Looking for opportunities

Six years to learn walking, 9 years before eating by mouth. This picture seems so ordinary, but his parents see more than meets the eye.

Originally posted 10 March 2017
Updated 16 January 2022
Available in Portuguese  https://pmsbrasil.org.br/procurando-oportunidades/

Success is very much about seizing opportunities. With all of David’s early issues caused by his Phelan-McDermid syndrome (PMS), we could not address everything at once. That said, we always looked for opportunities. For example, when he started climbing in the refrigerator we encouraged him to climb (under watchful eyes). See the picture here: Gene deletion versus mutation.

Like raising a child with a serious genetic disorder, science is about hard work and seizing opportunities. The discovery of penicillin is a classic example.  Alexander Fleming made his discovery in a moldy petri dish. The open dish left sitting in the sink was contaminated by a mold that killed bacteria in the dish. The mold in the dish was accidental, but Fleming’s observation was not. He was a scientist looking for ways to kill bacteria. A few years after the initial discovery, penicillin saved its first life: a child. We need to keep our eyes open for opportunities and we need to make the most of these opportunities. So how can we do that with PMS?

In March of 2017 a group of parents took on a challenge. I asked them to identify other children with PMS who were most like their own child. The goal was to find ways to “cluster” the characteristics of children with PMS, as described in my blog: Splitting, Lumping and Clustering. It was a lot of fun and, just as I suspected, there are groups of kids that are very similar to each other. The exercise on Facebook was an example of crowdsourcing. There are people who are experts at crowdsourcing studies. I would recommend someone expand this exercise into a real study. There is a lot to learn. Parents have insights into their children that medical researchers cannot. Categorizing how groups of children are alike and different could accelerate research, should someone want to take the opportunity

This blog is about untapped opportunities to look at categories of PMS (also called 22q13 deletion syndrome). There are special cases we should not overlook.

Matched deletions

I hear people say that no two deletions are exactly alike. That is not true. There are special cases where the deletions are exactly the same: 1) twins (yes, there are twins in our community), 2) unbalanced translocations (my son’s deletion and my niece’s deletion are exactly the same, as are several other children and adults in our extended family), and 3) germline deletions. I do not know any PMS family with multiple children from germline deletions, but I suspect they exist. In addition to exact matches, there are deletions in regions of the chromosome that have few genes, or few genes that affect the central nervous system. In these cases, nearby deletions may be equivalent in terms of genetic loss (see Understanding Deletion Size).

I have heard some physicians and scientists say “no two deletions are alike” even though they should know better. We need to exploit these cases to find out what matched deletions have in common and how they differ from each other. Those observations will hint at which aspects are purely due to the genes deleted and which are due to more complex interactions between genes and the environment. We need a more nuanced understanding of PMS.

Interstitial math

Most known cases of PMS are associated with terminal deletions of chromosome 22. “Interstitial deletions” (I call PMS Type 2) are far less common. What if we take each person with an interstitial deletion and compare them directly with those who have terminal deletions that start at the same spot on the chromosome? In such comparisons, both people would be missing the same interstitial genes, but not the remainder of the genes. What can we learn? It is a kind of A minus B experiment. It might or might not be very informative, but we won’t know until we take the opportunity. The data are already available in the PMS DataHub.

Pure SHANK3 deletions

If one copy of the SHANK3 gene is missing altogether and no other genes are affected, that specific case can examine the impact of loosing just the SHANK3 gene. But this specific circumstance (a pure SHANK3 deletion) is very rare and not systematically studied. I feel someone should take the opportunity to study these cases systematically. So far, there are studies of larger deletions with attempts to compare different deletion sizes. That is a helpful approach, but not quite the same. There are also scientists who argue strongly that pathogenic SHANK3 variants (sometimes called SHANK3 “mutations”) act simply by reducing the amount of SHANK3 protein. The theory predicts that pathogenic SHANK3 variants are the same as losing one copy of the SHANK3 gene, and nothing else. But the theory has not held up to scientific testing.

Various studies show that pathogenic variants of SHANK3 likely disrupt special “isoforms” of SHANK3 during fetal development, or may disrupt the normal operation of SHANK3 protein in the adult. The difference between having insufficient SHANK3 protein and having an errant (interfering) version of the SHANK3 protein, is important. Oversimplifying the effects of pathogenic variants may lead us down the wrong path when seeking ways to treat people with PMS.

There are opportunities to study the difference between deletions and variants. I describe this in my blog Gene deletion versus mutation. We need a study that specifically compares these two groups: people with SHANK3 mutations and people with complete (or nearly complete) SHANK3 deletions that are specific  enough to leave other, nearby genes, alone.  Once again, we parents seed these opportunities with family data in the PMS DataHub.

Where next?

I believe parents can be major contributors just by our ability to see similarities and differences in our children. The scientists and clinicians studying our children have all kinds of ideas, but frankly they can use a little guidance. I am not a fan of drug studies that mix kids with different pathogenic variants and different deletion sizes without first making the comparisons I have listed. There are more opportunities. We need to encourage greater discussion on the potential role of each important gene of PMS (see Which PMS Genes are Most Important).

Not every parent is interested in the detailed science. But, I encourage parents who take an interest to learn about the genes I discuss in my blogs. I also encourage scientists to think more critically about PMS. It is not a disorder of just one gene, and even in the cases of pathogenic variants of SHANK3, PMS may be a far more nuanced disorder than one of haploinsufficiency. We should always be looking for opportunities to address central questions, and we should always be cautious about our assumptions.

arm22q13

Some previous blogs

Splitting, Lumping and Clustering
Defining Phelan McDermid syndrome
Why don’t we have better drugs for 22q13 deletion syndrome?
What do parents want to know?
Is 22q13 deletion syndrome a mitochondrial disorder?
Educating children with 22q13 deletion syndrome
How to fix SHANK3
Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
22q13 deletion syndrome – an introduction

 

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Splitting, Lumping and Clustering

David happily watching his videos in the summer of 2009

If we want to find treatments for Phelan McDermid syndrome (PMS), first we need to figure out what is PMS.  That was spelled out in my blog: Why don’t we have better drugs for 22q13 deletion syndrome? My next blog addressed how to organize all the different genetic deletions and mutations so that we can define PMS (Defining Phelan McDermid syndrome).  Today’s blog addresses ways we can define different types of PMS.  If we don’t define different types, we are wasting our time experimenting with treatments.  For instance, some PMS kids talk fluently, some talk in short sentences, some can only say single words and many, like David, do not talk at all.  These and many other difference warrant different groups of kids when we test treatments.

Just as there is huge variation in abilities and behavioral characteristics, our kids have very diverse genetics.  Recent studies of rodents  show that not all with Shank3 mutations are alike.  In fact, drugs may work very differently on different Shank3 mutations. Anyone who has kept up with my blogs knows that deletions of different genes are likely to have very different effects on our children.  These difference are very important.

Useful drug testing is stuck right now until we develop a way to categorize people with PMS based on both phenotypic characteristics (symptoms and manifestations) and genotypes (deletions versus mutations and which genes are affected).

I have heard scientists who study Shank3 mice talk about “splitting” and “lumping”.  Splitting is breaking groups into subgroups.  Lumping is putting everyone/everything together into a single group.  Lumping has not worked and the growing consensus is that lumping will never work in our population. Splitting based on just one characteristic (e.g., deletion size) probably won’t work, either.  We need a more refined approach.  What we need is “clustering”.  Clustering is what mathematicians and scientists do when categorizing requires using many different characteristics at once.

Here is an example.  Let’s say you want to buy a car.  You might look at various cars and think about both price and gas milage.  You could make a graph something like this:

Similar types of cars have similar prices versus gas milage tradeoffs.  Race cars are more expensive, but get poor gas milage. Clustering is when you identify meaningful subgroups on a graph because the individual points are close together.  Each group is a cluster.  Even if not every car fits neatly into a cluster, you still have an organizational scheme that can be very helpful.

PMS needs meaningful groups.  Clustering can get complicated when there are more and more features that divide up the population.  However, computer programs can take care of the complexities.  What we need first is to identify which characteristics are important for grouping.  As a practical matter, researchers go back and forth. They consider characteristics, run a program that automatically clusters the data based on those characteristics, and then look to see if the clusters make sense.  That is what we need to do.

When we took David to the PMS Foundation Family Conferences in 2008 and 2010, we met a handful of kids that were remarkably like David (see photo of David, above).  What was it about those kids?  As I recall, they walked the same way, loved watching music videos, asked for help the same way, were nonverbal and all have relatively larger deletions.  Are those meaningful characteristics?  Will they help us divide PMS into different groups for meaningful drug studies? We need to find out.

arm22q13

 

Previous blogs

Defining Phelan McDermid syndrome
Why don’t we have better drugs for 22q13 deletion syndrome?
What do parents want to know?
Is 22q13 deletion syndrome a mitochondrial disorder?
Educating children with 22q13 deletion syndrome
How to fix SHANK3
Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
Science Leadership
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

Why don’t we have better drugs for 22q13 deletion syndrome?

David does not talk, although I am certain he would like to.  He has poor hand control.  He can just barely manage a spoon or glass of water with great effort.  Although he walks a lot, he is always at risk of falling.  There are so many things that are difficult for David.  It would be nice if we had a medication to make his life easier.

After years of drug testing on children with 22q13 deletion syndrome we are probably no closer to a treatment now than when it started.  This problem is not unique to 22q13 deletion syndrome; it is true for many, if not most neuropsychiatric disorders (see: Hope for autism treatment dims as more drug trials fail).  Recently, Rachel Zamzow wrote a very readable review about why autism clinical trials have failed (Why don’t we have better drugs for autism?).   Her review is in Spectrum, the on-line magazine affiliated with the Simons Foundation Autism Research Initiative (SFARI). Rachel identifies three problems that plague clinical trials: 1) bad design, 2) wrong measures and 3) too broad a range of participants.  While problems 1 and 2 are important, problem 3 is a major stumbling block for 22q13 deletion syndrome that I would like to address.

Clinical trials for 22q13 deletion syndrome are intended to treat defects or loss of SHANK3 (Kolevzon et al., 2014).  The problem with finding a treatment for SHANK3 is just as Rachel – and many others – have described.  If the subjects you are testing are too diverse, you will never see a clear impact of the drug you are testing.  The subjects recruited for these studies have either SHANK3 mutations or have 22q13 deletion syndrome with terminal deletions of different sizes.  This group is more diverse than many, perhaps all, of the other autism-related clinical studies that have failed.  Going on past experience in the field, this clinical group will not provide useable results. Here are the reasons why.

SHANK3 mutations are complicated

Early on, there was hopeful enthusiasm about hunting for a cure for people with 22q13 deletion syndrome.  At that time, SHANK3 mutations were lumped together with chromosomal deletions.  Importantly, SHANK3 mutations were thought of as simply a loss of SHANK3 function.  As it turns out, SHANK3 mutations are tremendously complicated. Different SHANK3 mutations can have very different effects on the gene, on the proteins it produces, on the neural development of the brain, and on the impact it has on both people and experimental animals.   The most recent and most thorough review of Shank proteins (Monteiro and Feng, 2017) says it clearly: “Indeed, the idea that isoform-specific disruptions [different mutations] will result in different phenotypic consequences (and even result in different disorders) has recently gained momentum.”  I can say with some pride that the momentum includes my June 2016 blog How to fix SHANK3, which makes that very same point.  You cannot lump together people with different SHANK3 mutations and expect to get a single clear result.

Too few patients have the same SHANK3 mutation

To date, no one has been able to find enough people with the same SHANK3 mutation to do a drug study.  You can find SHANK3 mutations in large autism databases, but these are not like a registry where you can call the patient up and ask them to participate.  There is no doubt that medical researchers would pull together a SHANK3 drug study population, if they could.  Autism is thought to be a polygenic disorder (like schizophrenia). Thus, we expect that many individuals from autism databases will also have mutations of multiple autism-related genes, not just SHANK3.  Finding a large enough group of people with one (or two) SHANK3 mutations to study drugs will probably never happen.

