Ionis Pharmaceuticals, Inc. (formerly Isis Pharmaceuticals, Inc.) announced today that it has launched a Phase 1 clinical trial for IONIS-DMPKRX. Ionis earned a $14 million milestone payment from Biogen Idec associated with this achievement. IONIS-DMPKRX is designed to reduce the production of toxic dystrophia myotonic-protein kinase (DMPK) RNA in cells, including muscle cells, for the treatment of Myotonic Dystrophy Type 1 (DM1).

"[IONIS]-DMPKRX is an example of the broad applicability of our antisense technology to develop novel drugs to treat patients with severe and rare disease. IONIS-DMPKRX is the first drug to enter our pipeline that is designed to target a toxic RNA, the first systemically administered drug to enter development from our Biogen Idec partnership and the second generation 2.5 drug to enter clinical development," said C. Frank Bennett, Ph.D., senior vice president of research at Isis.  "Myotonic dystrophy represents an ideal opportunity for antisense as the disease-causing gene produces a toxic RNA, which is not accessible by traditional therapeutic approaches but is uniquely accessible with our antisense technology. We look forward to rapidly advancing the development of IONIS-DMPKRX."

"Our collaboration with Biogen Idec has been very productive. [IONIS]-DMPKRX has rapidly advanced to the clinic, and we continue to make progress across the board in our drug discovery programs with Biogen Idec. All of these successes advance our neuromuscular disease franchise and translate into the potential for significant revenue as our drugs and programs progress," said B. Lynne Parshall, chief operating officer at Isis.

DM1 is a rare genetic neuromuscular disease characterized by progressive muscle atrophy, weakness and muscle spasms. DM1, the most common form of muscular dystrophy in adults, affects approximately 150,000 patients in the US, Europe and Japan. Patients with DM1 have a genetic defect in their DMPK gene in which a sequence of three nucleotides repeats extensively, creating an abnormally long toxic RNA, which accumulates in the nucleus of cells and prevents the production of proteins needed for normal cellular function. The number of triplet repeats increases from one generation to the next, resulting in the possibility of more severe disease in each subsequent generation. There are currently no disease-modifying therapies that address more than one symptom of the disease. IONIS-DMPKRX is designed to improve the underlying genetic defect that causes DM1.

"Myotonic dystrophy is a progressive and debilitating disease that affects thousands of patients for whom there are no direct therapeutic options. The innovative science behind IONIS-DMPKRX is compelling and targets the underlying genetic defect that causes myotonic dystrophy," said Molly White, executive director of MDF. "IONIS-DMPKRX has a chance to fill the therapeutic void for DM1 patients and transform the hopes and futures of thousands of patients and families."

07/09/2014

Myotonic community members often contact the Foundation with questions about genetic testing. Their questions range from how to find facilities that conduct genetic testing for myotonic dystrophy (DM) types 1 and 2 to whether or not they should be tested and how to assess the benefits and risks of having a genetic test.

The question of whether or not to be tested is best answered in partnership with a physician or genetic counselor who understands your family history, your personal circumstances and reasons for exploring genetic testing. Implications associated with DM genetic testing are financial as well as emotional. For this reason, the question of whether to get genetic testing for DM should be considered carefully.

We spoke with Carly Siskind, MS, LCGC, senior genetic counselor on the Stanford University Neuromuscular Disorders Team about the pros and cons of testing and the impact it can have on issues such as insurability.

What do you do as a genetic counselor?

Usually people are referred to me by a specialist such as a neurologist. Sometimes individuals call my office directly. My job is two-fold: to provide basic information about genetics and the implications of testing, and to be the person between the doctor and the patient to help interpret “doctor speak.”

I’d like to emphasize that while genetic testing can provide definitive answers with regard to the presence of disorders like DM, the decision of whether to pursue genetic testing is a personal one with significant potential impacts, both positive and negative. I encourage anyone considering genetic testing to consult with his or her physician or qualified medical professional first, and to consider working with a genetic counselor; we can be helpful in navigating this process.

Do most people understand how DM gets passed along?

