By the GRIN2B Foundation Board of Directors and Dr. Tim Benke, University of Colorado
GRIN2B parents, you’ve got questions; we’ve got answers.
As Admins of the GRIN2B Parent Support Group on Facebook, we pay close attention to the kinds of questions that are regularly asked. Many commonly asked questions are about school supports, sleep issues for our children, and comparing/contrasting of common symptoms. Our worldwide GRIN2B community does a great job of sharing experiences and counseling one another. But often times in our group, questions come up regarding medications and what types of genetic variants the individual children have. We try to chime in when we can and direct people to the appropriate resources, but as we are an all-Volunteer Board that also works full-time, we may miss some of these important questions.
We want parents to be properly informed about topics such as further genetic testing for your child, what medications can be tried and the risks involved AND information regarding (future) clinical trials. We, at GRIN2B Foundation, along with our partner GRIN-disorder organizations are working hard to make clinical trials for GRIN patients a reality in the not-so-distant future. But we are not there just yet, and parents need to make sure they have facts from medical professionals.
We compiled some commonly asked medical questions from our Facebook group and sent them off to Dr. Tim Benke, from our Medical Advisory Board. Dr. Benke is the Director of the Research-Neurosciences Institute at the University of Colorado as well as a Pediatric Neurologist at Children’s Hospital of Colorado. Dr. Benke is also the Principal Investigator of the GRIN Variant Patient Registry.
*Disclaimer – Any discussions that patients and families choose to have regarding the below topics should be done with a genetic counselor, genetic specialist and/or someone experienced and trained to discuss and explain these results with families and patients.
- How do I read my child’s genetic report?
Dr. Benke: I think all parents should keep a binder with their child’s health information. In that binder, create a section for the genetic report from the lab that performed the test. This report may look like this (I made this one up):
GRIN2B c.1234G>A (p.Gly411Asp) Heterozygous, de novo, pathogenic, see comments
The “comments” section will explain that after genetic analysis, there was a spelling change at the address of 1234 in the GRIN2B gene. This change is called a “mutation” or a “variant”; “variant” is now the preferred term. This type of variant is a missense variant.
There are different types of variants. There are nonsense variants and missense variants. A nonsense variant results in only part (if any) of the protein being made. Most nonsense variants are disease causing. The CFERV website has a nice discussion on this: http://functionalvariants.emory.edu/parents/index.html)
This missense address change resulted in a switch from G to A in the DNA at that address. This caused the protein, at protein address 411, to swap a glycine amino acid (Gly) for an aspartate amino acid (Asp). Simultaneous sequencing of the parents did not show this, meaning it is de novo or new to the child. (A new change like this can happen around the time the egg and sperm came together.) Since the child inherited two copies of the GRIN2B gene (one from each parent), the testing demonstrated that this was in only one of those copies (unable to say which one), so it is a heterozygous change. When the company looked up this change in their database and other databases, they found several reasons to state that it is pathogenic (disease causing). They may go on to say in the fine print, that with time their assessment of pathogenicity may change as databases grow. The report could also say: likely pathogenic, inconclusive, likely benign or benign. (See: https://www.invitae.com/en/variant-classification/).
If a variant was inherited from one of the parents, the variant may not be causing disease, especially if the parent does not have similar challenges as the patient. It is possible that a variant can have “variable penetrance” due to the slightly different background of the parent compared to the child. We do not know in GRIN disorders if “variable penetrance” happens. It does happen in other genetic epilepsies such as Dravet Syndrome. Until there is more research about variable penetrance in GRIN disorders, your clinical specialist may discuss if a GRIN variant is potentially causing disease. This assessment is based on the available evidence that can change with time and more research.
If a variant is causing disease, this means that when this patient uses their genetic instructions (DNA) to make the GRIN2B protein, 50% will be made correctly and 50% will not be made correctly. The 50% not made correctly will not work the same as the other. Both will likely be present at the same time.
- Gain of Function and Loss of Function are terms that keep coming up in our Parent community, what do they mean?
