David relaxing in the car during a drive through the countryside
Originally posted 10 April 2024
In 2022 a group of scientists associated with the Phelan McDermid Syndrome Foundation (PMSF) published a consensus paper that addressed a controversy around what constitutes Phelan McDermid syndrome (PMS). It was an important step toward defining the disorder, providing guidance to geneticists, and working towards an ICD code for diagnosis and medical reimbursement. Although the consensus opinion was not universal, the clarification was welcome. These experts split PMS into two mutually exclusive classes: “PMS-SHANK3 related” and “PMS-SHANK3 unrelated”. It is a simple, clear dichotomy. But genetics are rarely simple, as we shall see.
The thinking at the time was from the perspective of genetic testing. If the test result includes an abnormality of the SHANK3 “coding region”, then it gets the designation PMS-SHANK3 related. All other test results would be PMS-SHANK3 unrelated.
The people who hammered out this definition (distinction) are all experts. They were fully aware of all the possible ways genes can contribute to a disorder. There are dissenters, also experts, who feel that if SHANK3 is not involved, then the name PMS should not be applied. The consensus paper explains that people with interstitial deletions (PMS-SHANK3 unrelated) do not appear to have a syndrome different from PMS. There is also no transition from PMS to some other syndrome as deletion sizes get larger, whether or not SHANK3 is involved. There is no clear evidence that PMS-SHANK3 unrelated is some other type of disorder. The key concern for dissenters is that genes other than SHANK3 do not contribute to the disorder in the same way that SHANK3 causes PMS.
However, what if we can show that other genes on chromosome 22 contribute to PMS through their impact on SHANK3, or their impact on the molecules that interact with SHANK3? If the genes of 22q13.3 (the site of interstitial deletions) have such direct impact on SHANK3, then perhaps the term “PMS-SHANK3 unrelated” is misleading. If the biology of PMS-SHANK3 unrelated is highly related to SHANK3, then a distinction may not be warranted. The latest evidence suggests there are at least six genes that impact SHANK3 (and partner molecules), and these genes contribute to PMS when deleted. I would call these genes SHANK3 related.
The six genes and the details of how they interact with SHANK3 are discussed in a recent paper. The paper is open source (anyone can read it), but very technical. Full disclosure: I helped write the paper, so this blog post is undeniably biased. People who have read my previous blog posts should know that I have long been a proponent of looking closely at the many genes of PMS. The paper is a review of work done by scientists primarily between 2017 and 2024. Science is a continuous process and during this period enough information came to light to explain the tight relationships between six genes (PLXNB2, BRD1, CELSR1, PHF21B, SULT4A1, TCF20) and SHANK3. The first two genes are physically close to SHANK3 on chromosome 22, and thus are deleted in most deletions that include SHANK3. The remaining four genes are fairly evenly spaced across the last 4 Mb (megabases) of the PMS region of chromosome 22.
All seven genes (including SHANK3) impact brain development and all are involved with the process of inflammation. In this case inflammation includes “cellular stress”, “mitochondrial function”, and “recovery from injury”. These are all related processes, and all known to exacerbate PMS. So, impact on early development and response to stress and injury are features common to all of the genes.
Three of the genes (BRD1, PHF21B and TCF20) regulate what other genes do in the brain. For example, all three regulate the activity at synapses, the part of neurons that SHANK3 regulates. In addition, two more genes (PLXNB2 and SULT4A1) are directly involved in synaptic function. In fact, SULT4A1 not only regulates the same glutamate receptor as SHANK3, but it also regulates breakdown of SHANK3 at the synapse.
At this point it should be clear why PMS-SHANK3 unrelated may not really be unrelated to SHANK3. The six genes listed above join SHANK3 in shaping the development of the brain, the response of the brain to insults, and the operation of the synapses – the most important role of SHANK3 in the brain. It should also be clear why people with interstitial deletions typically have symptoms consistent with other cases of PMS. Likewise, it should also be clear why people with larger and larger deletions tend to have more severe PMS (see: Why PMS is worse for people with larger deletions).
The very close association between SHANK3 and at least six other PMS genes has a number of ramifications. First, while the distinction between SHANK3-related and -unrelated is sensible from the point of view of genetic testing, it may not be a good way to think about PMS as a disorder. Second, treatments that target SHANK3 will likely miss other relevant genes that tightly influence SHANK3, and thus may not produce very satisfying results in patients with chromosomal deletions. One way to think of the problem is that SHANK3 treatment in people with deletions essentially expands the number of PMS patients with interstitial deletions. It may be difficult to distinguish between a treatment that is not effective for SHANK3, and a treatment that is not effective because of other genes. Finally, when we are searching for an effective treatment for SHANK3 haploinsufficiency, maybe we should also look at treatments for one or more of the other genes. We may be missing out on developing additional valuable treatments.