Most people I’ve met with to discuss myotonic dystrophy are already fairly well educated on the topic, but I find it helpful to go through some of the basics nonetheless. DM is a complicated disease; for instance, if someone has not inherited the CTG (DM1) or CCTG (DM2) repeat expansion and both copies of this gene—one from each parent—are in the normal range, then he or she won’t develop DM or pass it on to future generations. Only if someone has the expansion will that person potentially experience symptoms or risk passing DM on to his or her children.

What are some of the pros and cons of genetic testing for DM?

If a person isn’t experiencing any DM symptoms but has a family history of DM, it is important to consider the consequences of being tested. Insurability is probably the biggest concern. Even if someone doesn’t have symptoms or a DM diagnosis, if his or her genetics indicate that there is potential to develop DM in the future, it can—and probably will—impact that person’s ability to secure life, disability and long term care insurance. While it is sometimes possible to get this insurance with a family history of DM, it is nearly impossible to get it with a diagnosis, and a diagnosis might still delay access to health insurance.

Because of this, we generally recommend that people think about setting up insurance policies prior to being tested in case their test comes back positive. Unlike health insurance, there’s no federal protection against discrimination for these other three types of insurance.

What are some of the reasons to consider getting tested?

Having a genetic test for DM provides information that, should the test result be positive, can assist the attending physician in anticipating complications, and can also allow for easier insurance approval for other diagnostic testing such as an EKG to detect arrhythmia, a common issue for those with DM. While clinicians typically use the repeat number to help inform diagnosis and care considerations, I have found that people in the community like to use this number as a way to help tell their personal stories and create connections to others living with DM.

Is the DM genetic test widely available? Where can people find a lab if they want to be tested?

There are a number of labs that conduct testing for DM, including academic or institutional settings such as Baylor College of Medicine, etc., and for-profit corporations such as Athena Diagnostics and Medical Neurogenetics. There are a number of labs in the U.S. that offer testing for DM. Prices can vary considerably depending on the lab and whether someone is being tested for DM1 or DM2. I would advise people to do their research in advance, and call their insurance plan before testing to see if the insurance company will cover a particular lab.

Insurance plans may request a CPT code to consider a request for genetic test reimbursement. CPT stands for Current Procedural Terminology and is generated by the American Medical Association for coding, billing and insurance purposes. CPT codes can be found on each lab’s website and are publically available.

The insurance company may also want an ICD-9 code, which is a diagnosis code used by the US Centers for Disease Control (CDC) and other international public health agencies to assist with disease reporting. ICD stands for International Classification of Diseases. The current diagnostic code for myotonic dystrophy is 359.21. Updated ICD-10 codes will be implemented in October 2014 and at that time the DM code will change to G71.1.

Insurance companies should be able to explain what percentage of the cost of a DM genetic test they will cover. Individuals with very low incomes can sometimes qualify for financial assistance programs.

What does the testing involve?

It requires a blood draw. It takes about three weeks to get the results, although timing varies from lab to lab. For those with DM in their family who choose not to get the genetic test, I recommend talking with their doctor about scheduling an EKG every year or two, at least until they’re 18 years old. While an abnormal EKG may not imply a DM diagnosis, when coupled with a family history of DM, an abnormal EKG should direct the physician to conduct additional follow up testing.

What do you discuss with individuals who have a family history of DM and are considering getting pregnant?

Reproductive options are definitely an issue we discuss when appropriate. The risk of having a child born with DM is 50/50 for each pregnancy. In this situation, prospective parents can think about options for testing the pregnancy. Tests can be run to look for the repeat expansion at 10-12 weeks of pregnancy through chorionic villus sampling (CVS) or by amniocentesis after 15 weeks.

It’s important to note that the expansions can grow and change as the baby develops. The number of repeat expansions found by CVS or amnio are not necessarily going to be the same number that the baby has when it is born, so those numbers can be used to determine whether the baby will be affected by DM, but not to predict disease severity. The implication for this type of testing would be that the parents can decide if they want to proceed with the pregnancy.

For those who don’t want to go that route, there’s also the option of pursuing in vitro fertilization (IVF) and getting the appropriate testing done before implantation to ensure the DM genetic mutation has not been passed along to the baby. This pre-implantation genetic diagnosis (PGD) test is done after the sperm and egg are fertilized and have developed up to eight cells. At that point, one or two of those cells are removed and tested for the DM repeat expansion. Doctors can then transfer to the mother only embryos that don’t have the expansion, so the disease won’t be passed on.