Individuals with 22q13 deletions are too diverse

Another approach might be to use 22q13 deletion syndrome patients with terminal deletions that remove SHANK3 altogether.  Every one of these patients would have exactly the same SHANK3 loss.  Further, there is a registry for 22q13 deletion syndrome patients that might help with recruitment (PMSIR).  While this seems appealing, it has its own flaw.  Just as the SHANK3 mutation population is likely to have other autism and intellectual disability genes complicating the picture, chromosome 22 is full of genes that likely contribute to autism, intellectual disability, hypotonia and other phenotypic traits associated with SHANK3.  Anyone who has read my other blogs has seen numerous examples of those genes (see Mouse models and How do we know which genes are important?).  Because of the densely packed genes near SHANK3 (see Understanding deletion size), it is unlikely that a big enough group of people with 22q13 deletion syndrome can be found with deletions that don’t involve other critical genes on 22q13.

Solutions

In her article, Rachel Zamzow discusses the N-of-1 Trials approach. We parents do this all the time.  We experiment with different medicines on our one child. N-of-1 design simply has the clinical researcher follow the child during the test.  I’m not a big fan of N-of-1.  I prefer a mixed experimental approach where research animal testing is done in tandem with human testing (see Have you ever met a child like mine?).

In their detailed review of Shank proteins and autism, Monteiro and Feng recommended that “..careful genotype-phenotype patient stratification is required before individual testing of specific pharmacological agents.”   That is, don’t test drugs until you understand the impact of the genes that have been lost.  If you have been reading my blogs, that should sound very familiar.

Two things must change before we can expect drug testing to bring meaningful results.  First, we need to organize Phelan McDermid syndrome, SHANK3 mutation syndrome(s), and chromosome 22q13 deletion syndrome into a meaningful “genotype-phenotype patient stratification”.  That is, we need to define different types and subtypes of the syndrome that was once called 22q13 deletion syndrome.   I proposed running an interactive session with parents and researchers in 2012, and for the session I put together a Power Point presentation called: “Defining PMS across Genotypes Phenotypes and Molecular Pathology.”  I was asked not to present my ideas.  Perhaps I will be given a chance, someday.

Second,  we must spend the time to characterize the genes that are near SHANK3 on chromosome 22 and understand (in experimental animals) how they might contribute to 22q13 deletion syndrome.  We need to study people with interstitial deletions, so we can isolate the effects of these genes. Efforts to explore the contributions of 22q13 genes has been lacking, yet they are a major impediment to the search for effective drug treatments.

22q13 deletion syndrome has left David completely dependent upon others for his day-to-day living. Both David and I have come to accept that.  What we cannot do for David is know where it hurts when he is sick or injured. If I had one wish for a new medicine, that medication would let David point to where it hurts. That medicine, or any useful medication, is not going to happen until someone takes the needed steps to remove the impediments that interfere with productive drug testing. It is clear where we need to go.  The question becomes, who will take us there?

arm22q13

 

Previous blogs

What do parents want to know?
Is 22q13 deletion syndrome a mitochondrial disorder?
Educating children with 22q13 deletion syndrome
How to fix SHANK3
Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
Science Leadership
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

What do parents want to know?

David dressed up in his birthday best

One nice thing about writing a blog is getting feedback.  While I embrace and benefit from all kinds of feedback, I admit being partial to positive feedback.  I get some nice comments from friend on my Facebook page.  I love the encouraging comments that get posted here.  There is another kind of feedback that is neither positive nor negative, but very informative.  It is the kind of feedback that we should all pay attention to.  Visitors vote with their mouse (or touchscreen).  Every time someone clicks on a blog link, WordPress adds one to a blog page counter.   Now that it is 2017, let’s see what the numbers recorded in 2016 tell us.

Frequency of access for all English Language pages of this blog in 2016

The most requested page is at the bottom of the graph: How do I know which genes are missing?  That is the number one question on parents’ minds.  It makes sense.  If your car breaks down, you want to know what caused it, even if you don’t know much about cars.  At the very least, you have some idea of what needs fixing.

Three more questions are virtually tied for 2nd place. Have you ever met a child like mine? and How to fix SHANK3 are discussions of SHANK3 and its relationship to the other genes of 22q13 deletion syndrome.  Together, with the most requested blog page, over one-third (35%) of mouse clicks on this blog are from people who want to understand how all the genes of 22q13 deletion syndrome operate together to produce the disorder.  The other blog page that is tied for 2nd place addresses the same topic from the opposite direction: How can the same deletion have such different consequences?

Taken together, nearly half (46%) of the information people want from this blog is to understand what genes are missing and why those genes matter.

Most visitors in 2016 already knew about 22q13 deletion syndrome.  Only about 4% of all clicks went to 22q13 deletion syndrome – an introduction. Fewer clicks went to learning about the author. (I can live with that!)  But, I think there are a other links that deserve attention.  Here is a suggestion for the new year:

Gene deletion versus mutation: sometimes missing a gene is better.
SHANK3 is not the only gene of 22q13 that can have serious consequences when mutated (modified, but not lost altogether).  Much is said about SHANK3 mutations, but 98% of people with 22q13 deletion syndrome are missing SHANK3 altogether. Understanding the difference may be crucial to finding cures.

I have dedicated the past few months to formal writing about 22q13 genes aimed at the scientific community.  That work has taken me away from this blog, but, hopefully, taken us all closer to effective treatments for our children.   That work is done for the moment and I hope to get back to this blog on a more regular basis.

arm22q13

 

Previous blogs

Is 22q13 deletion syndrome a mitochondrial disorder?
Educating children with 22q13 deletion syndrome
How to fix SHANK3Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
Science Leadership
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

Is 22q13 deletion syndrome a mitochondrial disorder?

David enjoying a walk in the park

Science is really interesting if you don’t let the details overwhelm you.  Scientists master huge piles of details, but they always step back to see the big picture. They are truly fascinated with science.  That fascination motivates their quest.  In this blog I will point out some really interesting facts so  you can share that fascination.

This blog is about organelles of the cell called mitochondria. If you look at cell with a high power microscope you will see something that appears to be another tiny organism living inside the cell.

Interesting fact #1: Mitochondria may have originally been single cell organisms that invaded larger cells.  Now, mitochondria are simply part of our cells.

Interesting fact #2: Mitochondria are the battery chargers of the cell. They turn sugars and oxygen into ATP.  ATP molecules are the rechargeable batteries used by the cell for nearly everything – from muscle contraction to digestion, growth and thinking.  Imagine what might happen if all your battery chargers were on the fritz. That would be a cell phone (mitochondrial) disorder with dramatic consequences.

Interesting fact #3: A paper published in January of this year shows that most people with 22q13 deletion syndrome have mitochondrial dysfunction (http://www.ncbi.nlm.nih.gov/pubmed/26822410).  Mitochondrial dysfunction affects more kids than any other problems except for intellectual and physical disabilities.

Interesting fact #4: Mitochondria are unique organelles because they operate using two separate sets of genes.  One set of genes is on the regular (nuclear) DNA.  The other genes are actually inside the mitochondria.  These genes, mitochondrial and nuclear, operate together.

Interesting fact #5: mitochondrial genes come only from the mother, whereas nuclear DNA is an even mix of both parents. The term “mitochondrial gene” is confusing.  Sometimes it means a gene from the mitochondrial DNA.  Other times it means a nuclear gene that is needed to help the mitochondria work properly.  In 22q13 deletion syndrome, a group of genes on chromosome 22 (nuclear DNA) are lost.  Many of these genes are important to normal mitochondrial function.  These are the mitochondrial genes I will discuss now.

The following genes impact mitochondria.  I have written on several of these genes previously.  This list includes the minimum size of a terminal deletion that would damage or delete the gene.  The list is borrowed from my earlier blog (How do I know which genes are missing?).

Mitochondria related gene	Deletion size (Kbase)
CHKB	235.47
CPT1B	240.05
TYMP	288.47
SCO2	292.86
SELO	600.85
GRAMD4	4,180.53
TRMU	4,484.77
ATXN10	4,973.16
KIAA0930	5,577.70
SAMM50	6,821.94
TSPO	7,655.23
MCAT	7,675.07
BIK	7,688.76
ATP5L2	8,177.87
NDUFA6	8,727.69
SMDT1	8,735.12
ACO2	9,289.48

This list has 17 genes.  About half the children with terminal deletions are missing 5.3 Mbases (5,300 Kbases) or more (see Understanding deletion size).  That means half or more of our children are missing at least 8 mitochondrial genes.  Have you met a child with a really large deletion?  They generally have multiple major health issues.  It is not clear which mitochondrial genes contribute the most to their problems, but even children with smaller deletions are outside the normal range of mitochondrial enzyme functioning.

Some of these genes likely have little or no impact.  I have written about ATXN10 (Gene deletion versus mutation: sometimes missing a gene is better.). Some people inherit a mutated copy of ATXN10 that has an extra sequence.  It is an unused sequence that gets stripped off when the protein is produced.  The protein is normal.  However, the excess stuff that gets stripped off interferes with another enzyme in the body.  The interference ends up poisoning the mitochondria and killing the cells (http://www.ncbi.nlm.nih.gov/pubmed/20548952).  Fortunately, this is not a gene we need to worry about.  Our children are missing ATXN10.  They don’t have the mutated gene seen in certain families. Likewise, based on what is know about the BIK gene, it is also unlikely to contribute to our children’s mitochondrial problems.

So, which mitochondrial genes are causing so many problems?  One gene is specific for muscle function (e.g., CPT1B). We should not be surprised if that mitochondrial gene contributes to hypotonia (see 22q13 Deletion Syndrome: hypotonia). However, most mitochondrial genes are essential for normal function in every part of the body.

The degree of damage mitochondrial genes can cause can be seen with SCO2.  Loss or damage to both copies is often fatal early in life (http://omim.org/entry/604377?search=sco2&highlight=sco2).  Perhaps more importantly, loss of just one copy of that same gene reduces essential enzyme activity and leads to behavioral defects in mice  (http://www.ncbi.nlm.nih.gov/pubmed/22900024).  Thus, SCO2 could be a major contributor brain dysfunction in many children.

We know that more than half of our kids are outside the normal range of mitochondrial function.  Many scientist believe that mitochondrial genes contribute to neurodevelopmental disorders (http://www.ncbi.nlm.nih.gov/pubmed/26442764), and that those disorders (e.g., 22q13 deletion syndrome) are associated with mitochondrial dysfunction (for example, http://www.ncbi.nlm.nih.gov/pubmed/26439018).   What is needed now is a more complete study that includes all of our children.  How? When the scientific paper came out in January describing mitochondrial dysfunction in our children, I was certain the researchers would be invited to the 2016 conference.  The head of that study was eager to come, present his results and gather more cheek swabs. What happened?

arm22q13

Previous blogs

Educating children with 22q13 deletion syndrome
How to fix SHANK3
Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
Science Leadership
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

 

How to fix SHANK3

David snacking on some cereal bar bits

Human – Mouse Partnership

Originally posted 16 June 2016
Updated 19 August 2022

Anyone who has read the majority of my blog pages knows that my goal is to help parents, scientists and other members of the Phelan-McDermid syndrome (PMS) community understand how the genetic landscape of chromosome 22 must shape our thinking if we are going to realistically pursue treatments. At least one company has proposed to tackle the challenging task of a genetic intervention. While very exciting, we also must be humble about the difficulty of making it work. This blog looks at the limitations of this approach.

If you have not read the earlier blogs, much of this one may seem foreign. This blog is based heavily on prior ones. Because of the overlap, I will omit most scientific references and simply recommend reviewing prior posts for supporting evidence.