Originally posted 30 December 2019
Updated 4 March 2022
Regression is one of the scariest words among the Phelan McDermid syndrome (PMS) community. It is not clear how common regression is in PMS. Recent and ongoing studies are addressing the problem. Regardless, regression does occur, can be unpredictable, and it can sometimes be devastating. After years of hard work helping your child navigate a world that is well beyond his or her comprehension, you then lose some or all of the gains to an invisible and unforgiving demon. In the PMS world, you either loath regression or fear its occurrence.
Three papers recently looked at regression in PMS, each taking a somewhat different approach. Reierson et al 2017 used scores from the Autism Diagnostic Interview-Revised (ADI-R) based on 50 patients seen in their clinical study. Verhoeven et al 2019 studied 24 adult PMS patients with psychiatric presentations and followed them over years. Kolevzon et al 2019 did a detailed literature review of PMS and SHANK3 mutations (denovo deleterious variants). This blog will be mostly a technical overview of the Kolevzon paper, with some additional notes from the Verhoeven study, both aimed at patients over 12 years of age and who present with psychiatric issues or sudden loss of function.
All of the authors have struggled to define “regression” and identify trends in occurrence and treatment methods. I am told there are important differences between “regression” and psychiatric decompensation. I’m not a psychiatrist so I won’t untangle the technical differences. But from what I read, either type of event can be a setback. Psychiatric decompensation seems to bring with it the likelihood of recovery and can repeat (episodes).
If you don’t want to read my full post, here some of the take-home messages.
1) There is yet no clear measure of how often regression or psychological decompensation occurs with PMS.
2) Relying on previous papers is difficult because not all descriptions in the literature provide sufficient detail about each patient.
3) Diagnosis of psychiatric illness is more difficult in people with low intellectual function and absent of functional language.
4) People with SHANK3 variants (“mutations”) and tiny chromosome deletions are a small minority of identified PMS cases. In general, they are more functional than others with PMS (higher intellectual function, better language and better motor skills). This minority are the patients who most often present with psychiatric problems. The great majority of patients with deletions greater than one or two Mb (more common PMS patients), show psychiatric problems far less often, at least given the limited research done so far.
5) When looking at the psychopathology overlap between SHANK3 variants and typical PMS patients, the common feature seems to be bipolar disorder.
6) In spite of the difficulties studying psychiatric problems in PMS, following best practices of psychiatric care used in the general population can be helpful for people with PMS.
7) Over-medication and certain medication choices can harm, rather than help.
8) Proper personal and social care of the adult with PMS is important for the successful management of regression and psychiatric problems.
9) These early studies provide a first, and perhaps hopeful picture, but much more work needs to be done to understand the natural history of PMS and the risks we face as parents.
Kolevzon and his co-authors specifically excluded individuals with interstitial deletions (called PMS-SHANK3 unrelated, explained here). That is, they chose to only study the category of PMS called PMS-SHANK3 related. The Kolevzon paper could have included interstitial deletions to look more closely at the causes of PMS regression. It is an unfortunate omission that I have warned against in the past (see: PMS, IQ and why interstitial deletions matter and We need to study interstitial deletions to cure PMS).
Most of the important information from the Kolevzon paper is in their Table 1, which is the result of an in-depth search for papers on PMS patients. Table 1 provides an excellent resource to the PMS research community despite ignoring interstitial deletions. I spent a few hours hunting down the original articles listed on the table and studying them. PMS science benefits greatly from this work. The table itself has a flaw that readers should be aware of. The reference numbers of the table are messed up. The following reference numbers are wrong (4-6, 8-12, 15-19, 22-25, 30, 32, 35-42). The author names and dates are correct, so there is no problem if you simply ignore the reference number and do a text search for the author name.
As noted above, it is difficult to define regression and the authors did not clearly describe the difference between regression and psychological decompensation. Presumably, experts in the field recognize the distinction. Regardless, the Kolevzon study required some rule set to execute a literature review. These authors chose: “Sudden change in the psychopathological presentation”. They focused on patients older than 12 years. The literature search looked for: 1) psychiatric decompensation, 2) loss of skill, 3) sudden behavioral change.
They found 56 cases, including 15 cases of ring chromosome, referred to as r(22). While it is laudable to dig deep into older medical literature and pull out r(22) cases, most of these predate the time when deletion size was easy to measure. There is another problem with evaluating r(22). Just as interstitial deletions can cause PMS without disrupting SHANK3, r(22) can cause developmental delays without disrupting SHANK3. In fact r(22) can cause developmental delays without disrupting any gene (see: Guilherme et al 2014). Also, as the Kolevzon paper points out, r(22) can lead to Neurofibromatosis 2 (NF2) disorder, which can be degenerative. The authors are careful to identify which results are dominated by r(22) cases. I think only the most recent cases (cases 19, 35 and 51) are helpful to the study. The earlier cases date from 1985 to 2007. One other case, 32, does not seem to provide much information.
