UA-45667900-1
Showing posts with label WES. Show all posts
Showing posts with label WES. Show all posts

Thursday 20 July 2023

Genetic testing results


Click on the picture above to read about the upcoming event in London. There are familiar faces appearing, like Agnieszka, Dr Boles and indeed me.

 


I am quite often sent genetic testing results. There are many types of tests ranging from inexpensive tests looking at SNPs to the expensive WES or WGS tests.

SNP = Snip = Single Nucleotide Polymorphism = a tiny genetic spelling mistake

WES = Whole Exome Sequencing

WGS = Whole Genome Sequencing

There is a small industry based around selling expensive supplements for SNPs.

We all carry thousands of SNPs and I think these tests may often raise issues that are not causal.  The results from WGS or WES can be much more insightful.  A good example being in the comment recently posted on this blog.

 

I've been following your blog for many years, it's a real blessing and the perfect place to come and read for us, parents of ASD kids. My boy, 9, has non-regressive autism, is largely non verbal (one word sentence) and has pronounced OCD symptoms (similar to excoriating disorder, but aimed at the environment), hyperactivity and severe gut problems, recurrent vomitting, gastroparesis, etc. The only thing that visibly stopped the hyperactivity and inappropriate laughing and helped him sit for longer periods of time and read his books or watch whole movies was 0.5/kg mg Naltrexone daily, as advised by this paper https://pubmed.ncbi.nlm.nih.gov/16735648/. Lower doses saw the OCD creep back. As for his WGS test results, I've found relevant the fact that he has four pathogenic mutations in the EIF4EBP1, also a de novo mutation in the PIK3R1 gene and multiple other mutations in the STAT3, HTR3a, MAPT and also HLA-DRB1, HLA-DQA1, HLA-A, HLA-B, HLA-C, NRG1, NRG2, SCN4a, CACNA1S genes, amongst many others. We recently tried a course of Azythromycin for immuno-modulation, which saw his OCD reduced further, also his academic interest and focus increased visibly. He responds very well to Ibuprofen, AlkaSeltzer gold, Propranolol, Sytrinol and Cromolyn, but a quite long trial of Bumetanide two years ago did nothing for him. After all trials of various protocols and individual drugs, his gut is still bad, very often food seems to have major difficulty to pass though his digestive tract, no matter how finely tuned his diet is or how many prokinetics he takes. Given your extensive knowledge, I've always wondered what your take on the underlying problem/genetic pathway might be in his case (microglial activation, MTOR activation, perhaps?) and what drugs/cocktail of drugs might work best for his specific genetics and symptoms. He is a smart boy, has self-taught reading, loves music and masters his iPAD like a pro and, unlike what we know about autism, loves being around people. I cannot give up on him. We live in the UK, not the best place to even talk about treatments for autism. Please, if it's not too much to ask, tell me what other medications you thing it might boost his cognition further and help him start talking and develop more skills. Sorry for the long post. And thank you for any advice and ideas you might have to offer.

 

It would be useful to know which of the above mutations are present in at least one of the parents.  There so many possibly causal mutations here; I expect some are actually not relevant. In other words, it is not as scary at it may appear to be.

I do like to start with the easy part, which will be the ion channels.  Dysfunctions in ion channels (channelopathies) are often treatable with existing drugs and there is a great deal of information on each one.

 

CACNA1S

This gene encodes the calcium channel Cav1.1.

This is known as an L type calcium channel, the other ones being Cav1.2 and Cav1,3 and Cav1.4.

These ion channels are extremely important to how your brain works.  Because they also play a role in how your heart works, numerous drugs have been developed, some are more specific to one type of channel (Amlodipine for Cav1.3, Verapamil for Cav1.2).

The individual channels interact with other sub-types, so a mutation in one sub-type can affect other subtypes.

Very interesting in this case are the GI problems. There were efforts made a few years ago to develop R-verapamil as a drug to treat IBS/IBD under the name of Rezular. Some readers of this blog have reported that the only thing that resolves their child’s GI problems is an L-type calcium channel blocker.

Note Memantine, which is an Alzheimer’s drug that was subject to a very large autism clinical trial in the US.  The trial was deemed a failure, but one reader told me that Memantine is the only drug she had found that solved her child’s GI problems.  Memantine has several different modes of action, and a little reported one is blocking L-type calcium channels.

 

https://www.mdpi.com/1648-9144/49/9/64

Conclusions. Our results suggest that the neuroprotective effect of memantine could arise not only through the inhibition of the NMDA receptor current but also through the suppression of the L-type Ca2+ current.   

 

You might expect/hope a geneticist would suggest treatment with a drug like Verapamil.

  

SCN4a

This gene encodes the sodium ion channel Nav1.4.

This is one of the genes associated with Hypokalemic Periodic Paralysis (HPP), that was covered extensively in this blog. Interestingly the above Cav1.1 is also associated with Hypokalemic Periodic Paralysis (HPP).

The other genetic cause of HPP is KCNJ2 (an inward-rectifier potassium channel Kir2.1).

The immediate recovery therapy is drinking a potassium supplement.

A common preventative measure is acetazolamide (Diamox). This drug has also been covered in previous posts. The proposed mechanism is that it “increases the flow of potassium” – not sure what that is supposed to mean.

Some common anti-epilepsy drugs block Nav1.4 (Lamotrigine, Phenytoin etc).

All of the above-mentioned drugs have been used in autism. In specific cases they have shown a benefit.

You could ask your doctor to cautiously try them one by one.

Interestingly, the drug that seems to help many with sound sensitivity is Ponstan.  This cheap drug that affects the flow of potassium ions was proposed by Knut Witkowski as a therapy for 2-3 year olds to prevent non-verbal severe autism. 

 

EIF4EBP1

Here you mention there are 4 pathogenic mutations.

This gene is a real mouthful, but regular reader might recall the odd looking eIF4E part appearing in some previous posts

“This gene encodes one member of a family of translation repressor proteins. The protein directly interacts with eukaryotic translation initiation factor 4E (eIF4E), which is a limiting component of the multi subunit complex that recruits 40S ribosomal subunits to the 5' end of mRNAs. Interaction of this protein with eIF4E inhibits complex assembly and represses translation. This protein is phosphorylated in response to various signals including UV irradiation and insulin signaling, resulting in its dissociation from eIF4E and activation of cap-dependent mRNA translation.”

eIF4E inhibitors for Autism – Why not Ribavirin?