Generally, it can be assumed that about 50% of these fertilized eggs have the expansion. IVF/PGD is a very expensive option, usually costing at least $20,000 per cycle so, depending on state and insurance plan, prospective parents may or may not be able to get some help with these costs.

Note: Athena Diagnostics offers qualifying patients a substantial discount on its regular price for clinical laboratory services. If you cannot afford to pay for testing ordered from Athena, you may obtain a 75% discount off the list price if you meet certain income guidelines. For more information, see call (800) 394-4493 and ask to speak with a Reimbursement Services Representative.

03/18/2014

Researchers at important academic labs around the US have recently published exciting new information about advances in DM research. The Thomas Cooper Lab at Baylor College of Medicine, Houston, TX released the results of a study that provided important new information on the specific changes that occur in the heart cells of people with DM.

The Mef2 Transcription Network is Disrupted in Myotonic Dystrophy Heart Tissue, Dramatically Altering miRNA and mRNA Expression
Kolsotra et al (Dr. Thomas Cooper’s lab)

A team of researchers at Baylor College of Medicine, under the supervision of Dr. Thomas Cooper, recently published a study examining the changes that occur in the heart cells of people with myotonic dystrophy (DM). Cardiac complications are common in DM, such as abnormal heart rhythms (arrhythmia) and problems with the electrical impulses in the heart that drive it to pump properly (cardiac conduction). The team was led by Dr. Aiunash Kalsotra, the recipient of a 2009 MDF postdoctoral fellowship award.

Given that heart problems are the second most common cause of death in DM, these researchers took a close look at the molecular changes that occur in DM heart cells in order to understand where things go off track. They show that a gene called MEF2 is reduced in DM, causing many small RNA molecules called microRNAs to be reduced. This collection of reduced microRNAs then causes many networks of other genes to be turned off or on inappropriately, and may be one of the reasons why we see cardiac issues in DM. Fortunately, they were able to show that by adding back MEF2 to DM1 cells cultivated in a dish, they could reverse the improper reduction of microRNAs. This study gives researchers a better idea of how the DNA repeat mutations associated with DM may cause symptoms in the heart.

For more information:

Click here to view a pdf of the full article

Click here to read the abstract

01/22/2014

Researchers from the University of Rochester recently summarized the “diagnostic odyssey” experienced by a group of 814 individuals with myotonic dystrophy enrolled in their national registry.  They focused on individuals with a confirmed diagnosis and with first symptoms starting after more than 4 weeks of age, and collected the age when the first symptom was observed, the type of symptom first experienced, any misdiagnoses, the age when a correct diagnosis was made, and the diagnostic tests administered.

This study found that members with myotonic dystrophy type 1 (DM1) experienced an average of 7 years delay to diagnosis, and members with myotonic dystrophy type 2 (DM2) had an even more stunning delay of 14 years to get a correct diagnosis.  On average, DM1 individuals experienced their first symptoms at age 26, whereas DM2 individuals had a later average age of onset at 34 years old.  In general, one quarter of the study’s myotonic dystrophy population experienced their first symptom before the age of 18.

Researchers determined that the type of symptom that manifests first can help dictate how quickly it will take for a correct diagnosis for DM1.  In DM1, members who reported weakness as their first symptom waited on average 6.6 years for their diagnosis, whereas a significantly longer delay in diagnosis was found when the first symptom was myotonia (7.6 years), fatigue (11.5 years), and sleep disturbance (15.6 years).  In DM2, a proper diagnosis was delayed on average 7 years, and was not significantly changed based on the type of symptom first observed.  The most common first symptom in DM1 was grip myotonia, followed by arm weakness, general weakness and leg weakness.  The most common first symptom in DM2 was leg weakness, followed by grip myotonia, general weakness and arm weakness.

Given that members with DM2 waited much longer for a correct diagnosis, the researchers further examined what caused the delay.  One quarter of DM2 members had an incorrect diagnosis, most often originating from a neurologist.  The most common misdiagnosis was limb-girdle muscular dystrophy, but others included chronic fatigue, fibromyalgia, arthritis, and multiple sclerosis.  DM2 members also underwent significantly more testing than DM1 members, having more EMGs, muscle biopsies and genetic testing.  Overall, 71% of DM2 members in this study had a confirmed genetic diagnosis, compared to 58% of DM1 members.