Dr. Benke: If a protein is not working the same, it could be either doing less of what it should (Loss of function) or more of what it should (Gain of function). The genetic report will NOT typically say whether or not it is a Gain or Loss of function.
A less than perfect analogy for understanding Gain/Loss is holiday lights. If you are like me, they come out of the box every year the same way I put them in the previous year—all strung together. They are hung up and you realize some strings are working normally and others are not. In a Gain of function situation, there are normal lights next to Gain of function lights that are too bright or blinking too much. In a Loss of function situation, there are normal lights next to loss of function lights that are blinking less or not turning on at all. In both situations, the homeowners’ association is complaining.
GRINs are critical proteins involved in the creation and maintenance of excitatory synaptic connections in most regions of the brain. They are necessary for neurodevelopment, learning and memory of everything from where we just parked the car, the smell of grandma’s house, how to move, how to see, and how much appendicitis hurt when we were a kid. They are also necessary for rhythmic things we don’t think about like breathing. When they are not working correctly, gain or loss, these key processes are affected.
- How do I determine if my child’s GRIN2B variant is Gain or Loss of function?
Dr. Benke: Determination of Gain OR Loss of function is a completely separate process from genetic testing. It is a research-type test. It is not meant to be used to make clinical decisions due to regulatory purposes. Laboratory tests that are used to make clinical decisions (for example, a blood test that shows you are anemic and need more iron) are governmentally regulated, tested and proven to be important for making clinical decisions (see https://www.cms.gov/Regulations-and-Guidance/Legislation/CLIA).
A computer program can sometimes guess at gain or loss of function. The gold standard is to test function in a special laboratory. In the research test to assess the functional status of a GRIN variant, a team of research scientists will test the variant in a special laboratory. Only a few laboratories in the world are capable of doing this testing. Dr. Steve Traynelis does this in his lab at Emory University in Atlanta, GA (http://functionalvariants.emory.edu/). Copies of human DNA that correspond to GRINs are kept in the lab; these are typically made by special bacteria in sufficient quantities. A specific GRIN variant is then engineered with modern genetic engineering techniques, also using these special bacteria. This engineered GRIN DNA is transferred to an “expression system” to make functional GRIN proteins. One expression system typically used is frog eggs. The function of the expressed GRINs are measured using special electrical equipment that measures the current that flows through the expressed GRIN proteins. GRIN proteins are electrical channels that turn off and on to let current flow (recall the holiday light analogy). This GRIN current is turned on when the expression system is exposed to the natural neurotransmitters that make the channel open (glutamate and glycine). The GRIN current can be reduced by natural inhibitors such as magnesium, zinc and protons. Comparisons of variant GRINs are compared to unchanged GRINs. In these assays, if the GRIN current is bigger than it should be, the variant is classified as a Gain of function; if the GRIN current is smaller than it should be, the variant is classified as a Loss of function. If the assays are mixed (some gain, some loss), then the variant is classified as “likely” gain or loss depending on whether there are more gains or losses. If there is no change, then the variant is reported as “inconclusive”.
- Why is it important information to know if my child has a Gain or Loss of function?
Dr. Benke: It is a partially proven theory whether or not the functional status of a missense GRIN variant is clinically important. Based on our understanding of the importance of GRINS in brain function, there is a vast quantity of evidence that loss of function variants are clinically important. There is growing proof-of-concept literature that suggests this theory is also correct for gain of function GRIN variants. The initial excitement began with the work from Dr Traynelis in a patient with very severe, medically refractory epilepsy with a gain of function change whose epilepsy responded to a GRIN blocking drug (https://www.ncbi.nlm.nih.gov/pubmed/24839611). It would seem to make sense that if you have a gain of function, then blocking GRINs might help. It would seem to make sense that if you have a loss of function, then enhancing GRINs might help. However, consider this scenario: the gain of function change caused the circuits to rewire during early development to try to correct the imbalance. As part of this rewire, the brain compensated with other GRINs that do not have the variant. By blocking GRINs, the circuits are now uncompensated again and this causes a serious side-effect like worsening seizures (or worse). A similar scenario is equally plausible with GRIN loss of function variants.