 

As you can see in the above post there are numerous ways to block elF4E. It is possible that the 4 mutations in your gene EIF4EBP1 could have the reverse effect in which case you would want to activate elF4E, not block it.

On the list, in my post above, is quercetin which is OTC and simple to try.

 

PIK3R1

A mutation in this gene can alter the PI3K/AKT/mTOR signaling pathway.

If this gene is causing a problem you might see some facial features a triangular face, a prominent forehead, small chin with a dimple, a loss of fat under the skin, prominent ears, hearing loss and delayed speech.

A mutation in this gene can lead to SHORT syndrome, which hopefully your pediatrician will have heard of.

 https://rarediseases.info.nih.gov/diseases/7633/short-syndrome

 

STAT3

STAT3 plays a key role in the immune system and elsewhere.

You can either have too much or too little STAT3.

In lay terms the immune system might end up either over-activated (hence benefiting from Ibuprofen and Cromolyn sodium) or under activated.

The immunomodulatory probiotics prescribed by gastroenterologists might be worth a try.

Lactobacillus rhamnosus GG

Lactobacillus plantarum 299v 

 

This might well reduce GI problems as well.

  

HTR3a 

This gene encodes subunit A of the type 3 serotonin receptor. It has lots of effects, but it may contribute to the vomiting.

It is associated with:

  • Motion sickness
  • Irritable bowel syndrome
  • Social phobia
  • Serotonin syndrome

For gastroparesis (impaired stomach's motility) the good drug seems to be Domperidone, which you should be able to get for free from your NHS doctor.

Another very popular therapy for gut dysbiosis of all kinds in some countries, but not the UK, is sodium butyrate. This has been mentioned in previous posts. It is an OTC supplement that will produce butyric acid in the gut and it helps restore a healthy mucosa. If you eat lots of fiber and have a healthy microbiome you would produce butyric acid naturally. The cheapest place in Europe to buy it is Poland, where they sell a product called Intesta Max (a weaker version is Intesta).  In the UK it is 3 times more expensive. Making friends with a Pole will save you money.

 

MAPT

The MAPT gene makes tau proteins.  There is a class of disease called tauopathy.

Tau Reduction Prevents Key Features of Autism in Mouse Models

 

Tau: A Novel Entry Point for mTOR-Based Treatments in Autism Spectrum Disorder?

 

As with the PIK3R1 mutation this will lead you to the idea of targeting mTOR signalling. You can inhibit this with Rapamycin, which has been used in autism.

 

Rapamycin/Sirolimus Improves the Behavior of an 8-Year-Old Boy With Nonsyndromic Autism Spectrum Disorder

 

One UK reader did get Everolimus prescribed on the NHS, but that was because the child was diagnosed with a genetic disorder called TSC. Several readers of this blog have tried Rapamycin as used in the Chinese case study.

If you do not have an over activated immune system, Rapamycin will cause the problem of an underactive immune system.

  

HLA-DRB1, HLA-DQA1, HLA-A, HLA-B, HLA-C,

 These genes all play a role in the immune system.

The human leukocyte antigen (HLA) system is a complex of genes in humans which encode cell-surface proteins responsible for regulation of the immune system.

The immune system uses the HLAs to differentiate self cells and non-self cells. Any cell displaying that person's HLA type belongs to that person and is therefore not an invader.

 

HLA Immune Function Genes in Autism

The human leukocyte antigen (HLA) genes on chromosome 6 are instrumental in many innate and adaptive immune responses. The HLA genes/haplotypes can also be involved in immune dysfunction and autoimmune diseases. It is now becoming apparent that many of the non-antigen-presenting HLA genes make significant contributions to autoimmune diseases. Interestingly, it has been reported that autism subjects often have associations with HLA genes/haplotypes, suggesting an underlying dysregulation of the immune system mediated by HLA genes. Genetic studies have only succeeded in identifying autism-causing genes in a small number of subjects suggesting that the genome has not been adequately interrogated. Close examination of the HLA region in autism has been relatively ignored, largely due to extraordinary genetic complexity. It is our proposition that genetic polymorphisms in the HLA region, especially in the non-antigen-presenting regions, may be important in the etiology of autism in certain subjects.

One specific HLA gene has been studied in autism.

 Inheritance of HLA-Cw7 Associated With Autism Spectrum Disorder (ASD)

Autism spectrum disorder (ASD) is a behaviorally defined disorder that is now thought to affect approximately 1 in 69 children in the United States. In most cases, the etiology is unknown, but several studies point to the interaction of genetic predisposition with environmental factors. The immune system is thought to have a causative role in ASD, and specific studies have implicated T lymphocytes, monocytes, natural killer (NK) cells, and certain cytokines. The human leukocyte antigen (HLA) system is involved in the underlying process for shaping an individual’s immune system, and specific HLA alleles are associated with specific diseases as risk factors. In this study, we determine whether a specific HLA allele was associated with ASD in a large cohort of patients with ASD. Identifying such an association could help in the identification of immune system components which may have a causative role in specific cohorts of patients with ASD who share similar specific clinical features. Specimens from 143 patients with ASD were analyzed with respect to race and ethnicity. Overall, HLA-Cw7 was present in a much greater frequency than expected in individuals with ASD as compared to the general population. Further, the cohort of patients who express HLA-Cw7 shares specific immune system/inflammatory clinical features including being more likely to have allergies, food intolerances, and chronic sinusitis as compared to those with ASD who did not express HLA-Cw7. HLA-Cw7 has a role in stimulating NK cells. Thus, this finding may indicate that chronic over-activation of NK cells may have a role in the manifestation of ASD in a cohort of patients with increased immune system/inflammatory features.

 

The therapeutic implication would be to look at immunomodulatory therapy.

At the simple level you have NSAIDs like Ibuprofen, but then you have the more potent drugs used to treat psoriasis, arthritis, IBD etc.

If you saw Dr Arthur Krigsman, the autism gastroenterologist, I guess he would prescribe Humira.  This is an injection you take every few weeks.  That very well might help your son in many ways. He does also come to Europe for consultations. You would need a colonoscopy.