The authors stressed the need for a timely diagnosis to facilitate addressing the short term medical needs of people with DM, because many of the symptoms are disabling and reduce quality of life.  They also note that it is “imperative to diagnose patients earlier in the disease course if promising experimental therapies can reverse or delay onset of symptoms and potentially ward off the progression of many disabling manifestations”.  Now that genetic diagnoses have been available for DM1 and DM2 since 1992 and 2001, respectively, and much effort is being put into educating the community and medical trainees about DM, hopefully the diagnostic odyssey for people with DM will be improved.

An abstract of the original article can be found here.

08/22/2013

Nicholas Johnson, MD, and researchers at the University of Rochester recently published an article in The Journal of Child Neurology that describes the impact of childhood and congenital myotonic dystrophy on quality of life. The authors interviewed 21 children with childhood and congenital myotonic dystrophy and 13 parents. After recording these interviews, the authors reviewed transcripts to identify the most important symptoms to parents and children. Overall, participants reported 189 different symptoms. 

Dr. Johnson and his colleagues found that many of the parents and children identified trouble speaking (dysarthria) as the primary symptom that impacted the children’s lives. Other participants identified learning difficulties and problems concentrating as life altering symptoms. Although many of the study participants did not identify a diagnosis of autism specifically, autistic traits, such as a narrow scope of interest, repetitive speech, inappropriate social responses, and inflexibility were reported. 

As expected, many of the symptoms affecting those with congenital and childhood myotonic dystrophy are different from the symptoms of adult-onset myotonic dystrophy type-1. Prior work by Chad Heatwole, MD, MS-CI, the senior author on this study, identified fatigue, hand and finger weakness, and difficulty walking as significantly impacting the quality of life of those with adult-onset myotonic dystrophy type-1. While these symptoms were also reported in children with myotonic dystrophy, their presentation and significance were different. 

Importantly, many symptoms of congenital and childhood myotonic dystrophy, such as communication difficulties, already have available treatments. The authors hope that by identifying the wide range of symptoms affecting children with myotonic dystrophy, doctors will be able to identify critical symptoms earlier and initiate timely treatment strategies.

Dr. Johnson and University of Rochester researchers, with the support of MDF, have used information from this study to develop a survey, which was distributed to US, Canadian and Swedish patients with congenital and childhood myotonic dystrophy. Results from this international group of patients will be used to further define and prioritize the most important symptoms to patients with congenital and childhood myotonic dystrophy. Ultimately this data will be used to help guide researchers in designing future therapeutic trials for these populations. 

06/25/2013

Maurice Swanson, Ph.D., Professor of Molecular Genetics and Microbiology at University of Florida, Gainesville, and a team of researchers have found that the muscleblind-like 2 (MBNL2) protein in the central nervous system (CNS) may be responsible for the neurological impacts of myotonic dystrophy (DM), providing hope for new treatments. Muscleblind is a type of protein that plays an important role in switching proteins typically found only in babies to proteins found in adults. If this switch isn’t made, an imbalance exists that leads to myotonic dystrophy.

Dr. Swanson states that the team’s work seeks to understand what causes myotonic dystrophy beyond the mutations in the DM1 and DM2 genes.

Dr. Swanson examined which genes were affected by loss of MBNL2 in the brain and found more than 800 affected genes. Many of them had one thing in common: the encoded protein could be made in both fetal and adult forms and MBNL2 appeared to regulate which version was created, according to an article in Neurology Today. One persistent concern that people living with DM1 and DM2 have is the effects of this disease on the brain. “People who don’t have DM usually feel refreshed after a night’s sleep. Myotonic dystrophy patients do not routinely achieve a normal sleep pattern; instead, they have an interrupted series of sleep-wake patterns that do not allow for deep, restful sleep cycles”.

Dr. Swanson created a mouse that lacks the MBNL2 protein as an animal model for DM effects on the CNS. These mice showed normal skeletal muscle structure and function. However, the mice did have DM-related sleep issues, such as a higher number of REM sleep episodes and more REM sleep in general, leading to less restful sleep. In mice lacking MBNL1, another member of the MBNL protein family, the skeletal muscle effects were similar to what is seen in DM. But the central nervous system was not affected, according to Dr. Swanson. 

