Due in part to information featured in previous blog posts, I’ve received several e-mails recently from individuals interested in learning more about genome-guided therapeutics for NF. The UAB NF Program is actively engaged in research initiatives in genomic-guided therapy with a focus on identifying approaches that will allow function to be restored to a non-functional gene or gene product. This therapeutic approach would represent an individualized treatment that is tailored to the specific genetic variant responsible for causing NF in an individual.  In this month’s blog, I’d like to discuss the subsets of the most common NF1 mutations and the genomic therapies currently being developed with the goal of restoring at least partial function to the NF1 gene.  

Neurofibromatosis type 1 is caused by a change in the genetic sequence in the NF1 gene, a large and complicated gene that contains a code for making a protein called neurofibromin.  All individuals have two copies of this gene, one inherited from each parent. In people with NF1, one copy of the NF1 gene is altered due to either inheriting the altered gene from a parent, or acquiring a new genetic mutation that occurs in the egg or sperm prior to conception, or from a mutation that occurs early in embryonic development (this results in segmental NF). For someone to develop NF1, a random genetic mutation must occur to the second copy of the NF1 gene in the tissue that will become the neurofibroma, café-au-lait spot, or other lesion. This is referred to as the “second hit” mutation. All individuals – with or without NF1 -- probably have some acquired mutations, which are random errors, that result in a few cells containing an NF1 gene alteration. These cells will not become neurofibromas, however, if only one NF1 gene copy is altered.  The problem for individuals with NF1, however, is that this “backup copy” of the NF1 gene is already altered, which is why a neurofibroma will develop.   Genes function in the cell to direct the production of proteins.  The key question is whether we can find a way to restore function to an NF1 gene that has been damaged by mutation, or perhaps restore function to the abnormal neurofibromin that in some cases is produced.

Therapies Focused on Blocking the Ras/MAPK Signaling Pathway

The majority of therapeutics developed so far for NF1 has focused on blocking the Ras/MAPK signaling pathway that is hyperactive in cells in which both copies of the NF1 gene have been impaired.  Neurofibromin regulates the activity of the Ras/MAPK cellular signaling pathway that helps to control cell growth and division. This pathway is also implicated in other diseases, such as cancer.  Several drugs have been developed that have shown promise in inhibiting components of the Ras/MAPK signaling pathway implicated in NF1 and other diseases. For example, selumetinib is one of a family drugs that has been developed as an inhibitor of one of the components of the pathway and has been shown to have efficacy in reducing the size of plexiform neurofibromas. The development of therapies that inhibit the over-activated Ras/MAPK pathway and other Ras-connected pathways opens new opportunities for treatment for NF1, cancer, and other disorders that share a similar mechanism.

Development of Genome-Guided Therapies Based on Genetic Mutations

While the development of therapies that target Ras signaling is an important approach to developing potential treatments for NF, the possibility of restoring function to mutated genes using genome-guided therapies has gained increasing attention from the NF scientific community and represents an area of focus for the UAB NF Research Program.  An advantage of this approach is that restoring function to the mutated gene might result in fewer side effects than with drug treatments that block Ras signaling.  On the other hand, Ras signaling seems integral to the mechanism of disease in all patients with NF1, whereas genome-guided treatments are based on the specific type of genetic mutation causing an individual’s NF1, and therefore one treatment will not work for all patients. There are thousands of different mutations in different patients with NF1. These mutations are distributed across the gene with no specific mutation predominating. There are, however, subsets of mutations that can be identified through genetic testing, which enable the development of specific approaches to restore function to specific types of mutated genes. In this way, rather than require development of thousands of drugs, one for each mutation, it may be possible to develop a handful, each of which targets a specific type of mutation.

Mutation Subsets

The thousands of mutations can be classified into a number of types.  A deletion mutation results in the total loss, or deletion, of the entire gene and usually produces a severe form of NF. Approximately 3% - 5% of NF mutations are of this type. There are currently no effective methods for replacing large genes, such as the NF1 gene, although this capability may be developed at some future point.

Another type of mutation, called a truncating mutation, causes a blockage or interruption in the formation of a protein.  Neurofibromin is comprised of a chain of 3,818 amino acids strung together in a unique sequence.  One type of truncating mutation, called a premature stop mutation (or nonsense mutation), inserts a signal that tells the protein production machinery in the cell to cease production of neurofibromin before the complete protein is made. Drug therapies currently in development have shown potential effectiveness in overcoming the effects of premature stop mutations. The UAB NF Research Program is currently testing drug compounds that read through the premature stop signals caused by these mutations, with the goal of allowing cells to produce a full-length, functional protein.  

A frameshift mutation is caused by insertions or deletions of a number of nucleotides in a DNA sequence that is not divisible by three.  When DNA is used by the cell to produce protein, the genetic information is read out in groups of three DNA elements, called “bases.”  Hence a specific building block of a protein (an amino acid) is inserted into the protein because of the presence of a specific three-letter base sequence in the gene.  If there is a loss or gain of one or two bases in the DNA sequence for that gene, the reading of the three-letter “words” is confused.  This results in the sequence of amino acids being significantly altered, and at some point there will be a premature stop in the sequence. These types of mutations may be hard to correct, but we are exploring an approach that would jump over the segment of a gene that contains a frameshift when the gene is being processed for reading the sequence and producing the protein. 

Splice-site mutations also result in a meaningless sequence that causes the production of a nonfunctional protein. A gene is encoded in segments, called exons, which code for the amino acids of a protein, separated by introns, which are intervening sequences.  The genetic code in the DNA of a gene is first copied into a molecule called RNA, which is then read out to instruct the production of a protein.  Initially, both the exons and introns are copied into the RNA, but then the introns are cut out and the exons spliced together to make the final “messenger RNA.”  The process of splicing is precisely controlled by the base sequence of the gene, and some mutations occur at the sites that control this process, and therefore disrupt splicing. It may be possible to restore the normal splicing pattern using medications that interact with the splicing system.  This may restore function to a gene disrupted by a splicing mutation; it is also the same method that might be used to jump over a segment with a frameshift mentioned above – this is called “exon skipping.”

Lastly, missense mutations result in the production of a full protein, although one amino acid in the sequence is incorrect. With some sequences, this error won’t cause a problem; however, if the error is related to the production of a critical part of a protein it may disrupt function.  We’re currently working to develop compounds that that interact with protein to restore its function, at least partially. This has been a useful approach to therapeutics in the treatment of conditions such as cystic fibrosis.  The exon skipping approach also might be useful here.

Gene Editing or Replacement

A final possibility is to try to get into the cell and actually correct the gene mutation, or perhaps even to replace the mutated gene entirely.  There has been a lot of interest in these kinds of possibilities, especially recently with the advent of the CRISPR/Cas9 system.  This system was developed based on a natural mechanism discovered in bacteria that protects bacterial cells from infection by viruses.  It has been modified to permit editing of DNA sequences, including potentially correction of gene mutations.  Our lab, and many around the world, are using CRISPR/Cas9 as an approach to creation of model systems that require producing a specific mutation of interest.  Applying this to the treatment of a genetic disorder is much more complicated, especially one like NF1 that affects a very large number of cells in the body.  This is, however, a new area of research, and one where we may see significant progress in the years to come.


We are beginning to see benefits from small molecule treatments that target Ras signaling, but in the long run we are likely to need many parallel approaches to effectively treat NF1.  Our group, and many others, are pursuing such approaches, including the development of genome-guided therapeutics.   It is likely that the eventual treatments of NF1 will require combinations of different approaches that will synergize with one another to control the symptoms of the disorder.