The only way to properly investigate GRIN drugs safely is part of research in an approved clinical trial. A clinical trial will determine whether a drug and what doses are safe. It will determine how often certain side-effects are observed.
It is unknown whether or not the functional status of a GRIN variant affects symptoms. Do gain of function changes always result in epilepsy or not? If not, why not? We would like to answer this question through the registry. Please enroll in the GRIN registry by emailing Jenifer Sargent at Jenifer.Sargent@childrenscolorado.org.
When you enroll in the registry, your variant information is sent to CFERV for functional testing. When testing is complete, we will notify you and provide you with a report. Please provide this report to your clinical specialist to discuss with you. If they have questions, they can contact us to discuss (tim.benke@cuanschutz.edu). Some variants have already been analyzed and the functional reports are available on-line (http://functionalvariants.emory.edu/database/index.html).
- What does it mean if the functional analysis findings are inconclusive?
Dr. Benke: “Inconclusive” can mean several things, including the variant does not change function. If this is the case, the variant may actually be benign and not disease causing. This might be very important information to discuss with your clinical specialist, as it may indicate that further genetic testing may be necessary; that is a clinical decision. Importantly, the variant was not found to change function in the testing assays used. With future research and additional testing assays, an impact on function may someday be determined. This is one of the many reasons why additional research is important. Through the registry, if we find that enough patients with an “inconclusive” variant all look very similar, then we can use this to provide evidence that this variant is still likely functionally important and will drive the development of additional testing assays. (Please enroll in the GRIN registry!)
- Once I get the results back from my child’s functional analysis, how do I use the information to help my child?
Dr. Benke: At the moment, the best way to help your child is to make sure that you have enrolled in the GRIN registry (Please email Jenifer.Sargent@childrenscolorado.org). The functional analysis should not be used to make treatment decisions. In exceptional cases of compassionate use for medically refractory epilepsy resulting in ICU or similar hospital-based care, your treating physicians can contact us to discuss (tim.benke@cuanschutz.edu) use of GRIN drugs.
We all hope that clinical trials will start in the very near future. As part of these trials, we think that GRIN functional status will be an important criteria for determining who can participate, as some clinical trials will use GRIN enhancing drugs and other trials will use GRIN blocking drugs.
- What are some drugs that could potentially be repurposed for either a Gain of function or Loss of function result? If I try one of these drugs now, would that preclude me from participating in a clinical trial down the road?
Dr. Benke: Use of a GRIN drug NOW may preclude you from participating in a clinical trial in the future; this is my opinion based on my experiences with clinical trials. These decisions are driven by the researchers, typically in industry or companies, that set up the clinical trial.
FDA approved drugs that are GRIN-blocking include memantine, ketamine and dextromethorphan. They are approved for other medical uses. Use of these drugs for non-approved uses are typically not covered by insurance, including Medicaid. There are no FDA approved GRIN-enhancing drugs. There are some dietary supplements (serine, glycine) that may be GRIN-enhancing, but it is unclear and not studied to know whether or not they are safe, effective or what doses actually cross into the brain to be helpful and safe. The few studies out there are often single or a few patients, unblinded, and not placebo controlled. These are not the standard for making informed treatment decisions or for FDA approval.
If individuals try things outside of a clinical trial, then how will the rest of the community benefit from this knowledge? How will the community be able to evaluate and trust the knowledge available? The current GRIN registry is trying to collect this information from the community, but this is not the data that would be considered by the FDA to approve a drug. This data is only to support efforts to consider and provide data to seek funding for a clinical trial. Please enroll in the registry by emailing Jenifer.Sargent@childrenscolorado.org.
- Why are clinical trials important and what is the general process?