By removing cases of PMS deletions that lack deletion size information, we can explore the impact of deletion size. What we discover is that psychopathology is almost exclusively diagnosed in patients with small deletions. The plot below uses the Kolevzon data from Table 1. It shows the frequency of diagnosis (number of cases) as a function of deletion size. SHANK3 variants are not included.
Psychiatric problems show up mostly with small deletions, which are uncommon.
To understand these results, remember that 95% of people identified with PMS have deletions greater than 1 Mb (see: Understanding deletion size). The cases found by Kolevzon are heavily weighted towards small and very small deletion. That is, most cases of psychopathology in the literature come from deletions that represent about only 5% of the PMS population.
If we merge the data from SHANK3 variants with the smallest deletions (size 0 to .9 Mb), the first bar more than doubles in size. We can conclude that psychopathology shows up almost exclusively when chromosome 22 is disrupted near the terminal end. That is, it shows up in the atypical cases where damage is limited to primarily SHANK3 disruption. Is this generally true for PMS or just for the sample of papers found during the literature search? We don’t know. For the moment, it is the best data available.
The authors suggest that because people with SHANK3 variants and tiny deletions are higher functioning, psychopathological changes might be easier to observe. Indeed, in their sample the cases of SHANK3 mutation and small deletions had both significantly higher intellectual function and motor skills. I suggest another possibility. It may be that loss of genes in the 2 – 4 Mb region of the chromosome help stabilize brain function. The most likely candidates in this region of the chromosome are BRD1, TBC1D22A, GRAMD4, CELSR1. I have discussed some of these genes in detail, in my other blogs (see 22q13: a hotbed for autism and intellectual disability genes?, and CELSR1: Do some people with PMS have more fragile brains?) Whether these genes seriously impact brain function, stabilize the brain, or both, there is no doubt that these genes greatly influence the outcomes of patients with PMS. The importance of these genes have come up in other studies, as well. These genes likely also contribute to the impact of PMS from interstitial deletions (see: Which PMS genes are most important?).
These initial results are concerning to families dealing with very small deletions or SHANK3 variants (limited primarily to SHANK3 disruptions). On the other hand, most of the Kolevzon study has less relevance to 95% of families. Still, there are 8 of the 56 patients that represent typical size deletions: P20, P21, P23, P24, P28, P33, P47 and P50. It is valuable to look specifically at these typical cases of PMS to see what might be more generally relevant to PMS families. For example, none of these 8 patients were diagnosed with psychosis and only one had tremor. Thus, psychosis and tremor might not be very relevant to most families. On the other hand, only typical PMS patients lost the ability to walk. Catatonia was seen in only one typical PMS patient (P50).
One feature in common between typical PMS patients and the uncommon cases of SHANK3 disruption is the incidence of bipolar disorder. Half of the typical PMS patients were diagnosed with bipolar disorder, as were many of the patients with SHANK3 disruption. This observation suggests a common mechanism, likely the SHANK3 gene. Do PMS patients with interstitial deletions have bipolar disorder? That would be an extremely valuable question to explore. Reviewing cases of interstitial deletions may support or refute the specific role of SHANK3 in bipolar disorder.
Verhoeven et al observed a similar overlap between typical PMS and cases of SHANK3 disruption. They state: “Based on actual psychiatric classification, in 18 patients, a diagnosis of atypical bipolar disorder was established of which symptoms typically started from late adolescence onward. In most patients, treatment with mood stabilizing agents in combination with individually designed contextual measures, and if indicated with the addition of an atypical antipsychotic, resulted in gradual stabilization of mood and behaviour.” Note the phrase “with individually designed contextual measures”. Proper social treatment of the person with PMS appears to be important for “recovery” (not a technical term).
Kolevzon et al discuss which treatments were more or less effective for both the SHANK3 disruptions and typical PMS patients. The overlap with Verhoeven is very encouraging. Together, the agreement between recent studies on regression and psychopathology provide improved guidance for the medical caregivers who may encounter bipolar disorder in PMS patients. The work of Kolevzon is especially valuable for cases of SHANK3 disruption (variants and very small chromosomal deletions). The number of cases where the PMS adult was stabilized was encouraging in both studies, which provides hope for families facing adult psychopathology.
It will take more studies to make strong statements about regression and psychological decompensation in Phelan McDermid syndrome. These early studies hint at the type of problems that do occur in PMS, the types of remediation that have been beneficial and the differences that appear to be associated with deletion size.
My son, David started life with some rather major problems. He was 6 years-old before he could walk and 9 years-old before he could eat by mouth. He never developed speech. His developmental trajectory has been slowly upward. At age 34, he has not had a regression or anything that could be called psychological decompensation. (His mom is qualified to recognize decompensation.) His deletion size is unknown, but I would wager it is not a small deletion. Does his stability come from missing certain genes?