Some British parents take their autistic kids with GI problems to Italy for treatment. You could ask the Thinking Autism charity who they go to see. One of these doctors presented at their conference in London in 2019.  He used some of Krigsman’s slides in his presentation.

 

NRG1, NRG2

Neuregulin 1 and 2 are implicated in brain disorders. NRG1 is well known as a schizophrenia gene, but it has been shown to be miss-expressed in autism as well.

NRG2 also plays a role in many neurological conditions.  

Neuregulins in Neurodegenerative Diseases 

The downstream effect of NRG1 is on epidermal growth factor (EGF). There are expensive cancer drugs like Lapatinib that are inhibitors of EGFR. 

As I have written in my blog, disturbed growth factors is a recurring feature of autism. This is why son many autism genes are also cancer genes. Don’t worry, this does not mean everyone with autism is going to get cancer.

 

Conclusion

Try and find a doctor who is interested to treat your son.

I think you will make great strides by treating the GI problems that you see every day.

I did meet an UK autism mother at that conference in London in 2019 who was told by her doctor that her son’s GI problems would not be treated in the UK and she should look abroad. She went to Italy and solved his problems.  It sounds so bizarre, I would not have believed it to be possible, had I not been talking directly to the mother.  I did talk to the Italian gastroenterologist at that same event.  Contact Thinking Autism and ask who was the Italian who presented in 2019.




Wednesday 2 February 2022

Genetic Mutations vs Differentially Expressed Genes (DEGs) in Autism

 

Genes make proteins and you need the right amount in the right place
at the right time.

I should start this post by confessing to not having carried out genetic testing on Monty, now aged 18 with autism.  When I did mention this to one autism doctor at a conference, I was surprised by her reply:- “ You did not need to.  Now there’s no point doing it”.

I got lucky and treated at least some of Monty’s Differentially Expressed Genes (DEGs) by approaching the problem from a different direction.

People do often ask me about what diagnostic tests to run and in particular about genetic testing.  In general, people have far too high expectations regarding such tests and assume that there will be definitive answers, leading to effective therapeutic interventions.

I do include an interesting example today where parent power is leading a drive towards an effective therapeutic intervention in one single gene type of autism.  The approach has been to start with the single gene that has the mutation and look downstream at the resulting Differentially Expressed Genes (DEGs). The intervention targets one of the DEGs and not the mutated gene itself.

This is a really important lesson.

It can be possible to repurpose existing drugs to treat DEGs quite cheaply.  Many DEGs encode ion channels and there are very many existing drugs that affect ion channels.

Entirely different types of autism may share some of the same DEGs and so benefit from the same interventions.

 

Genetic Testing 

Genetic testing has not proved to be the holy grail in diagnosing and treating autism, but it remains a worthwhile tool at a population level (i.e. maybe not in your specific case).  What matters most of all are Differentially Expressed Genes (DEGs), which is something different.

A paper was recently published that looked into commercially available genetic testing.  Its conclusion was similar to my belief that you risk getting a “false negative” from these tests, in other words they falsely conclude that there is no genetic basis for the person’s symptoms of autism. 

 

Brief Report: Evaluating the Diagnostic Yield of Commercial Gene Panels in Autism

Autism is a prevalent neurodevelopmental condition, highly heterogenous in both genotype and phenotype. This communication adds to existing discussion of the heterogeneity of clinical sequencing tests, “gene panels”, marketed for application in autism. We evaluate the clinical utility of available gene panels based on existing genetic evidence. We determine that diagnostic yields of these gene panels range from 0.22% to 10.02% and gene selection for the panels is variable in relevance, here measured as percentage overlap with SFARI Gene and ranging from 15.15% to 100%. We conclude that gene panels marketed for use in autism are currently of limited clinical utility, and that sequencing with greater coverage may be more appropriate.

 

To save time and money, the commercial gene panels only test genes that the company defines as autism genes.  There is no approved list of autism genes. 

You have more than 20,000 genes and very many are implicated directly, or indirectly, in autism and its comorbities. To be thorough you need Whole Exome Sequencing (WES), where you check them all.  

There are tiny mutations called SNPs ("snips") which you inherit from your parents; there are more than 300 million known SNPs and most people will carry 4-5 million.  Some SNPs are important but clearly most are not.  Some SNPs are very common and some are very rare. 

Even WES only analyses 2% of your DNA, it does not consider the other 98% which is beyond the exome.  Whole Genome Sequencing (WGS) which looks at 100% of your DNA will be the ideal solution, but at some time in the future.  The interpretation of WES data is often very poor and adding all the extra data from WGS is going to overwhelm most people involved. 

Today we return to the previous theme of treating autism by treating the downstream effects caused by Differentially Expressed Genes (DEGS).

Genetics is very complicated and so people assume that is must be able to provide answers. For a minority of autism current genetics does indeed provide an answer, but for most people it does not.

Early on in this blog I noted so many overlaps between the genes and signaling pathways that drive cancer and autism, that is was clear that to understand autism you probably first have to understand cancer; and who has time to do that!

Some people’s cancer is predictable. Chris Evert, the American former world No. 1 tennis player, announced that she has ovarian cancer.  Her sister had exactly the same cancer.  Examining family history can often yield useful information and it is a lot less expensive that genetic testing.  Most people’s cancer is not so predictable; sure if you expose yourself to known environmental triggers you raise its chances, but much appears to be random.  Cancer, like much autism, is usually a multiple hit process. Multiple events need to occur and you may only need to block one of them to avoid cancer. We saw this with a genetic childhood leukemia that you can prevent with a gut bacteria. 


Learning about Autism from the 3 Steps to Childhood Leukaemia


What is not random in cancer are the Differentially Expressed Genes (DEGs).

We all carry highly beneficial tumor suppressing genes, like the autism/cancer gene PTEN.  You would not want to have a mutation in one of these genes.

What happens in many cancers is that the individual carries two good copies of the gene like PTEN, but the gene is turned off. For example, in many people with prostate cancer, the tumor suppressor gene PTEN is turned off in that specific part of the body.  There is no genetic mutation, but there is a harmful Differentially Expressed Gene (DEG). If you could promptly turn PTEN expression back on, you would suppress the cancer.

Not surprisingly, daily use of drugs that increase PTEN expression is associated with reduced incidence of PTEN associated cancer.  Atorvastatin is one such drug.