“What we would like to do now is identify the specific cellular events that are abnormal in the DM brain and see if there is something we can do to treat these disease manifestations with focused therapy development. We would also like to understand the heart and muscle problems in DM. We have developed mice with DM-associated problems and we want to use these mouse models to develop effective drug treatments. Also, we want to understand what is so different about the congenital form of DM. Why does it manifest in babies and children? If we can develop animal models for congenital DM, then we can begin to address the important question of what goes wrong during fetal life,” explains Dr. Swanson.

Recently, therapy development for DM has accelerated and treatments based on anti-sense oligonucleotides will hopefully enter clinical trials in the near future. These new studies focused on the roles of MBNL proteins in CNS function should lead to alternative therapeutic strategies designed to reverse effects caused by expression of the mutant DM1 and DM2 genes.

04/16/2013

Circular RNA Primer

Cells contain a striking diversity of RNA types, many of which have been implicated in the pathogenesis of neuromuscular disease. Unlike most RNAs, circular RNAs (circRNAs) are single-stranded, covalently closed loops. CircRNAs arise via one of three mechanisms: (a) direct ligation of 5′ and 3′ ends of linear RNAs, (b) as intermediates in RNA processing reactions, or (c) via “back splicing,” when a downstream 5′ splice site (donor) is joined to an upstream 3′ splice site (acceptor). A variety of biologic roles for circRNAs have been identified.

Presence of circRNA in DM1 Skeletal Muscle

While aberrant RNA splicing represents a central disease mechanism in DM1, virtually nothing is known regarding the potential for dysregulation of circRNAs. Dr. Fabio Martelli (IRCCS Policlinico San Donato) and colleagues have recently published an analysis of dysregulation of circRNAs in DM1 patient skeletal muscle biopsies (Voellenkle et al., 2019). The authors show that specific myogenesis-associated circRNAs are altered in DM1 biopsies and in DM1 patient myogenic cell cultures.

The research team identified specific circRNAs through review of 30 published DM1 RNAseq databases—relative abundance of a circRNA to its linear counterpart was used as an initial filter, followed by comparison to a list of circRNAs that were previously identified in human or murine myoblasts. Thus, the analysis was not comprehensive, but geared toward identification of transcripts most likely to be dysregulated in skeletal muscle tissue. Candidate circRNAs meeting the investigators’ criteria then were validated using qPCR of skeletal muscles from DM1 subjects and age-/sex-matched controls. Primer specificity was confirmed and the possibility that results were due to a general increase in transcription in DM1 was excluded, and results were confirmed in independent muscle biopsies.

Taken together, four circRNAs—circCDYL, circHIPK3, circRTN4_03, and circZNF609—exhibited significantly increased circular-to-linear RNA ratio in DM1 muscles versus controls. The research team subsequently used receiver operating characteristic curve analysis and confirmed that a transcript’s circular-to-linear ratio could discriminate between DM1 and healthy controls. Finally, the increase in circular fraction for the four circRNAs correlated with a variety of clinical and molecular characteristics of study subjects. Circular fraction ratios correlated with both skeletal muscle strength and splicing biomarkers of disease severity.  Moreover, circular fraction was higher in the more severely affected DM1 patients. Induction of two of the dysregulated circRNAs (circCDYL and circRTN4) was also detected in plasma. Finally, analyses of DM1 myogenic cell lines identified a pattern of circRNA dysregulation that was, in part, similar to data obtained in patient muscle biopsies.

Potential Utility of Dysregulated circRNAs in DM1

The research team self-identified caveats and described these findings as pilot data. Any putative contributing role that dysregulated circRNAs may have in the pathogenesis of DM1 is currently unknown. Yet the discovery of specific, dysregulated circRNAs in DM1 skeletal muscle, if confirmed, may offer advantages for drug development efforts—circRNAs are exceptionally stable in that they are resistant to exonuclease degradation and their dysregulation was detectable in plasma and myogenic cell lines from DM1 patients. These traits make them amenable to use as pharmacodynamic biomarkers for clinical studies and trials in DM1.