When we moved David to a new group home two years ago, his mom and I were vigilant about managing the changes and constantly gauging his comfort. We were, frankly, scared that he would decompensate or regress. The transition was successful and David feels at home. But, what will be the next challenge and how will David respond? We don’t know.
David has both intellectual disability and some features of autism spectrum disorder.
Introduction
While SHANK3 has been identified as a major contributor to intellectual disability and autism spectrum disorder, there has always been a lingering question of why people with Phelan McDermid syndrome (PMS) have very similar symptoms (phenotype) whether or not SHANK3 is impacted. That is, people with terminal deletions that remove SHANK3 are not very different from people with so-called “interstitial deletions”. This observation is the main reason why the current definition of PMS says that SHANK3 is usually, but not always, impacted (see The 22q13.3 Deletion Syndrome).
Genes with a high “pLI” (genes that are very rarely mutated in the general population) are associated with intellectual disability. For 22q13 they are: SHANK3, MAPK8IP2, PLXNB2, TRABD, PIM3, ZBED4, BRD1, TBC1D22A, GRAMD4, CELSR1, SMC1B, PHF21B, PRR5, SULT4A1, SCUBE1, TCF20, SREBF2, and XRCC6. Most, if not all of these genes contribute to intellectual disability, although some are more certain. While 18 genes may seem like a lot of genes, remember that PMS can delete up to 108 genes. Still, it is impressive that nearly 17% of the genes of 22q13 potentially contribute to intellectual disability. The high number of these high pLI genes explains why intellectual disability is generally more severe with larger deletions, and it explains why interstitial deletions can also produce moderate to severe intellectual disability. But, what about autism spectrum disorder (ASD)? The frequency of ASD in PMS is an area of debate, various studies report anywhere from under 30% to over 70% incidence of ASD. The natural history study of PMS should narrow this range. Regardless, ASD is a component of the PMS population.
So far, three genes of 22q13 have been clearly associated with ASD: SHANK3, MAPK8IP2 and SULT4A1. It is probably no accident that these three genes also show up as high pLI genes.
What is new
Two new lines of research have identified new candidate ASD genes on 22q13. One candidate gene has arisen from studies of epigenetics, the regulation of genes. Genes occupy only about 1% of the DNA. Much of the remaining DNA is dedicated to controlling when and how often a specific gene is put to work. Gene regulation is essential for successful development of the brain (and all other body parts) and for responding to changes in the environment. By environment we can refer to the external environment (e.g., adapting to colder weather) or we can be concerned with the internal environment of the body (e.g., building muscle with increased exercise). There is earlier data suggesting a weak contribution of the gene BRD1 to ASD (see Which genes cause brain abnormalities in Phelan McDermid syndrome?). However, epigenetic studies have reinforced the connection between BRD1 and ASD (see Next-gen sequencing identifies non-coding variation disrupting miRNA-binding sites in neurological disorders). So, BRD1 can be considered an autism-associated gene.
Perhaps more exciting is the latest work on “second hits”. Second hits are when one gene is lost or damaged, and then the remaining copy of the gene is also affected. Second hits matter because some genes can operate just fine with only one copy. We have two copies of most genes in our DNA, one from each parent. When a person has a chromosomal deletion, which is what happens in over 95% of the cases of PMS, that leaves the person with only one copy of those genes. A second hit is when the remaining copy has a mutation, or even just an dysfunctional variant inherited from one parent. What do we know about second hits? A new paper has identified up to 4 genes of 22q13 that may contribute to ASD, given a second hit (see Recessive gene disruptions in autism spectrum disorder). One PMS gene stands out, KIAA0930 and three others are additional candidates, CHKB, ARFGAP3, and ZBED4.
This rounds-out the current full list of 8 ASD-associated genes: SHANK3, MAPK8IP2, CHKB, ZBED4, BRD1, KIAA0930, SULT4A1, and ARFGAP3.
Reveiw
PMS is most often a terminal deletion of 22q13. A deletion of the chromosome that includes SHANK3 is the most common deletion, but sometimes a deletion does not disrupt that gene. Interstitial deletions can disrupt up to 17 other genes likely to contribute to the moderate to severe intellectual disability that characterizes PMS. Infrequently, a second hit on one or more of 7 genes in the interstitial region of 22q13 may also contribute to autism spectrum disorder. In the overwhelming cases of PMS (called terminal deletions), many genes contribute to intellectual disability and, if autism spectrum disorder is part of the clinical picture, SHANK3 and one or two other genes may also be contributing to that aspect of the disorder.
Brain structure and functional abnormalities have been reported in Phelan McDermid syndrome (PMS) by a number of different investigators, including a a recent study associated with the Developmental Synaptopathies Consortium natural history study of PMS (Srivastava et al 2019). Prior studies have found abnormal formation of the cerebellum (Aldinger et al 2013), abnormal function of the cortex and amygdala (Philippe et al 2008), micropolygyria (Kurtas et al 2018), as well as commonly observed thinning of the corpus callosum and the presence of arachnoid cysts.