 

DEGs are what matter, not simply mutations

 

In many cases genetic mutations are of no clinical relevance, we all carry several on average.  In some cases they are of immediate critical relevance.  In most cases mutations are associated with a chance of something happening, there is no certainty and quite often further hits/events/triggers are required.

A good example is epilepsy. Epilepsy is usually caused by an ion channel dysfunction (sodium, potassium or calcium) that is caused by a defect in the associated gene. Most people are not born with epilepsy, the onset can be many years later.  Some parents of a child with autism/epilepsy carry the same ion channel mutation but remain unaffected. 

 

Follow the DEGs from a known mutation 

There is a vanishingly small amount of intelligent translation of autism science to therapy, or even attempts to do so.  I set out below an example of what can be done.

 

Pitt Hopkins (Haploinsufficiency of TCF4) 

The syndrome is caused by a reduction in Transcription factor 4, due to mutation in the TCF4 gene.  One recently proposed therapy is to repurpose the cheap calcium channel blocker Nicardipine. Follow the rationale below.

 

  means down regulated

↑ means up regulated


1.     Gene/Protein TCF4 (Transcription Factor 4) ↓↓↓↓

2.     Genes SCN10a  ↑↑    KCNQ1 ↑↑

3.     Encoding ion channels  Nav1.8   ↑↑     Kv7.1   ↑↑

4.     Repurpose approved drugs as inhibitors of Kv7.1 and Nav1.8 

5.     High throughput screen (HTS) of 1280 approved drugs.

6.     The HTS delivered 55 inhibitors of Kv7.1 and 93 inhibitors of Nav1.8

7.     Repurposing the Calcium Channel Inhibitor Nicardipine as a Nav1.8 inhibitor 


           

The supporting science: 

Psychiatric Risk Gene Transcription Factor 4 Regulates Intrinsic Excitability of Prefrontal Neurons via Repression of SCN10a and KCNQ1

  

Highlights

•TCF4 loss of function alters the intrinsic excitability of prefrontal neurons 

TCF4-dependent excitability deficits are rescued by SCN10a and KCNQ1 antagonists 

TCF4 represses the expression of SCN10a and KCNQ1 ion channels in central neurons 

•SCN10a is a potential therapeutic target for Pitt-Hopkins syndrome

  

Nav1.8 is a sodium ion channel subtype that in humans is encoded by the SCN10A gene

Kv7.1 (KvLQT1) is a potassium channel protein whose primary subunit in humans is encoded by the KCNQ1 gene.

  

Transcription Factor 4 (TCF4) is a clinically pleiotropic gene associated with schizophrenia and Pitt-Hopkins syndrome (PTHS).  

SNPs in a genomic locus containing TCF4 were among the first to reach genome-wide significance in clinical genome-wide association studies (GWAS) for schizophrenia  These neuropsychiatric disorders are each characterized by prominent cognitive deficits, which suggest not only genetic overlap between these disorders but a potentially overlapping pathophysiology.

We propose that these intrinsic excitability phenotypes may underlie some aspects of pathophysiology observed in PTHS and schizophrenia and identify potential ion channel therapeutic targets.

Given that TCF4 dominant-negative or haploinsufficiency results in PTHS, a syndrome with much more profound neurodevelopmental deficits than those observed in schizophrenia, the mechanism of schizophrenia risk associated with TCF4 is presumably due to less extreme alterations in TCF4 expression at some unknown time point in development

The pathological expression of these peripheral ion channels in the CNS may create a unique opportunity to target these channels with therapeutic agents without producing unwanted off-target effects on normal neuronal physiology, and we speculate that targeting these ion channels may ameliorate cognitive deficits observed in PTHS and potentially schizophrenia.

 

 

Disordered breathing in a Pitt-Hopkins syndrome model involves Phox2b-expressing parafacial neurons and aberrant Nav1.8 expression

Pitt-Hopkins syndrome (PTHS) is a rare autism spectrum-like disorder characterized by intellectual disability, developmental delays, and breathing problems involving episodes of hyperventilation followed by apnea. PTHS is caused by functional haploinsufficiency of the gene encoding transcription factor 4 (Tcf4). Despite the severity of this disease, mechanisms contributing to PTHS behavioral abnormalities are not well understood. Here, we show that a Tcf4 truncation (Tcf4tr/+) mouse model of PTHS exhibits breathing problems similar to PTHS patients. This behavioral deficit is associated with selective loss of putative expiratory parafacial neurons and compromised function of neurons in the retrotrapezoid nucleus that regulate breathing in response to tissue CO2/H+. We also show that central Nav1.8 channels can be targeted pharmacologically to improve respiratory function at the cellular and behavioral levels in Tcf4tr/+ mice, thus establishing Nav1.8 as a high priority target with therapeutic potential in PTHS. 

 

Repurposing Approved Drugs as Inhibitors of Kv7.1 and Nav1.8 To Treat Pitt Hopkins Syndrome

Purpose:

Pitt Hopkins Syndrome (PTHS) is a rare genetic disorder caused by mutations of a specific gene, transcription factor 4 (TCF4), located on chromosome 18. PTHS results in individuals that have moderate to severe intellectual disability, with most exhibiting psychomotor delay. PTHS also exhibits features of autistic spectrum disorders, which are characterized by the impaired ability to communicate and socialize. PTHS is comorbid with a higher prevalence of epileptic seizures which can be present from birth or which commonly develop in childhood. Attenuated or absent TCF4 expression results in increased translation of peripheral ion channels Kv7.1 and Nav1.8 which triggers an increase in after-hyperpolarization and altered firing properties.

Methods:

We now describe a high throughput screen (HTS) of 1280 approved drugs and machine learning models developed from this data. The ion channels were expressed in either CHO (KV7.1) or HEK293 (Nav1.8) cells and the HTS used either 86Rb+ efflux (KV7.1) or a FLIPR assay (Nav1.8).

Results:

The HTS delivered 55 inhibitors of Kv7.1 (4.2% hit rate) and 93 inhibitors of Nav1.8 (7.2% hit rate) at a screening concentration of 10 μM. These datasets also enabled us to generate and validate Bayesian machine learning models for these ion channels. We also describe a structure activity relationship for several dihydropyridine compounds as inhibitors of Nav1.8.

