initiate clinical investigational treatment by the FDA.2 Mila received the first dose at the end of January and was carefully monitored throughout her treatment. Two main factors were tracked during Mila’s dosing schedule: cerebral spinal fluid conservation and seizures.
Milasen is injected into the spinal cord, which means that cerebral spinal fluid (CSF) conservation is a good indication of Mila’s body accepting or rejecting the drug. Overall, the drug had a good CSF conservation rate, which was a monumental step towards a super effective treatment. The second metric measured was seizures. Seizures were a regular occurrence for Mila, happening 10-30 times a day. They were a super impactful part of her life and often impeded Mila’s ability to do things. The seizures varied in intensity and duration but trended towards longer seizures before treatment. Over the 300-day dosing schedule, Mila’s seizures decreased substantially both in duration and severity! This was a very promising result from treatment with ASOs, especially because Mila began to start talking, laughing, and feeling much better.
Tragically in February 2021, two years after the paper was published, Mila passed away. Her form of Batten’s Disease was already too advanced, and although the treatment significantly reduced symptoms, the damage was irreversible.6
What is the silver lining of Mila’s story? Batten’s Disease is just one of the estimated 7,000 rare diseases that we know of, and only a few rare diseases are tracked when a person is diagnosed. As far as we know, up to 30 million Americans live with a rare disease.3 Yet, these diseases do not have treatments because they are often unique to the patient, so corporations do not see financial incentives in developing drugs for rare diseases. Three in ten children who have a rare disease will not live to see their fifth birthday.6 In the last couple of years, there have been promising advances in whole-genome sequencing to the point where we can get patient sequences shortly after and sometimes before diagnosis. We are at the forefront of a revolution in patient-customized medicine. I hope that Mila’s groundbreaking, custom treatment will provide some scaffolding and background for future cases of patient-customized medicine. I look forward to a future where people like Mila will be able to get a custom medicine shortly after being diagnosed with a rare disease, making these rare diseases much less fatal.
1. Zineh, I. (2016, February 26). Personalized medicine: A biological approach to patient treatment. Federal Drug Administration. Retrieved from https://www.fda.gov/drugs/news-events-human-drugs/personalized-medicine-biological-approach-patient-treatment.
2. Kim, J. & et. al. (2019, October 24). Patient-Customized Oligonucleotide Therapy for a Rare Genetic Disease. The New England Journal of Medicine. doi: 10.1056/NEJMoa1813279.
3. (2021, January 26). FAQs About Rare Diseases.NIH – Genetic and Rare Diseases Information Center. Retrieved from https://rarediseases.info.nih.gov/diseases/pages/31/faqs-about-rare-diseases
4. Greenberg, D.S. & et. al. (2013). Alternative Splicing. Brenner’s Encyclopedia of Genetics. Retrieved from https://www.sciencedirect.com/science/article/pii/B9780123749840000437
5. Chen, I. (2019, November 14). An antisense oligonucleotide splicing modulator to treat spinal muscular atrophy. Nature Portfolio. Retrieved from https://www.nature.com/articles/d42859-019-00090-4 6.
6. Whitlock, J. (2022, February 9). A mother, shaped by tragedy, embarks on a mission to advance custom medicines. STAT. Retrieved from https://www.statnews.com/2022/02/09/julia-vitarello-mila-rare-diseases-custom-drug-development/