Reference:

Dysregulation of Circular RNAs in Myotonic Dystrophy Type 1.
Voellenkle C, Perfetti A, Carrara M, Fuschi P, Renna LV, Longo M, Sain SB, Cardani R, Valaperta R, Silvestri G, Legnini I, Bozzoni I, Furling D, Gaetano C, Falcone G, Meola G, Martelli F.
Int J Mol Sci. 2019 Apr 19;20(8). pii: E1938. doi: 10.3390/ijms20081938.

The Elusive Quantification of Repeat Length

In a disease that exhibits somatic mosaicism, somatic cell instability, and the consequent tissue-to-tissue variability in expanded repeat length, assigning an “actual progenitor repeat length” value to individual DM1 patients has been problematic. The connotations here are obvious—how do we use repeat length for essential drug development functions from molecular biomarkers to genotype-phenotype analyses to stratification of patients in interventional clinical trials, if the parameter is hard to pin down? A recent paper from a multi-site team (Universidad de Costa Rica, University of Texas MD Anderson Cancer Center, and University of Glasgow) attempts to determine the optimal body fluid/tissue to sample and thereby yield insight into the best path forward for clinical studies and interventional clinical trials (Corrales et al., 2019).

Saliva as an Accessible and Reliable Source for DM1 Mutation Testing

Dr. Fernando Morales and colleagues sought to build on their prior findings (Morales et al., 2012 & 2016) that used small pool-PCR (SP-PCR) to control for somatic instability in estimations of progenitor allele length measured in blood. The goal was to improve upon allele length correlations with age of DM1 onset. In their latest work, the research team reports out on comparison of saliva vs. blood as the analyte source for progenitor allele length determinations.

This report was based upon analysis of progenitor allele length in saliva and blood from 40 DM1 patients that had been characterized for age of onset; screening also assessed for presence of variant repeats and methylation of CTCF binding sites adjacent to the DMPK locus, as these may be modifiers of somatic instability. Modal allele length was slightly larger in saliva (529 repeats) than concurrently collected blood samples (486 repeats). Progenitor allele length then was estimated as the lower boundary of allele distribution from SP-PCR—values from the two sample sources were highly correlated and, again, were higher in saliva than blood (414 vs. 310).

Analyses showed that progenitor allele length estimated from blood samples explained 75% of the variation in DM1 age of onset, while that from saliva explained 66% of the variation. The authors suggest that the “true progenitor allele length” needed for genotype-phenotype studies and other preclinical and clinical development purposes is more likely reflected by the values obtained from blood samples.

Additional single molecule SP-PCR studies, excluding two CDM cases, revealed greater somatic instability in blood than in saliva. The research team also showed that the lower boundary of allele distribution was slightly higher in saliva than in blood, while the overall degree of somatic variation was typically lower in saliva than in blood. Finally, analyses of repeat variants and methylation levels as putative modifiers of somatic instability showed that neither were significant factors.

Blood or Saliva?

The authors of this paper summarize the compelling literature case against use of skeletal muscle samples (essentially the confounding effect of tissue-specific rate of somatic expansion) to estimate progenitor allele size, bringing the choice down to blood or saliva. These data show that somatic mosaicism is comparable in blood and saliva DNA from DM1 patients, while saliva is obtained by considerably less invasive means—a feature that is potentially vital for interventional clinical trials in CDM or repeated sampling to assess efficacy of a candidate therapeutic in either CDM or DM1.

References:

Analysis of mutational dynamics at the DMPK (CTG)n locus identifies saliva as a suitable DNA sample source for genetic analysis in myotonic dystrophy type 1.
Corrales E, Vásquez M, Zhang B, Santamaría-Ulloa C, Cuenca P, Krahe R, Monckton DG, Morales F.
PLoS One. 2019 May 2;14(5):e0216407. doi: 10.1371/journal.pone.0216407. eCollection 2019.

Somatic instability of the expanded CTG triplet repeat in myotonic dystrophy type 1 is a heritable quantitative trait and modifier of disease severity.
Morales F, Couto JM, Higham CF, Hogg G, Cuenca P, Braida C, Wilson RH, Adam B, del Valle G, Brian R, Sittenfeld M, Ashizawa T, Wilcox A, Wilcox DE, Monckton DG.
Hum Mol Genet. 2012 Aug 15;21(16):3558-67. doi: 10.1093/hmg/dds185. Epub 2012 May 16.