Brain abnormalities can provide important clues to understanding what goes wrong in PMS. They also could serve as “biomarkers”, biological measurements for early indicators of severity or evidence for treatment effectiveness. Readers of my blog will recognize that I spend a lot of time identifying which genes are most important in PMS. You need to know which genes are causing what problems to have any hope of finding effective treatments. My son, David (see picture), is waiting for new treatments.
So, what do the brain structural studies tell us?
SHANK3 variants can impact the proper development of white matter, but have minimal impact on the gray matter of the brain (Jesse et al 2020). Gray matter is where the neurons and their synapses reside. White matter is made up of the long, thin “axons” that travel together connecting one region of the brain with another region.
Aldinger and colleagues (Aldinger et al 2013) studied 10 subjects with PMS using x-ray images. Eight of the 10 subjects showed abnormality of the cerebellum (mostly gray matter) in addition to thinning of the corpus callosum (white matter) and enlargement of the cerebral ventricles (fluid space of the brain). Although there was no clear effect of deletion size, mutation of SHANK3 was not sufficient to cause cerebellar problems. They identified MAPK8IP2 and PLXNB2 as the more likely candidates for cerebellar malformation based on preclinical mouse studies.
The study by Philippe (Philippe et al 2008) had similar results from 8 PMS subjects. Three of the 4 subjects with small deletions (150 Kb or less) had no cerebellar or other major magnetic resonance imaging (MRI) results. The 4th subject with a small deletion had the least impressive positive finding. Thus, using MRI, there was a clear effect of deletion size, with small deletions have little or no effect. Four deletions of 1 to 9.3 Mb in size had stronger effects. Like the Aldinger study, SHANK3 did not seem to be a good candidate for most of their findings. In addition to cerebellar malformation, the group studied brain function using positron emission tomography (PET). They showed a group effect of amygdala dysfunction. Importantly, they used children with intellectual disability as their control group, a much stricter standard than other PMS studies.
The Srivastava study (Srivastava et al 2019) showed reduced size of the dorsal striatum, which is the opposite effect that loss of SHANK3 has in mouse models of PMS. Together, the results suggest that genes other than SHANK3 are driving brain malformation. SHANK3 can contribute to thinning of white matter, but is not the cause of other brain malformations.
Which genes might be driving the observed effects? Is there a smoking gun? To be a smoking gun, a gene that drives malformation should meet most, or all, of these criteria:
is strongly associated with a human neuropsychiatric or neurodevelopmental condition
causes reduced brain size in the striatum
impacts the amygdala
There are 6 genes on chromosome 22 that meet criteria 1 and 2. They have a high pLI score and are very frequently lost in PMS. They are located within 1 Mb of the chromosome terminus, which accounts for 95% of patients with a 22q13.3 deletion (see my blog Understanding deletion size). Those genes are: MAPK8IP2, PLXNB2, TRABD, PIM3, ZBED4 and BRD1. Of these, 4 genes meet criterion 3, being highly expressed in the cerebellum: MAPK8IP2, PLXNB2, ZBED4 and BRD1. Of these, 3 genes are associated with neuropsychiatric disorders. BRD1 is strongly associated with and schizophrenia. MAPK8IP2 is weakly associated with ASD (see my blog Which PMS genes are most associated with Autism?), and ZBED4 is weakly associated with schizophrenia. This leaves BRD1 as the strongest candidate gene for brain abnormalities/malformation in PMS.
Animal studies of BRD1 agree with the results from PMS imaging studies. Per Qvist and his colleagues in Denmark have been studying the BRD1 gene for some time. They have shown that loss of one copy of Brd1 in mice is sufficient to reduce cerebellum size, reduce striatum size and reduce the size of the amygdala (Qvist et al 2018). BRD1 is by far the strongest candidate gene for causing altered brain development, especially in gray matter.
The importance of BRD1 in PMS goes much further than structural brain anomalies. It has been known for some time that BRD1 impacts 100s of other genes through gene regulation (epigenetics), and the role of BRD1 shifts from development in utero (fetus) to a different role after birth (Dyrvig et al 2017). Now, a new epigenetics study of people with PMS Type 1 (terminal deletions) confirms that loss of BRD1 produces widespread changes in the human genome. BRD1 is a crucial gene lost in people with deletions greater than 1 Mb.
There is little doubt at this point that BRD1 plays a crucial role in PMS.
PMS is a contiguous chromosomal deletion syndrome, meaning that larger deletions interrupt more genes of importance. BRD1 is a critical gene for brain development. If we want to understand PMS, we need detailed studies that explore more genes like BRD1. One great way to study the impact of different genes is to look more deeply at the phenotypes and genotypes of people with interstitial deletions (PMS Type 2). A treasure trove of new information awaits these studies. Not exploring these candidate genes of PMS is a waste precious time. My son, David, and so many others, are waiting for these studies. We need to do the best possible science if we are ever going to find effective treatments.