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There are a remarkable number of optimistic-sounding mouse model papers on the SHANK3 gene. The number of model mice has passed one dozen. People who work on SHANK3 mice often describe their rodents’ behaviors as mouse analogs to human behaviors. When an unusual mouse behavior is “rescued” with a chemical compound, the implicit (sometimes explicit) suggestion is that mouse research is on a path to curing autism, PMS, or maybe even schizophrenia. Some researchers like to define PMS as a disturbance of SHANK3, which guarantees that any SHANK3 fix will fix PMS. This is not consistent with research that looks at the complete genetic landscape of PMS. Rodent research papers are often rather optimistic. Perhaps writing papers this way promotes optimism in the patient community. Optimistic papers help to keep financial donors excited. These are probably good things, but we need to recognize the limitations of the research.

From a practical standpoint, we need a strategy for fixing SHANK3 problems in humans that accurately reflect the science. We need a plan that has more to do with the human disorder than the rodent one, and more to do with  therapeutic benefit than a detectable statistical change. The plan needs to be based on what we know more than what we speculate. The plan needs to be about the patients, not the scientists, funding agencies or charity organizations.

One critical clinical concern is the relationship between SHANK3 deletions and SHANK3 sequence variants (sometimes referred to as SHANK3 mutations). Nearly all rodent studies are studies of SHANK3 mutations. The reason for so many rodent models is largely because different laboratories study different mutations. This is not such a bad idea because many of these mutations are designed to duplicate human SHANK3 gene variants. The mouse studies are virtually always described as studies of SHANK3 deletion. However, they are rarely complete removal of the SHANK3 gene (or removal of both SHANK3 genes). Among the many different SHANK3 mutations studied in mice, the behavioral, molecular, electrophysiological and drug effects differ widely. Importantly, a study that looked at total removal of SHANK3 protein from a mouse found less impact on the mouse behavior than many of the other SHANK3 mutation studies. The important message here: in rodents some mutations have a greater impact than complete SHANK3 deletion.

What about humans? Are SHANK3 variants different from SHANK3 deletion? That is, do people with SHANK3 variants have the same problem as people with 22q13 terminal deletions (complete deletion of SHANK3). In 2022 there was a paper that hints at the answer. After carefully looking at patients with small terminal deletions (under 0.25 Mb) and comparing them to patients with SHANK3 variants, they observed:

Although individuals with small deletions and SHANK3 variants showed similar findings in most of the categorical variables (Table 5), a remarkable difference was observed in “the ability to make sentences” between the two groups, with 30/65 (46.2%, Supplementary Table S3) of individuals with deletions below 0.25 Mb able to make sentences compared with 5/18 (27.7%, Table 1a) among those with SHANK3 variants.
Nevado et al 2022 Variability in Phelan-McDermid syndrome in a cohort of 210 individuals, Frontiers in Genetics

Significantly more people with complete deletions of SHANK3 were able to speak in sentences compared to people with SHANK3 variants. This and previous studies have shown that different human SHANK3 variants can produce very different impacts. Clearly, some of these variants interfere with the normal operation of SHANK3 in ways that are worse than just reducing the amount of available protein.

This effect is not difficult to explain. Some SHANK3 variants can produce improper SHANK3 proteins that wreck havoc with the assembly of the synapse. As an analogy, think about placing a bunch of defective nuts and bolts into the manufacturing process for a car or airplane. The production line may be better off omitting some hardware (or producing fewer products) than installing defective parts. The somewhat surprising conclusion is that we might want to to treat some SHANK3 variants by shutting down a defective SHANK3 gene.

If we are considering SHANK3 deletion as a treatment for SHANK3 mutation, then we better be prepared to treat SHANK3 deletion. Results from the first Shank3 complete knockout mouse provides a hint at how to treating human SHANK3 deletion. The most abiding and measurable effect of complete Shank3 deletion in the mouse is failure to engage and benefit from an operant conditioning task (lever pressing for a reward). This effect appears to be associated with abnormal ventral striatal function, which is consistent with many previous studies of the ventral striatum. Failure to explore and learn would be indicative of intellectual disability in humans, so it is of great interest to understand the exact relationship between the learning deficits in humans with pure SHANK3 deletions and mice with pure (complete) Shank3 deletions. Such an undertaking would require a very modern and somewhat novel strategy in the world of pre-clinical neuropsychiatric research.

The precise nature of the mouse learning deficit is not yet understood. Learning is a complex process and many aspects are very subtle. Even the reported rescue of learning in the Shank3 knockout mouse creates more questions than answers. These questions go to the heart of how SHANK3 loss might contribute to intellectual disability in humans. How can the details of learning deficits caused by SHANK3 deletion be dissected out? I do not believe it can be done in mice, but it is difficult to find humans missing the entire SHANK3 gene, but little else (pure SHANK3 deletion). Given the rarity of pure SHANK3 deletion, I propose that scientists could do an in depth study of one or two human volunteers with very small deletions. This research could be modeled after behavioral research methods from studies of nonhuman primates. These are studies where behavior from just one or two animals is studied in great detail.

The studies with the PMS volunteers would combine behavioral testing and advanced computational modeling. The results of each new test leads to a modification of the model, and the results of each new model identifies new things to test. These state-of-the-art computationally-based scientific learning studies are designed to incorporate variables that can be directly tied to equations describing an underlying theoretical framework of the learning process. Animal researchers are adept at designing learning tasks in ways that do not require verbal instruction. They are equally practiced at inferring the results without the need for verbal reports. Still, with the participation of a fluent verbal subject, researchers can work with the subject to help design tasks (games) that are interesting and engaging. Why not let the subject have fun?

As these learning tasks  begin to characterize the nature of the deficit seen in the subject/participant, they are then re-designed for testing in animal models. Current rodent models could be used with simple tasks, but more demanding tasks may require nonhuman primates. These studies could include fMRI and electrophysiological investigations. The technology of gene editing, common in mice, has reached farm animals and at least two species of nonhuman primates. As these methods become more mainstream, complete SHANK3 deletion could be a practical research option, especially in old world monkeys, species that shares important common features with human cortical evolution.

The goal of this scientist/participant research partnership is to develop a sensitive cross-species measure of learning ability that parametrizes the impact of SHANK3 dosage. Such a measure provides two invaluable assets to the development of treatments. First, animal models can be validated (or not) based on exquisite computational approaches that may be able to distinguish species differences from the influence of SHANK3 dosage. Second, interventions, either learning-based or pharmaceutical, could be tested using measures sufficiently sensitive to reflect the identified nature of the deficit. What can this human research/animal research partnership hope to produce? The first successes may be refinements to educational methodologies. The learning models could point the way to improvements in teaching strategies for our kids with PMS. Later, dare we hope, new pharmaceutical treatments could emerge.

We all hope that initial human trials of gene replacement therapy will provide a critical new treatment for our kids with PMS. Those trials may take many years to materialize. In the meantime, we need to better understand what we can expect from gene replacement, and the exact nature of learning deficits that arise from loss of SHANK3.

arm22q13

Previous blogs

Have you ever met a child like mine?
How do I know which genes are missing?
Mouse models
Science Leadership
How can the same deletion have such different consequences?
22q13 and the hope of precision medicine
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

22q13 deletion syndrome and science leadership

My previous blog (How can the same deletion have such different consequences?) ended with a discussion of science leadership.  I asked the question, “Why have we not benefited?”  That is, why have 22q13 deletion syndrome children like my son, David, seen little or no benefit after decades of research?  I have heard many reasons: limited number of patients, limited financial resources, limited knowledge, etc.  I frequently hear even less convincing phrases like, the research is “very promising” and “scientists are doing their best”.

However, these are not reasons for lack of results. These are reasons that emphasize the need for qualified, family-centric science leadership.  In the previous blog I identified another chromosome deletion syndrome (18q deletion syndrome) with all the same difficulties, yet their story is much different (See the scientific articles Making chromosome abnormalities treatable conditions and Consequences of chromosome 18q deletions).  Research on 18q deletion syndrome has been enormously successful in plotting a scientific course towards treatments.  Since writing that blog I discovered why.  Rather, who: Jannine Cody.  Dr. Cody got her PhD to develop treatments for her daughter with 18q deletion syndrome.  The success of 18q comes from the science leadership of Dr. Cody.  Dr. Cody is a qualified scientist without any other conflict of interest.  She is committed not only to every family, but also to the impact of each and every gene on the long arm of chromosome 18.  I encourage you to watch the YouTube and hear her story.  Science leadership makes all the difference in the world.

 

arm22q13

Comment la même suppression peut-elle avoir une telle variété de conséquences ?

[English version]

L’auteur, Andy, et sa cousine ont la même chromosome hybride 22 qui provoque le syndrome de la suppression 22q13.

Leurs enfants ont la même suppression mais un des enfants d’Andy est mort et l’autre, David, a failli mourir. La cousine n’a pas eu cette expérience et sa fille, contrairement à David, n’a pas pris 6 ans à apprendre à marcher, 9 ans à manger oralement et elle peut parler avec les phrases courtes. C’est la question la plus posée par les parents d’enfants atteints.

Si SHANK3 n’est pas la cause de la suppression 22q13, pourquoi y a-t-il un enfant avec uniquement la mutation SHANK3 qui ne sait ni parler ni marcher ? Et il y en a d’autres qui savent très bien marcher et parler. Si vous acceptez la dilemme que des suppressions similaires peuvent provoquer des résultats très différents, il n’y a plus l’argument du gène favori (SHANK3 aujourd’hui, demain autre chose probablement.)

Pour expliquer ces différences il y a plusieurs réponses :

1. Perte de l’hétérozygotie : Des petites erreurs génétiques ont lieu tout le temps pendant le développement et dans la vie adulte. Facteurs environnementaux – radiation, infections, coups de soleil, toxines. Le fait d’avoir les paires de gènes de deux parents différents nous protège des erreurs génétiques graves. Lorsqu’il manque des gènes à cause d’un syndrome de suppression, on n’a qu’une copie d’un gène alors il y a l’occasion parfaite pour des erreurs non-corrigées. Si ce gène est endommagé par la suite, les conséquences peuvent être graves. L’erreur peut être globale (corps entiers et détectable avec les tests génétiques) ou local (limité à une partie du corps ou du cerveau.) Quand l’erreur est locale elle n’est pas détectable et devient une différence inexpliquée.

2. Impression (ou empreinte ?) : Quand un des gènes est éteint comme le syndrome d’Angelman. Si la seule copie qui reste du gène s’éteint ça va provoquer des problèmes qui ne seront pas expliquées.

3. Impact de la mutation du gène : Souvent un gène qui est supprimé a moins d’impact qu’un gène qui a subi une mutation. Comme dans le cas des cellules de cancer ; on préfère qu’elles meurent au lieu de pousser avec des gènes modifiés. Si une suppression chromosomique ne frappe qu’une partie du gène, le gène peut commencer à créer des protéines qui empêchent le fonctionnement normal de la cellule et la petite suppression provoque des grands problèmes.

4. Les combinaisons de gènes : Quand il y a plusieurs suppressions et mutations on cumule les erreurs génétiques et chacune fait sa contribution. On a parfois des gènes qui, seuls, n’ont pas d’impact lors de leur suppression/mutation mais avec certains autres peuvent avoir des conséquences beaucoup plus importantes. On ne comprend pas encore ces combinaisons alors on utilise des termes comme ‘antécédents génétiques’ en attendant. La différence génétique principale entre David et la fille de la cousine d’Andy vient de la femme d’Andy et du mari de sa cousine ; des différences dans leurs antécédents génétiques.