A polymorphism in the MSH3 mismatch repair gene is associated with the levels of somatic instability of the expanded CTG repeat in the blood DNA of myotonic dystrophy type 1 patients.
Morales F, Vásquez M, Santamaría C, Cuenca P, Corrales E, Monckton DG.
DNA Repair (Amst). 2016 Apr;40:57-66. doi: 10.1016/j.dnarep.2016.01.001. Epub 2016 Mar 8.

RNA: A Popular Target in DM1

Given the novel disease mechanisms that are operative for myotonic dystrophy (DM), “drugging the RNA World” is a very popular theme in drug discovery and development for this disease. Small and large molecule approaches based on this strategy are in preclinical development for DM. Companies such as Expansion Therapeutics are moving toward interventional clinical trials to test the strategy. Dr. Steven Zimmerman (University of Illinois at Urbana–Champaign) and colleagues have recently published data in a model organism in support of a multivalent candidate therapeutic targeting both expanded repeat DNA and RNA in addressing the RNA gain-of-function in DM1 (Lee et al., 2019).

Multivalent Targeting of DM1 Expanded Repeats in DNA and RNA

Dr. Zimmerman and his colleagues have exploited their expertise in bioorganic, synthetic organic, and physical organic chemistry to design and evaluate multivalent ligands targeted to expanded CTG DNA and expanded CUG RNA in a preclinical discovery and development effort for DM1 therapeutics. The Zimmerman team describes the approach as exploiting “smart molecules designed to enter the cell nucleus, bind the target DNA or RNA specifically and operate to reverse the deleterious effects of the expanded repeats.” The efficacy of this strategy lies in the use of a single ligand that targets both transcription of expanded CTG repeat DNA and interactions of expanded CUG repeat RNA with Muscleblind to mitigate the consequences of DM1.

The research team first focused on rational optimization of their existing small molecule inhibitor that targets DM1 expanded repeat RNA. Dimerization of the inhibitor markedly improved its efficacy, but vastly reduced cell permeability—thus necessitating a strategy change to ensure drug-like properties of the candidate therapeutic. Utilization of a linker sequence improved cell permeability and allowed oligomerization of the original monomer to further enhance binding to CUG expansions. Moreover, optimization of their synthetic and purification procedures provided a means to move forward with further testing of the molecule.

Studies in HeLa cells transfected with 960 CTG-repeat DMPK showed the ability of the multivalent compound to disrupt the nuclear foci and rescue the mis-splicing characteristic of DM1 in a dose-dependent manner. Activity in suppressing nuclear foci in the HeLa cell model was 1000-fold greater than with the original monomeric compound. The oligomer also showed promising activity in DM1 patient-derived myoblasts.

A surprising finding with the oligomeric compound tested here was that it also targeted expanded CTG DNA—this dual mechanism capability in targeting both transcription of the expanded repeat DMPK gene and its toxic transcript could potentiate the activity of the compound. Finally, in initial model organism efficacy testing, the oligomer showed activity in a DM1 Drosophila climbing assay and in a liver-specific DM1 mouse model—supporting both the bioavailability and activity of the compound. Preliminary toxicity studies showed no liabilities.

Synthesis and Next Steps

Taken together, a multivalent, cell-penetrating compound with potential to knock down both production of toxic DMPK RNA transcript and its interactions with MBNL represents an attractive opportunity for therapy development in DM1. The model organisms that this compound has been tested in thus far, a DM1 fly and liver-specific DM1 mouse, are not ‘traditional’ models for therapy development in the disease, but establish an initial level of proof of concept. The strength of the Zimmerman group lies in bioorganic chemistry expertise and partnership with groups with strengths in DM1 disease mechanisms and preclinical models should help ascertain the value in moving forward with the novel compounds developed here.

Reference:

Intrinsically cell-penetrating multivalent and multitargeting ligands for myotonic dystrophy type 1.
Lee J, Bai Y, Chembazhi UV, Peng S, Yum K, Luu LM, Hagler LD, Serrano JF, Chan HYE, Kalsotra A, Zimmerman SC.
Proc Natl Acad Sci U S A. 2019 Apr 11. pii: 201820827. doi: 10.1073/pnas.1820827116. [Epub ahead of print]