In the previous blog we learned which Phelan McDermid syndrome (PMS) genes are most important. SHANK3 has often been touted as the gene that causes PMS, but SHANK3 rarely operates on its own and in some people has nothing to do with PMS (those with interstitial deletions). We learned that large studies of human populations identify 18 PMS genes that are impacted by “natural selection”. Loss of these genes are highly likely to cause problems, the problems that add up to PMS. The genes are:
The big question is, what do we know about these genes? That is, how might they be contributing to PMS? This blog is based on a paper that not only identified these genes, but also pulled together what is currently known about each. Although the paper describes each gene’s function in detail (for well-characterized genes), it also classifies genes into groups. Those groups are quite informative and help us understand why PMS has certain characteristics.
Genes that impact brain development
It is now very clear why PMS can occur with or without SHANK3. Of the 18 PMS genes that are likely to have a high impact on PMS, at least 7 impact brain development: SHANK3, MAPK8IP2, PLXNB2, BRD1, CELSR1, SULT4A1, TCF20.
PLXNB2 regulates the growth of neurons, especially early in development. PMS is a neurodevelopmental disorder, so nothing could be more important than regulating neuron growth.
CELSR1 is also crucial for neurodevelopment. Neurons are exquisitely organized into nuclei in the deep structures of the brain and into very precise layering in the cortex. For example, pyramidal neurons of the cortex are located only in certain layers of the cortex, with the dendrites reaching upwards and the axon pointing down. The axon often winds its way towards the white matter. CELSR1 is important for orchestrating the orientation of neurons to assure proper organization.
BRD1 regulates hundreds of other genes during development. It is highly associated with schizophrenia, as well as PMS. PMS individuals with terminal deletions greater than 1 Mb are missing BRD1 and the loss of BRD1 impacts the entire genome (see https://pubmed.ncbi.nlm.nih.gov/33407854/).
SULT4A1 was recently shown to impact the mitochondria in the brain (see New science: SULT4A1, oxidative stress and mitochondria disorder). Mitochondria convert food into energy for cells. Dysfunction of mitochondria explains why deletions that disrupt SULT4A1 can have a severe impact on neurodevelopment and adult brain function.
TCF20 is a very important gene for brain function. It can cause intellectual disability and other problems on its own. It probably explains why very large deletions in PMS can be more devastating than smaller deletions (see TCF20 may explain why some big deletions are worse than others).
Genes associated with sleep
There are three genes that have close association with sleep or sleep disturbance. SHANK3 impacts sleep in some individuals with PMS, but PIM3 (see this paper) and PRR5 (see this paper) have been identified in studies that explore which genes regulate circadian rhythms (so called, “clock” genes).
Gene associated with lymphedema
CELSR1, the gene important for proper orientation of cells during neurodevelopment, is also associated with inherited lymphedema. Presumably CELSR1 influences cell orientation and the structure in the lymph system during development.
Genes that have unknown function
We must recognize that just because a gene has never been closely studied, that does not mean it is unimportant. In fact, one genomic study makes a convincing argument that genes of unknown function are just as important as the well-characterized genes. PMS has 7 genes likely to be important, yet not well-studied: TRABD, ZBED4, SMC1B, PHF21B, SCUBE1, SREBF2, and XRCC6. The first two genes, TRABD and ZBED4, are of very special concern. One copy of each gene is missing in over 95% of individuals with terminal deletions. It is imperative we find out what these genes are doing and how loss impacts PMS.
Conclusion
This study of PMS genes was a critical step forward in understanding PMS. It provided a short list of culprits. It explains why interstitial deletions cause PMS and it identifies where our research efforts need to be focused. Most importantly, it provides new targets for therapeutics. Unfortunately, a lot of time has gone by without any serious effort to encourage research into the full array of PMS genes. The genes listed above warrant much more study. As a parent of a child with PMS, I strongly feel we should make an effort to encourage the scientists who study these genes. It is hard to understand why so many important genes of PMS are being ignored by the PMS community.
There have been a number of press releases and feel-good articles circulating among my 22q13 deletion syndrome Facebook friends celebrating the advancement of mouse models of “Phelan McDermid syndrome”. I am all for enthusiasm! However, cheering is more fun if you know how the game is played and what to cheer for. With that in mind, I would like to look carefully at models. What is a good model and what might it tell us?