5. Mosaïcisme et mutations somatiques : Pendant le développement les erreurs génétiques peuvent avoir lieu dans une petite partie du cerveau. Ces erreurs peuvent expliquer les variations comme les difficultés d’apprentissage. Ces mutations inaperçues peuvent avoir des incidences plus lourdes lorsqu’elles interagissent avec les 30 à 100 gènes manquants. L’impact de SHANK3 peut être amplifié par ces gènes modifiés ou perdus dans des parties spécifiques du cerveau.

6. Les régulateurs génétiques: Le projet génétique ENCODE tente de trouver tous les morceaux régulateurs des gènes de l’ADN. La plupart de l’ADN est composée de régulateurs de gènes. C’est plus facile à comprendre quand vous vous rendez compte que les cellules de foie, de peau, des intestins, du cerveau, ont toutes les mêmes gènes. Ce qui change est les gènes qui sont actifs ou inactifs. Les cellules de peau savent qu’elles sont cellules de peau et n’utilisent que les gènes de cellules de peau. L’ADN est réglé dans chaque tissu à associer la signature génétique nécessaire à fabriquer le tissu. Les suppressions chromosomiques 22 suppriment également les régulateurs de gènes. Les régulateurs sont très difficiles à repérer et pourraient être la cause de ces différences inexpliquées.

Compte tenu de la complexité et de nombreuses possibilités de variation inexpliquée , nous pouvons commencer à apprécier que la connaissance de la taille de suppression d’un individu ne fournit pas toutes les réponses. Cependant, grâce à des outils modernes, il existe des moyens pour étudier les effets de la taille de la suppression même avec une telle variabilité. Ces outils peuvent être utilisés pour démêler les gènes qui contribuent à chaque problème médical. Cela exige un engagement sérieux des parents à pousser les chercheurs et le personnel médical vers ces recherches, tirant pleinement parti des rapports génétiques. Trop l’accent sur un gène préféré entrave le progrès scientifique et médical. Ceux qui travaillent sur un autre chromosome, syndrome de délétion (suppression 18q syndrome) ont étudié leur syndrome à bon escient au cours des 50 dernières années. Ils se tournent vers la médecine scientifique pour les symptomes de 18q (Voir Création d’ anomalies chromosomiques). Les personnes 18q ont développé une feuille de route, que les gens atteints du syndrome de délétion 22q13 peuvent facilement suivre (Voir Conséquences du chromosome 18q suppression). J ai travaillé durement dans mes dernières recherches pour faire ce point , mais rien est plus convaincant que de voir d’autres prendre les devants avec une telle clarté et tel engagement. Pourquoi n en avons-nous pas profité ? La seule explication que je peux trouver est que la communauté du syndrome de délétion 22q13 manque de personnes qualifiées, dirigeant scientifique impartiale. Il y a un problème assez évident, avec des conséquences très tristes. Il n’y a pas plus de traitements pour David aujourd’hui qu’il ya 30 ans. Nous savons quels gènes sont portés disparus et pour beaucoup d’entre eux, nous savons ce qu’ils font (Voir Comment savons-nous quels sont les gènes 22q13 suppression: l’espoir de la médecine de précision). Ce que nous ne semblons pas savoir est comment rendre le travail de la science meilleur pour le bien de nos familles.

 

[My thanks to Betty Sepré for doing this translation.  That said, I take responsibility for any errors in typing, translation or content. Feel free to contact me with corrections. –  arm22q13]

How can the same deletion have such different consequences?

David at 3 days of age: David’s deletion size is exactly the same as his cousin’s, yet the deletion had more severe consequences.

Originally posted 19 January 2016
Updated 19 July 2021
Available in Portuguese  http://pmsbrasil.org.br/como-a-mesma-delecao-pode-ter-consequencias-tao-diferentes/

Introduction

There is a common question among parents of children with 22q13 deletion syndrome. Why is one child with a larger deletion able to talk, while another child with a much smaller deletion nonverbal? Unexplained differences in “phenotype ” (the manifestations of the disorder) can be confusing to parents. Some parents conclude that deletion size is unimportant, but that is incorrect. A sports a team can have good days and bad days. The team is judged on its overall record. Genetics are similar. A deletion in one individual can be more or less impactful, but it is the overall statistics of deletions that demonstrates the clear effect of deletion size.

The person-to-person variation is especially striking when we look at how the same genetic arrangement can have very different outcomes. My cousin and I inherited the same abnormal chromosome. Somewhere back in family history, tiny bits of chromosomes 19 and 22 got swapped (see Who is arm22q13?). My genetics include a hybrid and unhelpful chromosome 22 that has caused 22q13 deletion syndrome in my children (see Understanding translocations in 22q13 deletion syndrome: genetics and evolution). My equally unfortunate cousin had the same chromosome inherited from our grandfather. Despite inheriting the exact same deletion, our children turned out very differently. Both of my kids who received this chromosome were failure-to-thrive babies. One died a few days after birth and the other one almost died (see photo of David, who lived). My cousin had no such experience. Her daughter has 22q13 deletion syndrome, but without perinatal problems. Her daughter’s case of PMS is not as severe as David’s. For example, David does not talk at all but his cousin talks in short sentences. Why has virtually the exact same deletion had such different consequences? The effect is called phenotype variability.

Phenotype variability even occurs within the SHANK3 gene. Two people with the same pathogenic variant can differ vastly in their ability to walk or talk. Once we accept the dilemma that similar deletions can have very different outcomes, we are in the right frame of mind to try understanding this phenomenon.

Explaining variation

So, how do we explain these dramatic differences? The answer is: there are many answers. There are so many ways similar deletions can have very different outcomes, that it takes a catalog of explanations to cover them. Here we go.

1. Loss of heterozygosity (a.k.a., hemizygosity). Small genetic errors occur all the time during development and in adulthood. Environmental factors from cosmic radiation to infections, sunburn to environmental toxins, can create small errors. Cells have mechanisms to repair errors, but one important hedge against serious genetic errors is the fact that we carry two of every gene (one from mom and one from dad). When a person or a tissue in the body has only one copy of a gene, there is an unfortunate opportunity for errors to go uncorrected. 22q13 deletion syndrome is the loss of some or many genes on one chromosome. This creates hemizygosity (“half as many copies”) of those genes. Any uncorrected error in the sole remaining gene can have a dramatic effect. Loss of one gene, then damage to the remaining gene is sometimes called a “2nd hit”. The error can be global (whole body and detectable with genetic testing), or it can be local (limited to one small region of the body, or one region of the brain). When it is local, it is undetectable and becomes an unexplained difference.
2. Imprinting. Imprinting is when one of the two inherited genes is silenced (turned off). Angelman syndrome is an intellectual disability syndrome with a number of similarities to 22q13 deletion syndrome. It is caused by imprinting that turns off an important gene. Not much is known about how imprinting and chromosomal deletions interact, but obviously it would be a problem if the only remaining copy of a gene was inactivated through imprinting. It would be another unexplained difference.
3. Impact of a rare gene variant or partial deletion of a gene. A gene that is completely deleted can be less damaging than a gene that has an abnormal sequence of nucleotides (called a variant). An example of a dangerous variant is when a gene mutation produces cancer. Exposure to too much sun or other carcinogen leads to cells with damaged genes. Cells with damaged genes often die out. But, some may grow into a tumor. In the case of 22q13 deletion syndrome, cancer is extremely rare. However, there can be other problems when a gene is a pathogenic variant, or when a chromosomal deletion removes only part of a gene. A partially deleted gene can start creating proteins that interfere with normal cell operation (see When missing a gene is a good thing.) So, a bigger deletion could be actually be less pathogenic than a slightly smaller deletion if the smaller deletion disrupts just part of a gene. It is probably not very common, but a partial deletion or unfortunate pathogenic variant could cause of more severe problems.
4. Gene combinations. Most geneticists evaluate a deletion based on the impact of individual genes. But, current scientific studies suggest a lot more complicated things can happen when multiple genes are involved. Research in autism spectrum disorder, schizophrenia and other disorders show that these disorders are often caused when a large number of common gene variants combine together in an unfortunate way. Each variant contributes in a small way. In some cases, there are a few important genes, but they have little or no impact unless many other genes are also involved. These gene combinations are subtle and still poorly understood. Older research used the term “genetic background” to act as placeholder. More recent work has led to a newer concept called “polygenic risk”. The main genetic differences between my cousin’s daughter and my son are from our spouses, who each contributed different background genetics. Somehow, the genes from my spouse collectively lead to life-threatening problems as a newborn when mixed with his 22q13 deletion. My cousin’s daughter received a less threating set of background genes from her father.
5. Mosaicism and somatic mutations. Recent evidence shows that it is possible for a genetic error to occur in one small region of the brain. That is, some people have gene mutations that impact only certain areas of the brain. These events might explain many individual variations, including things like learning disabilities. In the case of 22q13 deletion syndrome, these silent mutations are likely to have a much more serious effect if they occur in the region of 22q13. In cases of pathogenic SHANK3 variants, the impact of SHANK3 may be greatly amplified by errors in other genes in specific brain regions. Blood tests often do not show mosaic/somatic errors that may occur deep in the brain.
6. Genetic regulators (elements). Since 2009 the ENCODE genetics project (https://www.encodeproject.org/) and others have sought to find the bits and pieces of DNA that regulate genes. Genes make up the minority of DNA. Most of DNA is comprised of gene regulators. This is very easy to understand when you realize that skin cells, brain cells, intestine cells and liver cells all have exactly the same genes. The difference is which genes are turned on and which are turned off. Skin cells know they are skin cells and only use genes necessary for the skin. Brain cells only use genes necessary for brain. The DNA is regulated in each tissue to match the needs of that tissue. Chromosome 22 deletions not only knock out genes, they knock out genetic regulators. A 4.7 Mb terminal deletion may not hit any more genes than a 4.8 Mb deletion, but it may hit a crucial regulator site. Gene regulators are not impossible to detect, but they can be difficult to study. Whole exome sequencing, a powerful tool for finding small genetic errors, skips over most of the gene regulators. Gene regulators are a likely cause of many unexplained differences.
7. Healing. DNA can be a very sticky substance. When a terminal deletion occurs (the most commonly observed occurrence in 22q13 deletion syndrome), the broken end of the chromosome can pick up various bits and pieces of DNA as it “heals” (re-seals the ends). In the extreme case the chromosome forms a ring by attaching to its opposite end. In other cases the end of the chromosome may pick up bits and pieces of DNA, sometimes copies of itself. The random junk at the end of a deletion could have an important impact, although this has not been carefully studied.
8. Positional effects. Positional effects is related to genetic regulators. The location of a gene relative to other genes and regulators on the same chromosome can be important. Enhancers, for example, are regulator elements that increase the likelihood that a gene will get used to produce its product (usually a protein). The distance between a gene and its enhancer may strongly influence how effectively the enhancer operates. Deletions, especially interstitial deletions, can change the distance between DNA elements and ultimately influence the impact of a deletion. Positional effects help explain why two similar size deletions might have noticeably different outcomes.

Final thoughts

Given the complexity and many opportunities for unexplained variation, we can begin to appreciate why knowing an individual’s deletion size does not provide all the answers. But, even with the wide person-to-person variation, studies have shown that larger deletions have a more serious impact than smaller ones. Some of my other blogs discuss specific genes and even ways to explore the influence of genes of unknown function. Parents should appreciate that it takes time to incorporate new science into medical practice. Genetic reports in the future will likely say a lot more about specific genes than reports today. If your genetic report provides a list of genes lost in a deletion, hold onto that list. Genes of “unknown significance” may someday be identified as important.

arm22q13

Some previous posts:

22q13 deletion syndrome: the hope of precision medicine
How do we know which genes are important
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
22q13 deletion syndrome – an introduction

22q13 deletion syndrome: How do we know which genes are important?