Conclusions:

This work could lead to the potential repurposing of nicardipine or other dihydropyridine calcium channel antagonists as potential treatments for PTHS acting via Nav1.8, as there are currently no approved treatments for this rare disorder.

  

Repurposing the Dihydropyridine Calcium Channel Inhibitor Nicardipine as a Nav1.8 inhibitor in vivo for Pitt Hopkins Syndrome

Individuals with the rare genetic disorder Pitt Hopkins Syndrome (PTHS) do not have sufficient expression of the transcription factor 4 (TCF4) which is located on chromosome 18. TCF4 is a basic helix-loop-helix E protein that is critical for the normal development of the nervous system and the brain in humans. PTHS patients lacking sufficient TCF4 frequently display gastrointestinal issues, intellectual disability and breathing problems. PTHS patients also commonly do not speak and display distinctive facial features and seizures. Recent research has proposed that decreased TCF4 expression can lead to the increased translation of the sodium channel Nav1.8. This in turn results in increased after-hyperpolarization as well as altered firing properties. We have recently identified an FDA approved dihydropyridine calcium antagonist nicardipine used to treat angina, which inhibited Nav1.8 through a drug repurposing screen.

 

All of the above was a parent driven process.  Well done, Audrey!

Questions remain.

Is Nicardipine actually beneficial to people with Pitt Hopkins Syndrome? Does it matter at what age therapy is started? What about the Kv7.1 inhibitor?

 

Conclusion 

Genetics is complicated, ion channel dysfunctions are complicated; but just a superficial understanding can take you a long way to understand autism, epilepsy and many other health issues.

There is a great deal in this blog about channelopathies/ion channel dysfunctions.

https://epiphanyasd.blogspot.com/search/label/Channelopathy

Almost everyone with autism has one or more channelopathies. Most channelopathies are potentially treatable.

Parents of children with rare single gene autisms should get organized and make sure there is basic research into their specific biological condition.  They need to ensure that there is an animal model created and it is then used to screen for existing drugs that may be therapeutic.  I think they also need to advocate for gene therapy to be developed.  This all takes years, but the sooner you start, the sooner you will make an impact.

Very likely, therapies developed for some single gene autisms will be applicable more broadly.  A good example may be the IGF-1 derivative Trofinetide, for girls with Rett Syndrome. IGF-1 (Insulin-like growth factor 1) is an important growth factor that is required for proper brain development. In the brain, IGF-1 is broken down into a protein fragment called glypromate (GPE). Trofinetide is an orally available version of GPE.

The MeCP2 protein controls the expression of several genes, such as Insulin-like Growth Factor 1 (IGF1), brain-derived neurotrophic factor (BDNF) and N-methyl-D-aspartate (NMDA).  All three are implicated in broader autism. 

https://rettsyndromenews.com/trofinetide-nnz-2566/

In girls with Rett Syndrome the genetic mutation is in the gene MeCP2, but one of the key DEGs (differentially expressed genes) is the FXYD1; it is over-expressed. IGF-1 supresses the activity of FXYD1 and hopefully so does Trofinetide.  Not so complicated, after all!

Medicine is often driven by the imperative to do no harm.

In otherwise severely impaired people, perhaps the imperative should be to try and do some good.

In medicine, time is of the essence; doctors in the ER can be heard to say "Stat!", from the Latin word for immediately, statim.  

How about some urgency in translating autism science into therapy? But then, what's the hurry? Why rock the boat?

On an individual basis, much is already possible, but you will have to do most of the work yourself - clearly a step too far for most people.








   



 






Thursday 9 March 2017

Gene Primer for people interested in Autism


Today’s post is a standalone introduction to genetics, as relevant for lay people interested in autism.
The scientists among you will likely know all this and more, but many more people are talking about genes and genetic testing these days.
Genetics is the domain of some very clever scientists and to fully understand this field would require a great deal of effort.  Often the cleverest people are the least able to explain things to the rest of us. Some equally clever researchers think that there will be so many possible autism genes that it is better to focus on the much smaller number of shared affected pathways.  I stand with the latter group.
Confused? He should be.

As is often the case, even a basic understanding of the principles does allow you to draw meaningful practical conclusions from the ever-expanding pool of research.
In most cases of idiopathic autism there is over/under expression of a very large number of genes.  Most of these genes do their job and produce perfectly functional proteins, just too much or too little. If you ran a test to sequence all those genes, most would come up as normal.  A variation in the expression in one gene can affect the expression of numerous others.
Ultimately what matters is, in each part of the body, which genes are turned on when they should be off and vice versa.
Some well-known syndromes result from missing copies, or extra copies, of certain genes.  These are called CNVs (Copy Number Variants). This is relatively easy to validate with existing tests.
The most common type of genetic variation among people is an SNP (single nucleotide polymorphism) and it represents a difference in a single DNA building block, called a nucleotide.  There are online database that log the effect of individual SNPs, which you can look up.
There are 12,932 known SNPs for the CACNA1C gene, which expresses the calcium ion channel Cav1.2. Ion channels are little gates in your cells that open and close as part of the signaling process that controls your body.  When these gates are faulty they might stay open, or stay shut, or you might just have too many of them; and you end up with a problem called a channelopathy. The Cav1.2 ion channel is known  to go wrong autism, bipolar and schizophrenia. If you want to give that gate a nudge to shut it, you use a channel blocker. A complication is that these same gates appear all over your body and a Cav1.2 blocker will affect them all. This is why some people get side effects.
There can be genetic mutations that result in a variation in the structure of the protein that is expressed.  This can be good (protective), or it can be bad.  This does not seem to underlie autism.
The great majority of research concerns the exome, a tiny part of the wider genome; the exome holds the information needed to express proteins.  The remaining 98% was thought to be “DNA junk”, but this appears not to be the case.  Variations in the 98% do matter, because they include things like silencers and enhancers that affect gene expression.  Only recently have scientists paid attention to the 98%; so much remains unknown.
Until recently, scientists thought that human diseases were caused mainly by changes in DNA sequence, infectious agents such as bacteria and viruses, or environmental agents. Now, researchers have demonstrated that changes in the epigenome also can cause, or result from, disease.
The epigenome consists of a record of the chemical markers attached to your DNA, like bookmarks that turn on or turn off particular genes.  These changes can be passed down via transgenerational epigenetic inheritance.  Changes to the epigenome result in changes to the function of the genome. 
Epigenomics/epigenetics, will become a vital part of efforts to better understand the human body and to improve health. Epigenomic maps may someday enable doctors to determine an individual's health status and predict a patient's response to therapies. 
Genetics can be made to sound bewildering complex:-
An analogy to the human genome stored on DNA is that of instructions stored in a book:
  • The book (genome) would contain 23 chapters (chromosomes);
  • Each chapter contains 48 to 250 million letters (A,C,G,T) without spaces;
  • Hence, the book contains over 3.2 billion letters total;
  • The book fits into a cell nucleus the size of a pinpoint;