I started out in engineering, where modeling is very important. Let’s imagine modeling an airplane. The first issue of modeling is, what aspects of the plane are motivating us to build a model? In this case we might want to learn about how the airplane will perform if the tail is damaged in flight. We can construct a scaled-down miniature with wings, tail, etc. and omit the inside furnishing (cockpit, storage compartments, etc.). If the miniature has the exact same shape and moving wing/tail parts as the real airplane, we would say our model has construct validity. That is, it is constructed in a way that reflects the original plane. The next step is to put our model into a wind tunnel and see if it flies. The model is held on a wire. We can adjust the wing flaps and other control surfaces. If the model tries to rise at the same wind speed as the real plane, and tries to bank left or right with the same amount of wing flap adjustment as the real plane, we can say this model has face validity. That is, with regard to what we are testing, the model behaves much like the real plane. We are finally ready to benefit from all our work. Let’s take the model for a test flight and then poke a hole in the tail. What settings of the flaps and other control surfaces allow us to keep control of the plane? We may struggle with this for weeks or months hoping we can learn to control our crippled model plane. If we find a solution, maybe we have found a way for pilots to rescue their plane in the event of a similar emergency. If this scheme works, our model has predictive validity. Thus, we measure a model’s worth by:
does it model what we want?
is it constructed in a way that tests what is important to us? (construct validity)
does it perform in a way that mimics what we already know? (face validity)
will model manipulations tell us how the real thing will respond? (predictive validity)
I hope you are getting a picture of what an animal model should do. Let’s look at a mouse model of “Phelan McDermid syndrome” or “PMS”. I use quotes because different scientists have different definitions of “PMS”. See Introduction. For this blog, I will omit the quotes, but remember that there are numerous definitions floating around.
The definition of PMS is important for modeling. The definition of PMS tells us what people claim to be modeling. Some scientists define PMS as a deletion or mutation on chromosome 22 that involves SHANK3. That is fine with me, but that omits the rest of 22q13 deletion syndrome, since there are numerous cases of “interstitial deletions” that don’t affect SHANK3. So, SHANK3 might not be a good choice of model for many families. It depends on the deletion and it depends on what is causing your child the greatest difficulties. There are currently mouse models for 11 different PMS genes for deletions of 1 Mbase or larger. Every gene on this list is relevant to 95% of patients (See Understanding deletion size.) The 11 genes are BRD1, CHKB, CPT1B, MAPK8IP2, MAPK11, MAPK12, NCAPH2, PANX2, PIM3, SHANK3, TYMP. Most of these mouse models are very well studied. If you go further up the chromosome you find other well studied genes with mouse models, like ATXN10. So, the choice of gene is all about what aspect of a disease or syndrome you wish to study.
SHANK3 is popular not because of PMS. It is popular because is has been associated with autism. There are under 1,000 identified people in the USA with PMS, but there are an estimated 36,500 children born each year in the USA with autism. Parents of 22q13 deletion syndrome children should appreciate that researchers who study Shank3 mice are tapping into the national (and world) autism crisis. Our children are a convenient source of subjects, which is why the big national study officially excludes PMS families with interstitial deletions that do not affect SHANK3. Children with SHANK3 mutations are of greatest interest even though they technically do not have 22q13 deletion syndrome (that is why the name PMS was created). Note that only 1/3 of children with PMS have autism, so patients with SHANK3 mutation and autism are the most valuable research subjects.
Although SHANK3 is of great current interest, autism is caused by hundreds of genes. Most parents don’t realize that many 22q13 genes are autism-related or suspected to contribute to autism. Some of the autism genes on chromosome 22 are BRD1, CELSR1, CHKB, MAPK12, PANX2, BRD1. Further up the chromosome (associated with somewhat larger deletions) you can find CELSR1, WNT7B, TCF20, EP300 and others.
Now we understand that Shank3 mice need to be models of something. From the above lists of genes, it is pretty clear that a mouse missing only one gene is not a universal models of either 22q13 deletion syndrome or autism. Both conditions involve a large number of genes. The Shank3 mice are single gene models. The mice are fashioned after super-rare cases of people with specific SHANK3 mutations. Let’s see how these models stack up in terms of construct, face and predictive validities.
All of the published papers so far describe Shank3 mutations and microdeletions, not deletion. I am not going to cite the specific papers here. I have written brief reviews on most of these papers. Contact me if you would like specifics. Some of the mouse Shank3 models use deletions that reflect mutations found in actual patients. For those specific patients (often just one or two), the genetic manipulations have construct validity. That is, the mouse gene has been changed in a way very similar to the human gene. As for the rest of us PMS parents, 95% of our children are missing the SHANK3 gene altogether, along with 30 to 100 more genes. So, these mice do not have very strong construct validity for our children. It is well known that mutations can be very different from deletions. (see Gene deletion versus mutation.) Of course, mutating a single gene may not be very helpful to understanding your child when so many other important genes may be involved. (see How do we know which genes are important?) There is also the problem of using a mouse to model a human. The gene is mutated in an animal that lacks brain areas that are crucial to human behavior, like the granular prefrontal cortex. If these brain areas are important to autism, then the construct validity is weaker.