When I get on Facebook I look for pictures of our 22q13 deletion syndrome kids.  Every time I see one I give it a “thumbs up”.  It warms my heart to see other parents share their pride in their children, even if our children are peculiar in some way.  Our snapshot may capture a funny posture or gait.  David is almost always looking away from the camera.  Some 22q13 kids are captured chewing on “non food items”.  We post photos to show our pride in accomplishments that would have been easy for most other kids.  Our children are usually not very photogenic, except to families and, of course, other parents of kids with 22q13 deletion syndrome.

There are some pictures that we don’t put on Facebook in deference to families that could not appreciate them.  There are pictures of feces on the sofa, self-inflicted injuries, frightening hospital scenes, and even pictures after an early death. The reality of 22q13 deletion syndrome is often not pretty.  However, our goal is not to provoke a reaction. We simply want to share joy or commiserate with our community, like all parents.

David, like the overwhelming majority of children with 22q13 deletion syndrome, has many things wrong.  He is missing more than one or two genes and the impact is pretty obvious.  Ninety-seven percent of children with terminal deletions are missing from about 30 to 200 genes (see Understanding deletion size).  Science can help us find ways to help our children.  The first step is to find out which gene causes which problem.  Fortunately for our children, science has a bunch of relatively new tools to help create this “genotype-phenotype map“.

First things first.  Let’s have a look at the list of genes that are lost with a 22q13 terminal deletion (the most common type of deletion).

This is a list of genes organized by deletion size.  The deletion size on the left corresponds to the list of missing genes of the same color on the right.  A 1 Mb deletion will delete all genes in dark brown, starting from RABL2B and ending with ALG12 (33 genes).  The next group (reddish brown) are missing if your child has terminal deletions of 5 Mb or more (16 more genes, giving a total of 49 genes).  That covers about half of all common terminal deletions.  Terminal deletions have been observed for sizes up to about 9 or 10 Mb.  The genes above that are usually missing only with certain interstitial deletions.

Ok, so now we have our list.  The crucial question is, which genes do what?  In the past few years scientists have built some rather clever and remarkable tools for figuring this out.  Here are some tools and some examples of how they can be used.

Comparing 22q13 genes with known genetic syndromes

Online Mendelian Inheritance in Man (OMIM) is a database of genes and the problems associated with them.  By choosing a trait like poor body temperature control (poor thermoregulation) or low muscle tone (hypotonia), you can find out what genetic disorders have that feature.  From that information, you can identify which genes are involved.  Sound complicated?  Not at all. If you go to the Human Phenotype Ontology web site and type in “abnormal muscle tone” it does the entire cross-reference in a few seconds.  Click the tab for “genes” and you get a list.  I did just that.  I found which genes match the list of 22q13 genes and highlighted them here.

What is interesting about this list is that only two genes are directly involved with the synapses of the brain (SHANK3 and MAPK8IP2).  Other genes linked to hypotonia have other important functions. One gene is important for the synthesis of neurotransmitters (SULT4A1). Some genes affect white matter and peripheral nerves (ARSA and SBF1).  Another gene affects the muscles directly (CHKB).  Some genes affect many organs (ALG12 and NAGA).  As I see it, each gene is an opportunity to find a treatment for our children.  If one gene is complicated and hard to study, there are other genes that might lead more quickly to important benefits, like new treatments.

Comparing 22q13 genes with genes that work specifically in the brain

If we are interested in behavioral problems and intellectual disability we can benefit from a recent scientific study that has created a list of genes that are specialized for the brain (Pandey et al., 2014).  Using a “gene expression atlas,” these researchers identified genes that are either used (expressed) at a very high level in the brain, or used much more in the brain than anywhere else.  The logic is simple, if the brain treats these genes as important, then they must be important.  

Only 4 genes show up.  These are obviously 4 genes that deserve careful research to help people with 22q13 deletion syndrome.  Two of these genes, MAPK8IP2 and SULT4A1 also appeared in the hypotonia gene search.

Comparing 22q13 genes with genes that evolved for a specific purpose

One of the most interesting new methods for understanding the role of genes comes from the study of how humans evolved.  I have already written about the value of this approach (see Is 22q13 deletion syndrome a ciliopathy ?).  There is an interesting website that automates the process of studying evolution. This approach, called “forward genomics” is more difficult to use than the previous two examples, but this method may solve some important problems.  I am very interested why David gets too hot in the sun and too cold after a bath.  That is, why does he have problems regulating his body temperature.  By studying the scientific literature on which animals are good at body temperature regulation and which animals are not, this web site will tell me which genes are involved.  My job is to read textbooks and papers to find out how well each of 27 species of animals regulate their temperature.  Once I do that, I can ask the website to scan the genomes of these species and identify which genes are associated with the emergence (or loss) of the ability to regulate body temperature. It is a fascinating approach and I am very eager to learn the results. The results may open the door to lowering the risk of febrile seizures.

Other methods

There are other methods for finding genes that affect our children in specific ways.  For example, gastroesophageal reflux was such a serious problem for David that he required major abdominal surgery (Nissen fundoplication).  A comparison of reflux with known genetic conditions (similar to the hypotonia example) provided no new information about 22q13 deletion syndrome genes.  However,  the search did produce a list of 48 reflux genes. How can we use the reflux gene list to learn more about 22q13 genes?  First, there is an analysis method called “guilt-by-association“.  This analysis will indicate which 22q13 deletion syndrome genes naturally operate in concert with the reflux genes.  A even more complex analysis tool for protein-protein interaction can identify which 22q13 genes have chemical interactions with reflux genes. I expect one or more 22q13 deletion syndrome genes will be associated with reflux after these analyses.

Tremendous progress has been made in the understanding of how genes contribute to disorders.  The best way science can help our children is by identifying the many different genes that cause the many different problems.  That is Step One and modern methods make that step much easier and more informative that the old methods of the past.  Step Two is to find treatments.  Many of these genes have been studied in great detail.  Some have related treatments either already in use or suggested by researchers.  As a parent, I want to see new treatments found for David.  As a researcher, I don’t understand why we are not taking these potentially fast tracks to treatment.

 

arm

Previous posts:
22q13 Deletion Syndrome: hypotonia
Understanding gene size
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding translocations in 22q13 deletion syndrome: genetics and evolution
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

22q13 Deletion Syndrome: hypotonia

A Sunday walk with David

Originally created 29 August 2015
Updated 28 November 2021
Available in Portuguese  https://pmsbrasil.org.br/sindrome-da-delecao-22q13-hipotonia

The joys of walking

Although he is a bit unsteady at times, David loves to walk. David began a day program after high school and he was assigned an aid new to the program. After one month the aid nearly quit! Keeping up with David’s constant motion — usually walking — forced the aid to become an athlete. After working with David for twelve years, she looks back at the experience in an appreciative way. David brought fitness into her life and the two of them developed a deep affection for each other. They enriched each other’s lives in many ways. Health from walking was an important one.

David has 22q13.3 deletion syndrome, also known as Phelan-McDermid syndrome (PMS). David, like many others with PMS, was born a “floppy baby”:  A general medical reference to an abnormal condition of newborns and infants manifested by inadequate tone of the muscles. It can be due to a multitude of different neurologic and muscle problems. See also Hypotonia. At age one, after daily work-outs and multiple physical therapy sessions each week, David developed the strength to lift his head and arms. He gradually learned to sit up, drag himself by his arms, and then crawl. Countless hours of therapy in a clinic and at home went into each milestone. We pushed him constantly for six years. Each time he improved, we “raised the bar”. Once David gained strength and basic skills, his mom, Carol, would exercise David at the grocery store by having him hold onto the side of the shopping cart as she pushed. One day, fascinated by a stack of bright red apples in the produce section, David let go of the cart and walked eight steps on his own to reach the stack of applies. Carol was caught completely by surprise. David reached the apples and everything ended up on the floor. The store staff came running and found Carol holding David, crying tears of joy. After six very long years, David had learned to walk on his own. Now, David I go on weekend walks together (see photo). Every time I walk with David, it warms my heart to watch him

Hypotonia 

Having very low muscle tone interferes with normal growth and development in many ways. Muscle tone is important for breathing in newborns (Lopes et al., 1981). David was born prematurely and he was on a ventilator for weeks. Low muscle tone slowed his recovery. Muscle tone is important for normal cognitive development and function (e.g., Jongsma et al., 2015). Gastroesophageal reflux plagues many children with PMS (including David) and is likely caused by low tone of the esophageal sphincter (Hershcovici et al., 2011). Other gastrointestinal problems likely result from muscle tone problems of smooth muscles. The most obvious problem with low muscle tone, however, is delayed or absent walking. Walking requires stable standing, which requires sufficient tone to hold the body erect. Building strength in David’s abdominal, back and leg muscles took years of work.

What is muscle tone and what interferes with normal tone? For skeletal muscle, “Muscle tone refers to the resistance that an examiner perceives when moving someone’s limb in a passive manner” (Mitz and Winstein, in Neuroscience for Rehabilitation, 1993). Normal muscle tone disappears when someone is knocked unconscious, or when the muscle itself is unable to support contractions. Diagnosing the cause of hypotonia in infants can be complex, especially in the presence of a genetic syndrome (Bodensteiner, 2008). In genetic syndromes that include both hypotonia and intellectual disability, the hypotonia is often diagnosed as “central hypotonia”: hypotonia caused by problems with the brain or spinal cord. However, the hypotonia associated with PMS may be from multiple causes. Certainly, it is not caused by any one gene. No single gene deletion or mutation has been identified that always causes hypotonia, and no one gene is essential for hypotonia. There is also no doubt that infant hypotonia is far more common in children with somewhat larger deletions (Sarasua et al., 2014, figure S1).

The severe hypotonia so often seen in infants with PMS may arise from multiple sources. Since finding ways to treat hypotonia could help children with PMS, understanding the causes will open the door to improving their lives.

Genes that directly affect synapses 

If your child with PMS was seen by a pediatrician or pediatric neurologist, it is likely the physician concluded that the hypotonia was of central origin (see, Bodensteiner, 2008). Although the conclusion would be based on accepted clinical practice, it would actually require a battery of tests to rule out other sources. Without other signs of major muscle or metabolic problems, the physician may be wise to avoid the additional tests that would be necessary. Right now, such testing is best done as part of a research study.

Which genes might contribute to low muscle tone of central origin? One obvious source of central hypotonia is a problem with synaptic proteins. For chromosomal deletions of 22q13.3, two proteins coding genes are nearly always deleted together: SHANK3 and MAPK8IP2. I have found only one published clear case where MAPK8IP2 and more proximal genes were deleted without impacting SHANK3 (Vondráčková et al., 2014). That patient had hypotonia. Thus, hypotonia can be caused without impacting SHANK3. What is lacking in PMS research are more studies of children with so called interstitial deletions. (See my blog: PMS, IQ and why interstitial deletions matter). Generally, hypotonia created by the deletion of SHANK3 is less than with deletions of any larger size. If we include pathogenic variants of SHANK3, we know that hypotonia with a SHANK3 variant is much less prevalent (33%) than hypotonia in patients with terminal deletions of 22q13.3 (65% to 75%), whether or not SHANK3 is involved in the deletion (Vondráčková et al., 2014).

Genes that affect brain development

In PMS, hypotonia of central origin is likely caused by the genes essential to normal to brain development. A review of PMS genes showed that 18 genes that are deleted in PMS patients are associated with brain development (Mitz et al., 2018). Of these, 10 genes are associated with reproductive fitness (e.g., necessary for normal health) based on their “pLI” scores: SHANK3, MAPK8IP2, PLXNB2, TUBGCP6, BRD1, TBC1D22A, CELSR1, SULT4A1, TCF20 (see Supplementary Table S2 of Mitz et al.). Since that study, The gene PHF21B has been added to the list as an epigenetic regulator of development (Basu et al., 2020). Thus, genes across the nearly entire 22q13.3 region associated with PMS are critical genes that participate in normal brain development. Any, and likely all, contribute to both the intellectual disability and the hypotonia of PMS.