Dr. Benke: We need clinical trials to understand which drugs are safe and effective. They are needed so that your clinical specialist can feel confident that what they are recommending to you will be safe and effective. Your clinical specialist needs to understand the side-effects that are common to a drug in order to let you make an informed choice when you weigh together the risks and benefits of a therapy. We need clinical trials to get FDA approval for drugs so that their cost can be covered by insurance and Medicaid.
A clinical trial is the research process by which subjects/caregivers consent/assent to participate in a defined clinical research study designed to produce information that can be used to understand (study) or help (trial) the population as a whole. This data is peer-reviewed and published to advance the knowledge and effective and safe treatments of the disease. Trials are interventions.
You do not HAVE to participate in a clinical trial unless you WANT to participate: participating involves consent (in some cases using de-identified data you “opt out” versus “opt in”). You can WITHDRAW at any time. A review board must approve the trial before any subjects are contacted or enrolled; international rules of standards and ethics are applied.
There are phases to trials:
Phase 1: Testing healthy volunteers, multiple doses. May be in patients with advanced stages of a disease or diseases with no known treatments.
Phase 2: Testing on patients for efficacy and safety. (Where GRIN drugs will likely start.)
Phase 3: Confirmatory: Testing on patients for efficacy, effectiveness, and safety
FDA typically will approve for use after Phase 3. This process can take years.
Clinical trial definitions:
Placebo controlled: a placebo (“sugar pill”) is used to make sure the effects are real
Randomized: Who gets placebo or drug is a flip of the coin.
Double blind: Subjects and local Study personnel do not know who is getting drug.
Open Label: Everyone is getting drug and there is no blinding
Cross-over: Those getting drugs then get placebo and vice-versa. Usually Randomized and Double blind too.
Extension: Access to drug after trial ends (usually an open label continuation of the trial).
Gold standard that FDA uses to approve drugs: randomized, double-blind, placebo-controlled
The GRI Center of Excellence has partnered with the RARE Bear program, a grassroots community-driven outreach program for kids with rare disease. Community volunteers create one-of-a-kind teddy bears for one-of-a-kind “rare” kids. Each patient who visits the GRI COE will be gifted with their own “Rare” Bear. 
In our Private GRIN2B Parent Support Group on Facebook, a parent recently commented that most GRIN2B Parents have read the blog post I wrote back in 2016 – 
We are working with the team at GRIN Therapeutics to help design this upcoming Clinical Trial and determine the best locations to have trial sites. But to do this, we need to know which patients are classified as Gain of Function. And for that to happen, 
Please, please, pretty please put these at the top of your list (if you haven’t already done them).
Making room for everyone is at the heart of our Foundation. Everyone comes to us at a different place; some have babies who are being diagnosed, and some are finally putting a name to what their adult children have lived with their entire lives. There is no right or wrong way to process the diagnosis or get involved with our community. But, for those who are ready to get involved in whatever way possible, this blog is the resource you are looking for.
The recipient of GRIN2B Foundation’s 2019 research grant is Dr. Caitlin M. Hudac, an Assistant Professor at the University of Alabama for her research project entitled, “Linking Brain and Behavior: A GRIN2B Biomarker.”
During most vacations, our family is always the outlier. When things go wrong or when breakdowns occur, it can feel like we’re alone on an island. But having another GRIN2B partner family and a group of passionate researchers excited to see us calmed my anxiety and helped create a great getaway for everyone in our family. Like dorks, every member of our two families wore our GRIN2B t-shirts. Everywhere we went, people asked us questions about our “bee” shirts.
Sam and his family could not have been more gracious and patient with us. They were excited to speak with us and spend time with our families. Following the tour, they took us to lunch. After we bid them goodbye, the balance of the day was our own. Our family and the Criders went to a children’s indoor playground, had pizza in the hotel and then spent some time with the kids by the pool. Normal family stuff, but for me, this time it was free from the usual anxieties. Our kids behaved wonderfully, but having another family with us that also experiences the highs and lows that come with a GRIN2B diagnosis made all the difference. 
Samuel Kwon, PhD