To be fair, a lot of work goes into to creating a knockout mouse. I don’t have first-hand experience, but I work down the hall from a colleague who is an expert. He works hard and I can read the frustration in his brow on tough days. Just making a mouse is not good enough, you have to prove you have modified the right gene in the right place without messing up the rest of the genome. Then you cross-breed, back breed and then do more validations. My hat is off to those people who make a living this way. When done, the mouse has construct validity in that the targeted gene has been modified.
Face validity is a huge problem with mouse models. Our model airplane ascends and descends, banks and aerodynamically behaves much like its real-life big brother. We know the flight behaviors that are important and we can directly (although not perfectly) compare our model plane to the real thing. How do we compare our model disease to the real disease? Generally, the first step is to compare normal mice to our genetically modified mouse. We note what is different between the two. Then, we compare normal human subjects (“developmentally typical”) to those with the syndrome or disorder. The question then becomes, do the mouse differences seem to reflect the human differences? Mouse models of kidney function and cancer have been very successful with face validity. Urine output and tumor size are easier to measure than social behavior and eye contact in mice. Biopsies of human kidney and many tumors are also much easier than brain biopsies. It is no surprise, then, that mouse models of neuropsychiatric disorders are hard to validate.
The differences between normal and mutated mice can be observed in brain structure, chemical signatures, cellular changes and gene expression. For the most part, there is very little human data for comparison. There are studies with human post mortem tissue that can be helpful, but most of that tissue is from normal human brain. As you might guess, screening for gene expression and other changes in a donated brain from a PMS patient will have issues. Most patients have too many genes involved. One helpful approach is induced pluripotential stem cells (iPScs), grown from skin samples or hair follicles. One might even be fortunate enough to find a donor who’s genetic mutation matches the mouse (or vise versa). However, iPScs do not undergo normal development, so developmental studies are impossible at the moment. They also don’t produce circuits like an intact brain. These are all limitations that impact face validity.
The one common feature you can examine between mice and humans is behavior. There are two extreme viewpoints in this regard. At one extreme, some scientists argue that any behavioral difference between the normal mouse and the genetically modified mouse is “autism-associated” because of the construct validity. At the other extreme, a few scientists argue that the mouse behavioral difference should reflect the standard manual that defines neuropsychiatric disease (DSM-5). The remaining scientists are in the middle until it comes to publishing a paper. Then, they make the argument that their animals’ behavior somehow reflects the behavior of a person with autism. It is my observation that the more popular or prestigious the journal, the more their mouse sounds like a miniature human.
A major problem with Shank3 mice is that, almost universally, they don’t have a significant aberrant behavior. How can you study behavior when it seems normal? There are two solutions, but both have problems. With one approach, researchers find a mutation spot on the gene that is extra good at producing a pathological behavior (e.g., ignoring other mice, not learning, rubbing its fur until the skin is damaged). Another approach is to mutate both copies of the gene. Each of these solutions creates a new problem. In the first case, we are no longer modeling PMS or autism, because this behavior only shows up in very specific mutations. In the second case, we have lost construct validity. We were never trying to model mutation or loss of both copies of SHANK3. That would likely be a different syndrome (just as mutation is different from deletion).
In the end, current models of SHANK3 mutation (autism and SHANK3 mutation aspect of PMS) provide information about molecules and neurons, but are very limited models of the human diseases. They don’t necessarily say much about 22q13 deletion syndrome in general. It is interesting to note that mouse models of other genes of 22q13 deletion syndrome may be more promising. Several mouse models have clear consequences with only one gene mutated or deleted. For example, the Cpt1b mouse model is used to study cold tolerance and insulin sensitivity, and the autism and schizophrenia-associated gene, BRD1, is under intense research right now by a group in Denmark. In both cases the behavioral effects are clear when only one copy of the gene is affected.
We are not done yet, because the most important and most difficult type of validity is predictive validity. That is, does your mouse model tell you what will happen in your human subjects? Given all the problems with making a mouse model of a neuropsychiatric disorder, it is no surprise that the models have not lead to translational (treatment) gains. (See Hope for autism treatment dims as more drug trials fail).
I have heard many people in the research field try to sound hopeful to parents. I believe that hope, when anchored in reality, is very helpful and important. But, too many people have alternative reasons for fanning the flames of hope by knowingly or unknowingly misrepresenting the size of strides in research. My recent blog (How can the same deletion have such different consequences?) touches on why so many parents are mislead by researchers, writers and support organizations. The subsequent blog (22q13 deletion syndrome and science leadership) explains that it does not have to be this way if we bite the bullet and hire qualified leaders.
As a parent, my heartstrings feel the tug of hope every day. The urge to follow those feelings is managed by having a deeper understanding of the research world, and remembering that too many people are eager to give those strings an extra tug.
arm22q13 