Genes that may affect the environment of the central nervous system

We sometimes forget that the brain must have lot of things working properly for synapses to operate. For example, the brain is about 2% of our total body weight, but it uses up 20% of the oxygen we breathe (Rolfe and Brown, 1997). So, the blood flow from the heart, nutrition from the gut and oxygen from the lungs are of critical importance to human brain function. Any missing gene that might affect the brain’s ability to process energy in the mitochondria may impact synaptic function. Note that studies of rats and mice might be misleading. The rat brain, for example, uses only 3% of the oxygen they breathe for brain function. These mammals are not nearly as sensitive to the “energetics” of brain function as humans. The genes of 22q13.3 used by mitochondria appear to have a mixed impact on people with PMS (Frye et al., 2016). Beyond hypotonia of central origin, the same genes that affect the energy supply for the central nervous system can affect muscles directly. Muscles come in three flavors, skeletal, cardiac and smooth. They are all major users of energy.

SCO2 and TYMP are two mitochondrial PMS genes that are lost with relatively small deletions of 22q13.3. Individuals missing one copy of SCO2 and/or one copy of TYMP seem to do fine (Pronicka et al., 2013). However, if the remaining copy of either SCO or TYMP has an unusual variant, the results can be profound (Vondráčková et al., 2014). Of greater concern for most people with terminal deletions of 22q13.3 is the SULT4A1 gene. SULT4A1 is one of the few genes that has been implicated in intellectual disability and hypotonia based on a study of interstitial deletions. Recently, it has been shown the SULT4A1 protein is crucial for mitochondria function in the brain.

Conclusions

Most clinicians will conclude that hypotonia in children with PMS is of central origin. This is a good assumption, but further research is needed to look for more direct effects on muscle. There is strong evidence that many PMS genes contribute to central hypotonia, and central hypotonia occurs with all genotypes of PMS (see The four types of Phelan McDermid syndrome). On average, larger deletions lead to greater hypotonia. Developing broad and effective treatment for hypotonia will require understanding more about each gene’s contribution to maintaining healthy muscle tone (see 22q13 deletion syndrome: the hope of precision medicine).

arm22q13

Understanding gene size

Father’s Day gifts from David

Originally posted 14 July 2015
Updated 28 March 2021
Available in Portuguese https://pmsbrasil.org.br/entendendo-o-tamanho-de-um-gene/

David has a terminal deletion of chromosome 22 caused by a balanced translocation.  Like nearly everyone with 22q13 deletion syndrome (Phelan-McDermid syndrome), he is missing a lot more than one gene.  What, exactly, does that mean?

DNA and genes

Each gene is made up of many “bases”.  DNA has two strands (strings) that grip each other tightly. Imagine a bunch of bar magnets threaded onto a string like pearls. Now, in your mind, take two of these strings and hold them near each other.  Slowly bring them close together.  When they get near, the north poles of magnets from one string will start to find the south poles from the other string. When the magnets come together, opposite poles will grab each other. Anywhere north faces north, or south faces south, that pair will repel each other until one flips around and the opposites unite.  DNA is made of chemical strings that have opposite poles. These opposites find their mate and the two DNA strands lock together. Each time a north meets a south you get a “base pair”.

Magnets can only make one type of partnership (north attracted to south).  DNA actually has two kinds of partnerships from four chemical bases.  The bases are abbreviate T, A, G and C.  T and A attract each other. G and C attract each other. If you make a string like this: -T-A-G-G-C-A-, the matching string will always look like this: -A-T-C-C-G-. That is, the strings stick to each other in this way:

-T-A-G-G-C-A-
-A-T-C-C-G-T-

Voilà! You have a small strand of DNA. This miniature DNA has 6 base pairs. The order of the base pairs describe the protein that this segment of DNA makes. The lower strand is kind of mirror of the upper strand. If you know what is on one strand you can always figure out the other strand. Thus, we now know a bunch of properties of DNA:  1) The sequence of base pairs describes how to make a protein, 2) DNA is strongly stuck to itself, 3) DNA keeps a mirror copy of itself available at all times, and 4) the length of the DNA can be measured by counting the number of base pairs. There is a lot more to learn about DNA, but this is enough to discuss gene size.

Big genes are easier to find

In a previous posting I explained that 95% of all people with 22q13 deletion syndrome are missing at least 1 megabase (Mb) from their chromosome (see Understanding deletion size). 1 Mb means 1,000,000 (1 million) base pairs along the two parallel strands of DNA. Genes are segments of the long strings, like chapters in a book. And, like many books, some chapters are long and some are short. There are 32 genes in the distal 1 Mb of 22q13, many of which influence brain function. Chromosome deletion syndromes are inherently difficult to study because so many genes are involved. It is hard enough to study and understand the impact of losing a single gene. It is much harder to study and understand 22q13 deletion syndrome, where many genes are missing.

This problem with studying multiple genes is not unique to 22q13 deletion syndrome. It shows up in neuropsychiatric disorders like autism and schizophrenia, each of which have hundreds of associated “risk factor” genes. Autism, for example, results from various combinations of these many genes (see review by Gratten et al., 2014). Chromosomal deletions are known to operate in a similar way (see contiguous gene syndrome). Each missing gene weakens the normal operation of the brain. No one gene needs to be “dominant” for the combined loss to be devastating, especially when so many brain-related genes are missing at once.

Not everyone thinks of 22q13 deletion syndrome this way. Much of the current thinking about the genes lost in 22q13 deletion syndrome focuses on one or two genes that code for synaptic proteins. The term “synaptopathy” has been used a lot recently, but that word originates from the study of the inner ear where they are able to clearly demonstrate the relationship between synaptic function and hearing loss (Sergeyenko et al., 2013). The relationship between genes and function is not nearly as clear in 22q13 deletion syndrome. Synapses are involved, but the synapse may be only one site of dysfunction (see Is 22q13 deletion syndrome a ciliopathy?). For many years no one thought primary cilia were important. Now, ciliopathies are a recognized type of brain dysfunction despite the fact that synapses are also involved. Science often goes off in a wrong direction; it is part of the process. The other thing to remember about 22q13 deletion syndrome is that it is a neurodevelopmental disorder. Something goes wrong during the growth and maturation of the brain. There are so many things that can go wrong with too few or too many neurons connecting between two sites in the brain, neurons connecting to wrong places, wrong proportions of excitatory and inhibitory neurons, etc. Human neural development is one of the most complex processes in the animal kingdom. Errors in neurodevelopment are not just problems with synapses.

There is another reason that synaptic genes have taken the spotlight. The synaptic genes of 22q13 are relatively large genes. This is the theme of our blog.

In general, large defects are easier to notice than small ones. If we look at the history of 22q13 deletion syndrome, the first cases were discovered in people with very large deletions and with the most “severe” phenotype (symptoms).  As research in 22q13 deletion syndrome advanced, smaller and smaller deletions were identified and studied. The gene that gets most attention is a large gene that has a large effect when disrupted. So, why does size matter?

Genes lost in a 1 Mb deletion of 22q13 sorted by their sizes (mRNA size).
Right click on the graph to see a full size image.

The pie chart shows the 32 genes that are missing from about 95% of patients with 22q13 deletion syndrome. The genes are sorted in order of size. The largest gene is SBF1 and the second largest is SHANK3. The genes continue in descending order of size in a counter-clockwise direction. Although the reality is a bit more complex, it is generally true that the likelihood of a gene getting accidently modified, or otherwise disrupted, depends on the gene’s size. This pie graph shows that the 10 largest genes account for half of the “protein-coding” DNA in the first 1 Mb. To put it another way, you are twice as likely to disrupt SHANK3 than it neighbor MAPK8IP2, simply because SHANK3 is twice as large. SHANK3 is 16 times larger than SYCE3. So, when studying gene disruption, SHANK3 can show up more often simply because it is big. Scientists are aware of this size effect. They have developed gene disruption scores that take into account the size of a gene (i.e., probability of loss-of-function intolerance, pLI).

As I noted above, no one has carefully studied the impact of a complete deletion of SHANK3 without disrupting other genes involved in brain development and function. It may seem surprising, but a damaged gene can actually have a more severe impact compared to deleting the gene altogether (see When missing a gene is a good thing). A pathogenic variant of SHANK3 (an atypical and harmful version of the gene often resulting from damage) can contribute to 22q13 deletion syndrome, especially when SHANK3 is the only gene affected. But SHANK3‘s contribution to 22q13 deletion syndrome when many genes are missing is remains poorly understood. There are other 22q13 genes that have severe neurodevelopmental consequences after deletion whether or not SHANK3 is involved. Future blogs will discuss some of these genes in detail.

The take-home message is that certain genes are more likely to come under the microscope (literally and figuratively) simply because they are larger genes. Being large makes a gene easier to study (usually), but it does not necessarily confer importance. Measures like pLI have been developed to separate size from importance. This measure was used in a study of Phelan-McDermid syndrome that is discussed in the blog Which PMS genes are most important?.

When a gene gets popularized in the scientific literature, lots of papers are published on that one gene, at least for a while. Scientists will focus on genes that get them grants and publications. That is how science typically works, even if it is not necessarily the best approach to finding effective treatments that families really need. The direction of science can be influenced by patient groups, but choosing the right direction requires a deep understanding of the science (the current state of research), science (the discipline) and scientists (who do science).

arm

Previous posts:
Gene deletions versus mutations: sometimes missing a gene is better.
Is 22q13 deletion syndrome a ciliopathy?
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

Is 22q13 deletion syndrome a ciliopathy ?

David wearing his Roots, Rock, Reggae tee shirt.

What makes humans special?

There are different ways we, as humans, think of ourselves as special.  Of course, our children are special.  That goes without any further discussion.  But, as a species, special seems to come in a number of flavors.  Much of it centers around our flexible thinking and strategic planning. For example, one theory is that our brain allows us to plan and execute cooking.  Cooking provides concentrated nourishment essential for brain expansion (Warneken and Rosati, 2015). Of course, spoken language with complex syntax and hypothetical context is unique among humans. Other than being a funny-looking primate (upright walking and rather hairless), most of what makes us human is related to our brains: intellectual ability, sophisticated social interactions, symbols, language and future planning.

In the prior posting (“Understanding translocations in 22q13 deletion syndrome“) I explained how evolution opens new opportunities by duplicating a gene (creating paralogs).  One gene can remain in its original job and the other copy can mutate to find a new role. Once a gene has diverged into two genes, another wonderful door opens.  We have about 20,000 genes in our DNA, but we have about 800,000 ways to regulate when and where those 20,000 genes are used. So, not only is a paralog free to mutate, but where it is used and when it is used are now open to evolutionary pressures. In our example from last time, hypothetical gene L1 kept its original function while L2 took on a new role.

Even science makes mistakes

Not long ago, scientists comparing single-cell organisms with more recent animals noted that the whip-like tail (called flagella) used by bacteria, euglena and sperm to “swim”, did not exist on most cells of complex organisms, like mammals. There are a few places where we mammals use flagella. We use a version called microvilli to stir foods in the intestine.  Flagella are used by sperm.  Mostly, however, our bodies don’t use flagella. Yet, nearly every cell in our body has a tiny hair-like appendage structurally similar to flagella.  They are called cilia.  Scientists once decided that these were unimportant leftovers (fastigial organelles) from our swimming ancestors.  They were wrong.

Primary cilia in neurodevelopmental disorders
Enza Maria Valente, Rasim O. Rosti, Elizabeth Gibbs & Joseph G. Gleeson
Nature Reviews Neurology 10, 27–36 (2014)

Cilia are no laughing matter

Now that science is older and wiser we know that cilia are critically important organelles.  They are the nose of the cell, sniffing out molecules in the extracellular environment.  Most cilia it seems, have lost their hardware for swimming.  In doing so, they have gained the ability to provide specialize chemical interaction with other cells in the environment.  Free from their motile past, cilia have evolved into critical components of the body, including the rod and cone photoreceptors that give us the ability to see.  Cilia are very much like synapses (see drawing).  They have complex networks of proteins that create a special assembly for cell-to-cell communications.  Proteins are selectively transported to and from the membrane and the membrane has receptors like a synapse.  Also like synapses, mutations or deletions of genes that build or regulate cilia function can lead to partial or complete neuronal (i.e., brain) dysfunction (Metin and Pedraza, 2014; Valente et al., 2014).