 Or you can make it quite simple:-

·        The latest Boing 747 has 6,000,000 parts, and I expect many special extra ones in the new Air Force One variant.
·        Less than 25,000 different genes are needed to make a human and many of these appear not be essential.
·       As with jumbo jet you don't just need all the components, but you have to know where to put them and the correct sequence. Is the human equivalent of these instructions in the exome? or elsewhere in the genome?

I prefer simple.

There are different forms of genetic testing; simple ones test for a specific dysfunction, like Fragile-X.  This is relatively straight forward because the lab knows exactly where to look.
When you start looking for unknown dysfunctions the big risk is that you will find very many variances, the great majority of which have no relevance whatsoever.  You need to only consider the variances that are relevant.  Who decides which are relevant?  Beyond the very well-known risk genes, it becomes hugely subjective.
This interpretation is subjective because no one knows for sure the complete list of genes that could relate to autism.  It is likely to be a sub-set of the so-called "essential genes". The Simons Foundation suggest around 700 genes, but the list keeps growing. The AutsimKB database maintained by Peking University currently contains 3075 genes (99 syndromic autism related genes and 3022 non-syndromic autism related genes), 4964 Copy Number Variations (CNVs).
Some people will learn important things about their autism from today’s  genetic testing, but many will not.
Two things need to change, better data and better analysis.
I did check the analysis from one well-known US testing laboratory.  All I did was look at the genes they highlighted as being relevant to autism and then look at the supplementary list of variance present in one test, but that they considered irrelevant to autism.  Just using google I could find evidence that some of those irrelevant genes were actually potentially relevant.
Even if you find you have a flagged autism gene, this may or may not help you. 
If you have a defect in your mitochondrial DNA then you can say “for sure” that you have a particular type of mitochondrial disease and try and optimize your therapy.
If you have a variance in a gene associated with people with autism who remain non-verbal, you might be better off not knowing.  There is a case often quoted of the mother who made great efforts to get her child to talk, only to find later that a “non-verbal” mutation existed.  She might not have bothered had she made the genetic testing at a younger age.  I actually know a child diagnosed with a supposed “non-verbal” mutation and I think it is a really stupid diagnosis.  


A Future World

In a future time you might analyze the genome and epigenome of both parents and the child. This would be automatically be compared with the results from tens of millions of other people.  This would then reliably predict the possible dysfunctions that might develop in the child.  Having predicted the dysfunctions and the probability of them actually occurring,  a list of personalized therapies, some preventative, would be provided.


Conclusion
The science is not quite there yet. There is much work to be done on the exome, the wider genome, let alone the record of all those tags on it, which is the epigenome.
Current genetic testing can confirm known single gene autism syndromes.
Current genetic testing can identify known single gene metabolic and other disorders that can contribute to autism.
For idiopathic autism current genetic testing may, or may not, tell you anything that leads to a useful therapy.  Given the choice, take the best genetic testing available, just realize its limitations.
Now for the science:-


Understanding Genetics
There are some good introductions to genetics that are available for free.
Wikipedia gets quite complicated
This book is available for free:-


  

DNA

DNA is the hereditary material in humans. Nearly every cell in a person’s body has the same DNA. Most DNA is located in the cell nucleus, where it is called nuclear DNA, but a small amount of DNA can also be found in the mitochondria, where it is called mitochondrial DNA. 

The information in DNA is stored as a code made up of four chemical bases:

adenine (A), guanine (G), cytosine (C), and thymine (T).

Human DNA consists of about 3 billion bases, and more than 99 percent of those bases are the same in all people. The order, or sequence, of these bases determines the information available for building and maintaining an organism, similar to the way in which letters of the alphabet appear in a certain order to form words and sentences. 

DNA bases pair up with each other, A with T and C with G, to form units called base pairs.  Each base is also attached to a sugar molecule and a phosphate molecule. Together, a base, sugar, and phosphate are called a nucleotide. 

Nucleotides are arranged in two long strands that form a spiral called a double helix. The structure of the double helix is somewhat like a ladder, with the base pairs forming the ladder’s rungs and the sugar and phosphate molecules forming the vertical sidepieces of the ladder. 

An important property of DNA is that it can replicate, or make copies of itself. 

Each strand of DNA in the double helix can serve as a pattern for duplicating the sequence of bases. This is critical when cells divide because each new cell needs to have an exact copy of the DNA present in the old cell.





Mitochondrial DNA

Although most DNA is packaged in chromosomes within the nucleus, mitochondria also have a small amount of their own DNA. This genetic material is known as mitochondrial DNA. 

Mitochondria are structures within cells that convert the energy from food into a form that cells can use. Each cell contains hundreds to thousands of  mitochondria. 

Mitochondria produce energy through a process called oxidative phosphorylation. This process uses oxygen and simple sugars to create adenosine triphosphate (ATP), the cell’s main energy source. A set of enzyme complexes, designated as complexes I-V, carry out oxidative phosphorylation within mitochondria. 
In addition to energy production, mitochondria play a role in several other cellular activities. For example, mitochondria help regulate the self-destruction of cells (apoptosis). They are also necessary for the production of substances such as  cholesterol and heme (a component of hemoglobin, the molecule that carries oxygen in the blood). 

Mitochondrial DNA contains 37 genes, all of which are essential for normal mitochondrial function. Thirteen of these genes provide instructions for making enzymes involved in oxidative phosphorylation. The remaining genes provide instructions for making molecules called transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs), which are chemical cousins of DNA. These types of RNA help assemble protein building blocks (amino acids) into functioning proteins.


Genes
A gene is the basic physical and functional unit of heredity. Genes, which are made up of DNA, act as instructions to make molecules called proteins. In humans, genes vary in size from a few hundred DNA bases to more than 2 million bases. The Human Genome Project has estimated that humans have between 20,000 and 25,000 genes.