Cilia are part of every neuron in the brain. They regulate where neurons go and which other neurons they contact during the dance of prenatal brain development.  They regulate brain growth, respond to inflammation and probably regulate many of the changes associated with puberty and aging.  Cilia are so important that there are now a series of syndromes recognized as ciliopathies: diseases/syndromes clearly caused by the deletion or mutation of a gene used within the cilia.  Is 22q13 deletion syndrome a ciliopathy?  There is reason to suspect so.

RABL2B somehow makes us human

There is a gene sitting on the very end of chromosome 22 that is deleted in every case of 22q13 deletion syndrome caused by a terminal deletion. Let me repeat that: every child with a terminal deletion is missing RABL2B.  Until recently, scientists had no idea the importance of this gene. RABL2A and RABL2B are paralogs of RABL2 found in nearly all mammals.  In rodents there is one role that has been studied: this gene is necessary to build working sperm tails (Lo et al., 2012). Male mice missing RABL2 do not seem to be very impaired, but they are sterile because of defective flagella.  Before the importance of cilia was fully understood, researchers studying 22q13 deletion syndrome assumed that RABL2B was not an important brain gene.  However, now we know two new things about RABL2B.  First, RABL2A and RABL2B can only be found together in primates, and only in primates with big frontal lobes: chimpanzees and humans (Wong, 1999).  Large frontal lobes might have been impossible without the duplication of RABL2 into the two paralogs RABL2A and RABL2B.  However, humans are much smarter (IQ equivalent) than chimpanzees.  And, only in humans, RABL2B is expressed at a disproportionately higher level in the brain than RABL2A (Kramer et al., 2010).  RABL2B is not a sperm motility gene (although it may contribute to sperm motility).  It is specialize for assembly of the cilia in the brain.  It is somehow important for what makes humans, human.

How does one study a gene so unique to humans?

Some genes are easy to study.  If my research was in molecular genetics, I would look for a large, popular gene that has a known function and can be knocked out in rodents.  Why bang my head against the wall working on a difficult gene?  RABL2B is difficult.  It does not exist in rodents.  It is small, so it is harder to pick up on gene microarrays/chips.  The RABL2A paralog is so similar to RABL2B that distinguishing between the two genes is difficult. RABL2B expression is a critical feature of brain function in humans, and expression regulation is more difficult to study than simply mutating a gene to see what happens. There are fewer tools for studying cilia than studying the nucleus or the synapse.  All-in-all, figuring out the exact role of RABL2B in the human brain will be inconvenient.  Molecular biologists steer away from unpopular genes that are difficult to study.  Unless we incentivize the study of RABL2B, we are unlikely to ever learn what damage its deletion does to our children.  Treatments and drugs aimed at other genes could end up being a waste of time if we are not prepared to understand and mitigate the impact of RABL2B.

A connection between RABL2B and IGF-1?

A very recent study of cilia (in fat cells) has demonstrated a link between IGF-1 receptors and cilia. As fat cells mature their cilia develop IGF-1 receptors, presumably to regulate fat cell growth (Dalbay et al., 2015).  It is not a stretch to expect that cilia in the brain have similar IGF-1 receptors. That means, giving IGF-1 to patients with 22q13 deletion syndrome may be ineffective if loss of RABL2B interferes with cilia formation and IGF-1 detection.

RABL2B and seizures

Cilia sample the extracellular environment (fluid space between neurons) and allow the neurons and other brain cells (e.g., astrocytes) the opportunity to respond to pathological conditions.  Seizures are an example of a pathological condition that impacts the extracellular environment:  seizures are associated with excess extracellular glutamate.  Extracellular glutamate can be damaging (Coulter and Eid, 2012).  From this perspective it is not surprising that ciliopathies are often associated with seizures (e.g., Oral-Facial-Digital Syndrome Type I).  About 30% of individuals with 22q13 deletion syndrome experience seizures.  Given the severity of seizures in some of our children, it seems likely that loss of RABL2B either precipitates seizure activity, or worsen seizures events by interfering with a cell’s response to extracellular chemical events. Many people with 22q13 deletion syndrome take seizure medications, which work for a while, then cease to provide benefit. Perhaps problems with cilia are responsible for making these medications lose effectiveness over time.

Why are there not clinical cases of RABL2B causing 22q13 deletion syndrome?

Thus far, there are no cases of 22q13 deletion syndrome where only RABL2B has been mutated or deleted.  (Although, every case of terminal deletion is missing RABL2B.)  One possible case of RABL2B deletion was reported, but further examination showed it was a more typical larger terminal deletion (see Patient 31 in Bonaglia et. al, 2011).  The problem in finding  a patient missing only RABL2B is two-fold.  First, most genetic tests cannot find a mutation or microdeletion in RABL2B.  The gene is small and most arrays/chips do not sample that gene sufficiently (if at all).  Exome sequencing can find a mutation or deletion, but geneticists have already convinced themselves that RABL2B is not an important gene.  Geneticists are likely to treat any mutation of RABL2B as irrelevant.  That is rather ironic, considering it would be relevant to the one-thousand patients with 22q13 deletion syndrome who are missing this gene.

 

arm

Previous posts:
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

Understanding translocations in 22q13 deletion syndrome: genetics and evolution

My dad

That’s my dad.  No, not the dapper gentleman standing in the back.  He is the diapered baby sitting in front.  One interesting thing about this circa 1924 photograph, I can tell  you unequivocally, is that both males in the photograph are carriers of 22q13 deletion syndrome.  I can say this even though neither man was ever genetically tested and neither ever had children with a known diagnosis of 22q13 deletion syndrome.  How can I be so sure? I spent several years contacting relative and having them tested.  It was an interesting and often challenging undertaking.  My motivation was to warn family members and at least prepare them for the possibility of having to raise a son or daughter with 22q13 deletion syndrome.  Along the way I received “thank you” from some family members and angry words from others.  Some family members simply did not want to know.  It is a story for some future blog, I suppose.

From this family research I was able to deduce that Joe, my dapper grandfather, was a carrier of 22q13 deletion syndrome with a “balanced translocation” of genetic material between chromosome 22 and chromosome 19.  Some of my grandfather’s siblings were carriers and at least two of Joe’s sons were carriers.  My dad was one of the carriers.  He had four sons, including me, and I am a carrier.  My dad did not have children with 22q13 deletion syndrome, but I had two.

The power of genetic principles

I know my dad was the carrier (not my mom) because a few of dad’s relatives are carriers. (See the line chart on my page “Who is arm 22q13?“.)   I was able show that my grandfather was a carrier using similar family evidence.  Genetics and inheritance follow certain rules and those rules can be used to peer into the past.  Genetics and evolution are two different aspects of the same rules, and understanding them can be very powerful tools for understanding where we come from and where we might be going.

Somewhere between 15% and 24% of all children with terminal deletions inherit that deletion from a carrier parent.  If your family has carriers, nature has provided a curious way to remove carriers from future generations: have small families.  This graph shows why.

(right click on graph to enlarge in a new window)

The main graph has three colored lines. (Ignore the small “inset” graph with bars; it provides details some researchers might want to see.)  The green line on the main graph represents what happens when people in the extended family have relatively large families (4.4 children, on average).  The black line shows the same process when the average family size is less (3.6 children per family).  The red line shows the impact of small families (1.5 children per family, on average).  What impact are we talking about?  The beginning of the graph starts today.  The end of the graph shows what happens after 10 to 50 generations from today.  Since most people assume 25 years for each generation to pass, the first 10 generations will take 250 years.  Here is the point.  If people have only small families, we can expect carriers to disappear (reach 0.0 on the scale) from the population in fewer than 10 generations.  However, if people choose to have large families (green line), carriers are unlikely to ever disappear (green line never reaches zero).

Let me be clear.  I am not advocating for any specific choice.  This is not about ethics.  In a sense, these are God’s rules.  They are inferred from the statistics of inheritance in the same way quantum tunneling is inferred from the statistics of nuclear emission.  I worked with a member of my family to generate this graph using a mathematical simulation.  I wanted to know how long 22q13 deletion syndrome has been in our family.  The answer comes from the green line.  Historically, my European ancestors had large families.  My great-grandfather had six children.  His children had an average of 4.8 children each.  These numbers suggest that the translocation could have existed in our family for tens of generations.