Every person has two copies of each gene, one inherited from each parent.

Most genes are the same in all people, but a small number of genes (less than 1 percent of the total) are slightly different between people.

Alleles are forms of the same gene with small differences in their sequence of DNA bases. These small differences contribute to each person’s unique physical features.

Genes are made up of DNA. Each chromosome contains many genes.

Chromosome

In the nucleus of each cell, the DNA molecule is packaged into thread-like structures called chromosomes. Each chromosome is made up of DNA tightly coiled many times around proteins called histones that support its structure.

Chromosomes are not visible in the cell’s nucleus—not even under a microscope - when the cell is not dividing.
In humans, each cell normally contains 23 pairs of chromosomes, for a total of 46.

We inherit 23 of our chromosomes from our mother (in the egg), and the other 23 from our father (in the sperm), so that we have 23 pairs of chromosomes, and therefore two copies of each gene.

Twenty-two of these pairs, called autosomes, look the same in both males and females. The 22 autosomes are numbered by size.

The 23rd pair, the sex chromosomes, differ between males and females. Females have two copies of the X chromosome, while males have one X and one Y chromosome.

Down syndrome is caused by possessing three copies of chromosome 21 instead of the usual two.

Fragile X syndrome is caused by a defect on chromosome 23, typically due to the expansion of the CGG triplet repeat within the Fragile X mental retardation 1 (FMR1) gene on the X chromosome.

Males with a full mutation display virtually complete penetrance and will therefore almost always display symptoms of FXS, while females with a full mutation generally display a penetrance of about 50% as a result of having a second, normal X chromosome. Females with FXS may have symptoms ranging from mild to severe, although they are generally less affected than males.



Do all gene mutations affect health and development?

No; only a small percentage of mutations cause genetic disorders—most have no impact on health or development. For example, some mutations alter a gene's DNA sequence but do not change the function of the protein made by the gene.
Often, gene mutations that could cause a genetic disorder are repaired by certain enzymes before the gene is expressed and an altered protein is produced.
Each cell has a number of pathways through which enzymes recognize and repair mistakes in DNA. Because DNA can be damaged or mutated in many ways, DNA repair is an important process by which the body protects itself from disease.

A very small percentage of all mutations actually have a positive effect. These mutations lead to new versions of proteins that help an individual better adapt to changes in his or her environment. For example, a beneficial mutation could result in a protein that protects an individual and future generations from a new strain of bacteria.

Because a person's genetic code can have a large number of mutations with no effect on health, diagnosing genetic conditions can be difficult. Sometimes, genes thought to be related to a particular genetic condition have mutations, but whether these changes are involved in development of the condition has not been determined; these genetic changes are known as variants of unknown significance (VOUS). Sometimes, no mutations are found in suspected disease related genes, but mutations are found in other genes whose relationship to a particular genetic condition is unknown. It is difficult to know whether these variants are involved in the disease.

The types of possible gene mutations

The DNA sequence of a gene can be altered in a number of ways. Gene mutations have varying effects on health, depending on where they occur and whether they alter the function of essential proteins. The types of mutations include:


Missense mutation

This type of mutation is a change in one DNA base pair that results in the substitution of one amino acid for another in the protein made by a gene.


Nonsense mutation

A nonsense mutation is also a change in one DNA base pair. Instead of substituting one amino acid for another, however, the altered DNA sequence prematurely signals the cell to stop building a protein. This type of mutation results in a shortened protein that may function improperly or not at all.


Insertion

An insertion changes the number of DNA bases in a gene by adding a piece of DNA. As a result, the protein made by the gene may not function properly.


Deletion

A deletion changes the number of DNA bases by removing a piece of DNA. Small deletions may remove one or a few base pairs within a gene, while larger deletions can remove an entire gene or several neighboring genes.

The deleted DNA may alter the function of the resulting protein(s).


Duplication

A duplication consists of a piece of DNA that is abnormally copied one or more times. This type of mutation may alter the function of the resulting protein.


Frameshift mutation

This type of mutation occurs when the addition or loss of DNA bases changes a gene's reading frame. A reading frame consists of groups of 3 bases that each code for one amino acid. A frameshift mutation shifts the grouping of these bases and changes the code for amino acids.

The resulting protein is usually nonfunctional. Insertions, deletions, and duplications can all be frameshift mutations.


Repeat expansion

Nucleotide repeats are short DNA sequences that are repeated a number of times in a row. For example, a trinucleotide repeat is made up of 3-base-pair sequences, and a tetranucleotide repeat is made up of 4-base-pair sequences. A repeat expansion is a mutation that increases the number of times that the short DNA sequence is repeated. This type of mutation can cause the resulting protein to function improperly.






Change in the number of genes (CNVs)

People have two copies of most genes, one copy inherited from each parent. In some cases, however, the number of copies varies—meaning that a person can be born with one, three, or more copies of particular genes. Less commonly, one or more genes may be entirely missing. This type of genetic difference is known as copy number variation (CNV).

Copy number variation results from insertions, deletions, and duplications of large segments of DNA. These segments are big enough to include whole genes. Variation in gene copy number can influence the activity of genes and ultimately affect many body functions.

Copy number variation accounts for a significant amount of genetic difference between people. More than 10 percent of human DNA appears to contain these differences in gene copy number. While much of this variation does not affect health or development, some differences likely influence a person’s risk of disease and response to certain drugs.

Changes in the number of chromosomes

Human cells normally contain 23 pairs of chromosomes, for a total of 46 chromosomes in each cell. A change in the number of chromosomes can cause problems with growth, development, and function of the body's systems. These changes can occur during the formation of reproductive cells (eggs and sperm), in early fetal development, or in any cell after birth. A gain or loss of chromosomes from the normal 46 is called aneuploidy.

A common form of aneuploidy is trisomy, or the presence of an extra chromosome in cells. People with trisomy have three copies of a particular chromosome in cells instead of the normal two copies.

Down syndrome is an example of a condition caused by trisomy. People with Down syndrome typically have three copies of chromosome 21 in each cell, for a total of 47 chromosomes per cell.