Promise

In my prior posting (“Understanding deletion size“) I promised to discuss a brain gene that is missing in 100% of the cases of terminal deletions.  I realize that explaining its importance will first require explaining a bit about evolution.  So, the rest of this blog will set the stage for judging the importance of a brain gene.

~~~~~ INTERMISSION ~~~~~
There is a lot of material here, so you are welcome to take a break before reading the second part.

Evolution:  There ain’t no missing link

Earlier I noted that genetics and evolution are closely related.  Describing evolution is simple in the same way that describing police work is simple. The task seems like it should be easy to explain, but the devil is in the details.  Many of the principles are not obvious at first, and both requires a lot of study.

Consider this make-believe story. A farmer has two children.  The son grows up to be a christian missionary in Africa and the daughter becomes an international arms dealer.  Their divergent lives lead to divergent branches of the family.  Years after dad passes away, two great-grandchildren meet.  One lives in a hut, is very religious and dresses modestly.  The other shows up on a yacht.  They are very different, but connected through a common ancestor (the farmer).  Evolution works the same way. The Chimpanzee is our closest living relative species. However, there was never a species halfway between Chimpanzee and Homo sapiens.  We share a common ancestor.  Some primate, extinct now, had members that experienced very different genetic and environmental events and each evolved into a different species.  These two offshoot species each underwent their own evolutionary history.

Primates (e.g., monkeys, apes, chimpanzees, humans) are special for a lot of reasons, but most notably for the development of higher brain function through the evolution of a new type of prefrontal cortex.  The new cortical areas help manage uncertainty, understand complexity and better imagine the future (Wise, 2008).  Rodents do not have an equivalent to the granular prefrontal cortex of primates.  Importantly, this area has undergone its greatest expansion in humans.  Two genetic features drive this type of dramatic specialization of brain function in humans: changes in the genes (either new ones or altered ones) and changes in when, where and how the genes are expressed.

Paralogs and gene expression

Here is a hypothetical example.  Let’s say a very early microorganism has a gene that we will call gene L.  Gene L is required for movement through its water environment.  Gene L is needed for “swimming”.   It is so important that the organism cannot survive any mutation of the gene. However, one day there is a genetic error during cell division and an offspring ends up with 2 copies of gene L (duplication event).  The new copy of gene L is somewhat “liberated”.  It can mutate and change without interfering with swimming, since the old copy of gene L is still available to do its job.  We name the two genes L1 and L2.  They are “paralogs” of the original gene L.  In our hypothetical case, gene L1 allows the organism to swim, and L2 is “free” to mutate and change.   One thousand years later, an L2 mutation event allows the organism to detect light in the environment.  The evolution of vision has just begun!  Thus, “duplication events” are crucial to evolution.  They copy important genes so that one copy can continue its original job and the other can do something new.  Sometimes, the two paralogs are very similar to each other, but are used differently in some crucial way.  L1 and L2 don’t have to be very different as long as having two different versions opens the door to new evolutionary opportunities.

Primate paralogs

The gene I will discuss next time is a paralog that only exists in ourselves and our very closest primate relatives.  That is, you and I carry a pair of genes that were duplicated and then evolved for specialized use in only the largest and most developed brains.  Moreover, humans have the most specialized use of the gene, and its specialization takes place in our brain. From an evolutionary point of view, this is a very special gene.  This gene is missing from every child with a terminal deletion, 98% of all known cases of 22q13 deletion syndrome. What critical functional role does it play in the human brain and how does that impact our children?

 

arm

Previous posts:
Understanding deletion size
Can 22q13 deletion syndrome cause ulcerative colitis?
Can 22q13 deletion syndrome cause cancer?
22q13 deletion syndrome – an introduction

 

 

 

Understanding deletion size

David has numerous other problems in addition to intellectual disability

Originally posted 4 June 2015
Updated 1 April 2021
Available in Portuguese http://pmsbrasil.org.br/entendendo-o-tamanho-da-delecao/

Probably everyone living with 22q13 deletion syndrome knows that it is much more than a disease of the brain. My son, David, is not unusual in that regard. He has flaky toenails, gastrointestinal (GI) problems, and poor temperature regulation. 22q13 deletions affect the entire body. I worry about painful conditions that he is unable to express to me (see Can 22q13 deletion syndrome cause ulcerative colitis?) or other medical condition that may shorten his life. That said, as parents we primarily see our child’s future most influenced by his intellectual disability: the loss of typical cognitive development. What causes this defining feature of 22q13 deletion syndrome?

Terminal deletions

Somewhere around 95% of individuals identified with 22q13 deletion syndrome have terminal deletions, where the chromosome has a piece broken off the end. About 10% of the deletions are inherited (unbalanced translocation), but the rest are not (de novo). The remaining individuals have interstitial deletions: the broken and missing material is somewhere inside the chromosome without affecting the end of the chromosome. 22q13 deletion syndrome was not originally associated with a single gene. But, disruptions of the SHANK3 gene became included under the umbrella name “Phelan-McDermid syndrome” (abbreviated as PMS) or “Phelan-McDermid deletion syndrome” (PMSD). The term PMS has been used inconsistently, sometimes excluding interstitial deletions and sometimes not. For a long time I avoid using the PMS name (see 22q13 deletion syndrome – an introduction). There is another reason to omit discussing single gene mutations (properly called “pathogenic variants”) when discussing a contiguous chromosomal deletion syndrome like 22q13 deletion syndrome. Single gene variants can have very funny and unpredictable effects. See my explanation (Gene deletion versus mutation: sometimes missing a gene is better). A variant can have no effect (benign), it can be a weak effect because we normally have two of each gene, or it can have a very strong “dominant negative” effect. A dominant negative means that the variant gene is worse than losing the gene altogether. Thus, variants of a gene like SHANK3 may have different effects, but the individuals with SHANK3 variants may not be representative of most people we know who have 22q13 deletion syndrome. Most of the people identified with PMS have a chromosomal deletion syndrome. There is important overlap, but there are also important differences. This article discusses chromosomal deletions rather than SHANK3 variants.

Even small terminal deletions cause a major loss of genes

What most people do not understand about chromosome 22 is that the 22q13 area is rich in genes near the terminal end. That is, deleting a small part of the end removes a lot of important genes. Here is a chart based on the most complete published study to date (Sarasua et al., 2014) and the most complete listing of genes available.

(Right click on the graph and open to a new window to see it full size.)

The graph has two lines drawn across the 22q13 region of the chromosome. The scale on the bottom is distance from the end of the chromosome. Zero is the terminal end of the chromosome (the end of the DNA). The numbers 1 through 12 are the distance in megabases (Mb) from the terminal end. Thus, small deletions are on the left, larger deletions are on the right.

The line in blue, shows how many people have a deletion of at least a certain size. The scale on the left shows the percentage of the population. For example, about 97% of documented cases of 22q13 deletion syndrome have deletions that are 1 Mb in size or larger (red arrow at 1 Mb). People with very small deletions are actually uncommon. It is far more common to find people with 1 Mb deletions or larger. In green, you can see how many genes are involved with each deletion size. The thick red arrows show that the same 97% of cases are missing a whopping 25% of the known genes in this region of the chromosome. The green line jumps up rapidly in the first 1 Mb. After the green line jumps up, it flattens out for a long stretch of the chromosome. From a genetics standpoint, people with 2, 3 or even 4 Mb deletions are not very different from people with 1 Mb deletions. So, 22q13 deletion syndrome is a syndrome of many genes for most people.

This chart also helps explain why the effects of deletion size have confused people (including scientists) for so long. There are so few cases of small deletions and so many genes, that researchers have never been able to tease out how individual genes contribute to the disorder (although many claims have been made). It has been confusing to families that deletion size does not easily explain difference among their children. Here we see one reason. Deletions smaller than 1 Mb are rare and terminal deletions between 1 and 4 Mb add very few additional genes. It makes sense given the shape of the green line. About 30% of the population has essentially the same size deletion.

You might ask, what kind of genes are in the “gene rich” 1 Mb part of the chromosome? Are they important to the hallmark trait of 22q13 deletion syndrome, intellectual disability (cognitive dysfunction)? The answer is a resounding, yes! There are 31 genes in the first 1 Mb and 10 of these are related to brain function. Thus, 97% of 22q13 deletion syndrome patients are missing 10 or more “brain genes”. An investigation into which PMS genes are the most likely to cause problems after a deletion narrows this list and provides a roadmap for research (see Which PMS genes are most important?). Genes likely to affect IQ are mapped in detail in this blog PMS, IQ and why interstitial deletions matter.

Brain genes

The 22q13 region has at least 19 different genes that affect the brain and 10 reside in the terminal 1 Mb region. These genes sculpt the developing brain, protect it from damage, regulate excitability (e.g., avoid seizures), maintain healthy tissue and regulate cell death. In my next blog I discuss one gene that is a real mystery. This gene is found only in advanced primate species (e.g., humans, chimpanzees). Moreover, it has a unique, specialized role in the human brain. We still do not know enough about this gene, but we should not ignore it, either!

arm

Previous posts:

Can 22q13 deletion syndrome cause ulcerative colitis?

Can 22q13 deletion syndrome cause cancer?

22q13 deletion syndrome – an introduction

22q13 deletion syndrome – an introduction

Originally posted 20 May 2015
Updated 3 April 2021
Available in Portuguese http://pmsbrasil.org.br/sindrome-da-delecao-22q13-uma-introducao/

Definition of 22q13 deletion syndrome

22q13 deletion syndrome has been defined different ways. Here is one that was on Wikipedia for a long while:

“…a genetic disorder caused by deletions or rearrangements on the q terminal end (long arm) of chromosome 22.”

In some research reports the role of one gene (out of 108 genes that may be deleted), SHANK3, is emphasized.

22q13 deletion syndrome is often called Phelan-McDermid syndrome (PMS) or Phelan-McDermid deletion syndrome (PMDS). 22q13 deletion syndrome was first observed as a terminal deletion of chromosome 22 (Nesslinger et al., 1994). Other major deletions in that region are still considered 22q13 deletion syndrome by some, but not by others (for example, see this letter to the editor). The term PMS was introduced by The Phelan-McDermid Syndrome Foundation, which adopted that name for the Foundation. An attempt to build a scientific consensus on the definition of PMS in 2012 ended in disarray. I am told that new discussions have been underway since early 2020. As of 2021 there has been no announcement from the Foundation.

Different scientists and clinicians often use the term PMS differently. Some use PMS to describe any intellectual disability or autism caused by deletion or mutation of SHANK3, excluding cases of interstitial deletion or pathogenic variants of other genes. For example, “…deletions and mutations that lead to a loss of a functional copy of SHANK3 cause Phelan-McDermid syndrome” (Betancur and Buxbaum, 2013). This follows the “SHANK3 hypothesis of PMS”, which is widely cited, but as yet, unproven. Other experts in the field define PMS as deletion or mutation of any gene(s) in the 22q13 region, as long as the individual presents with manifestations typical of 22q13 deletion syndrome: “Almost all of these deletions include the gene SHANK3…” (Original text of Phelan and McDermid, 2012, strangely, it was changed by the editor more than a year after publication). When this blog was originally written (May of 2015), the Foundation web site (http://22q13.org) avoided a detailed operational definition. Rather, it described “typical causes” that focused on SHANK3, somewhat echoing the SHANK3 hypothesis of PMS. This blog space discusses the roles of SHANK3 and the other 107 genes that are involved in deletions that lead to 22q13 deletion syndrome. It goes into great detail and discusses the science behind each relevant gene. Along the way it teaches three things related to science: 1) the science behind our understanding, 2) how science works and 3) the human component of scientists doing science.

Science, medicine and PMS

My son, David has a 22q13 deletion, but he also has an overdose (trisomy) of chromosome 19. Is that PMS? Who knows? Most people say “yes,” but it is hard to diagnose without a consensus definition. This brings into focus the role of clinical geneticists. They operate under certain principles and rules, although somewhat different rule sets prevail in different settings (e.g., U.S. versus European). Recently, I was asked by a mother to make sense of a  genetic report from a prestigious university-affiliated hospital. The report had errors owing to the confusion between what constitutes PMS and the phenotype characteristics of 22q13 deletion syndrome. Because of those errors, the recommendations were flawed. Another parent showed me their child’s genetic report with a conclusion that seemed absurd. It centered around the problem of guessing whether or not a radically different version (“variant”) of a gene interfered with the gene’s normal function.

In fact, much of medical genetics is guesswork. One goal of my blog is to inform parents, scientists and clinicians. I separate facts from conjecture. I introduce concepts from the latest scientific studies and how they may apply to PMS. The scientist’s perspective is often very different from the clinician’s, especially when it comes to genetics. Of course, the parent’s perspective is different from the others. Parents mostly interact with clinicians. This blog is an opportunity for parents to incorporate science into understanding their child’s disorder. Very few geneticists ever see even a single case of PMS. Faced with a potential case of PMS, a geneticist may read one or two sources and then render his/her professional decision. Unbeknownst to the geneticist, the sources may not be ideal selections for the case in front of them. This blog will supply a more complete view of PMS to help guide their research and their decision making. Scientists who study PMS see the disorder from either a technical perspective, or as a clinical practitioner. Living the life of a PMS parent paints a different picture. It is a case study (my child) in unfathomable depth and under infinite magnification. Scientists need to hear from parents. Rarely does that occur in the vernacular of science. Scientists have much to learn from parents who speak their language fluently.

This blog space

As with the definition of PMS, much of science can be messy. Most discussions of science oversimplify the realities. This blog is dedicated to those parents, clinicians and researchers who try to make sense of genetic defects that can arise from the q13 region of chromosome 22, whatever you call it. It is aimed at parents who live with it and deal with it every day. It is for anyone who wants to understand more. All sources of information are biased and perhaps my bias suits you, or at minimum, is informative.

There are only about 2 to 3 thousand identified cases of 22q13 deletion syndrome world-wide. That leaves little opportunity to find a parent who has raised a child and is also a scientific expert in the field. There are a few. As one of the few, I felt obligated to start a blog. So, that is what I have done. You can click on Who is arm22q13 to learn a little more about me or explore my social media profiles listed below. Please read this disclaimer and keep it in mind as you read my blog posts. When I write my blog, I am not a representative of any organization. I represent only myself.

DISCLAIMER:

This blog will be full of statements: scientific, personal, experiential, opinionated, political and maybe even hostile. All of the statements are mine, alone (except when explicitly indicated otherwise). None of these statements are to be taken as a reflection of my employer or any other organization. I make no pretense of representing any organization or employer. Any errors, misstatements, wrong facts, wrong attributions or other misdeeds are my sole responsibility. One of my past employers is an agency of a government. I do not represent or speak for the agency or the government either on or off the record. This blog is a personal undertaking for the benefit of my fellow parents who must deal with the often painful reality of raising a child with 22q13 deletion syndrome. They deserve all the help and love we can offer. 

arm

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