Monosomy, or the loss of one chromosome in cells, is another kind of aneuploidy. People with monosomy have one copy of a particular chromosome in cells instead of the normal two copies. Turner syndrome is a condition caused by monosomy.  Women with Turner syndrome usually have only one copy of the X chromosome in every cell, for a total of 45 chromosomes per cell.

Rarely, some cells end up with complete extra sets of chromosomes. Cells with one additional set of chromosomes, for a total of 69 chromosomes, are called triploid. Cells with two additional sets of chromosomes, for a total of 92 chromosomes, are called tetraploid. A condition in which every cell in the body has an extra set of chromosomes is not compatible with life.


Changes in the structure of chromosomes

This type of change is usually associated with cancer.

Changes that affect the structure of chromosomes can cause problems with growth, development, and function of the body's systems. These changes can affect many genes along the chromosome and disrupt the proteins made from those genes.

Structural changes can occur during the formation of egg or sperm cells, in early fetal development, or in any cell after birth. Pieces of DNA can be rearranged within one chromosome or transferred between two or more chromosomes. The effects of structural changes depend on their size and location, and whether any genetic material is gained or lost. Some changes cause medical problems, while others may have no effect on a person's health.

Changes in chromosome structure include:


Translocations

A translocation occurs when a piece of one chromosome breaks off and attaches to another chromosome. This type of rearrangement is described as balanced if no genetic material is gained or lost in the cell. If there is a gain or loss of genetic material, the translocation is described as unbalanced.


Deletions

Deletions occur when a chromosome breaks and some genetic material is lost. Deletions can be large or small, and can occur anywhere along a chromosome.


Duplications

Duplications occur when part of a chromosome is copied (duplicated) too many times. This type of chromosomal change results in extra copies of genetic material from the duplicated segment.


Inversion

An inversion involves the breakage of a chromosome in two places; the resulting piece of DNA is reversed and re-inserted into the chromosome.
Genetic material may or may not be lost as a result of the chromosome breaks. An inversion that involves the chromosome's constriction point (centromere) is called a pericentric inversion. An inversion that occurs in the long (q) arm or short (p) arm and does not involve the centromere is called a paracentric inversion.





Isochromosomes

An isochromosome is a chromosome with two identical arms. Instead of one long (q) arm and one short (p) arm, an isochromosome has two long arms or two short arms. As a result, these abnormal chromosomes have an extra copy of some genes and are missing copies of other genes.


Dicentric chromosomes

Unlike normal chromosomes, which have a single constriction point (centromere), a dicentric chromosome contains two centromeres. Dicentric chromosomes result from the abnormal fusion of two chromosome pieces, each of which includes a centromere. These structures are unstable and often involve a loss of some genetic material.


Ring chromosomes

Ring chromosomes usually occur when a chromosome breaks in two places and the ends of the chromosome arms fuse together to form a circular structure. The ring may or may not include the chromosome's constriction point (centromere). In many cases, genetic material near the ends of the chromosome is lost.

Many cancer cells also have changes in their chromosome structure. These changes are not inherited; they occur in somatic cells (cells other than eggs or sperm) during the formation or progression of a cancerous tumor.


Changes in mitochondrial DNA

Mitochondria  are structures within cells that convert the energy from food into a form that cells can use. Although most DNA is packaged in chromosomes within the nucleus, mitochondria also have a small amount of their own DNA (known as mitochondrial DNA or mtDNA). In some cases, inherited changes in mitochondrial DNA can cause problems with growth, development, and function of the body’s systems. These mutations disrupt the mitochondria’s ability to generate energy efficiently for the cell.

Conditions caused by mutations in mitochondrial DNA often involve multiple organ systems. The effects of these conditions are most pronounced in organs and tissues that require a lot of energy (such as the heart, brain, and muscles).

Although the health consequences of inherited mitochondrial DNA mutations vary widely, frequently observed features include muscle weakness and wasting, problems with movement, diabetes, kidney failure, heart disease, loss of intellectual functions (dementia), hearing loss, and abnormalities involving the eyes and vision.

Mitochondrial DNA is also prone to somatic mutations, which are not inherited. somatic mutations occur in the DNA of certain cells during a person’s lifetime and typically are not passed to future generations. Because mitochondrial DNA has a limited ability to repair itself when it is damaged, these mutations tend to build up over time. A buildup of somatic mutations in mitochondrial DNA has been associated with some forms of cancer and an increased risk of certain age-related disorders such as heart disease, Alzheimer disease, and Parkinson disease. Additionally, research suggests that the progressive accumulation of these mutations over a person’s lifetime may play a role in the normal process of aging.

What does it mean to have a genetic predisposition to a disease?

A genetic predisposition (sometimes also called genetic susceptibility) is an increased likelihood of developing a particular disease based on a person's genetic makeup. A genetic predisposition results from specific genetic variations that are often inherited from a parent. These genetic changes contribute to the development of a disease but do not directly cause it. Some people with a predisposing genetic variation will never get the disease while others will, even within the same family.

Genetic variations can have large or small effects on the likelihood of developing a particular disease. For example, certain mutations in the BRCA1 or BRCA2 genes greatly increase a person's risk of developing breast cancer and ovarian cancer. Variations in other genes, such as BARD1 and BRIP1, also increase breast cancer risk, but the contribution of these genetic changes to a person's overall risk appears to be much smaller.

Current research is focused on identifying genetic changes that have a small effect on disease risk but are common in the general population. Although each of these variations only slightly increases a person's risk, having changes in several different genes may combine to increase disease risk significantly.

Changes in many genes, each with a small effect, may underlie susceptibility to many common diseases, including cancer, obesity, diabetes, heart disease, and mental illness.

In people with a genetic predisposition, the risk of disease can depend on multiple factors in addition to an identified genetic change. These include other genetic factors (sometimes called modifiers) as well as lifestyle and environmental factors.  Although a person's genetic makeup cannot be altered, some lifestyle and environmental  modifications (such as having more frequent disease screenings and maintaining a healthy weight) may be able to reduce disease risk in people with a genetic predisposition.


Genes direct the production of proteins

Most genes contain the information needed to make functional molecules called proteins. (A few genes produce other molecules that help the cell assemble proteins.)

The journey from gene to protein is complex and tightly controlled within each cell. It consists of two major steps: transcription and translation. Together, transcription and translation are known as gene expression. During the process of transcription, the information stored in a gene's DNA is transferred to a similar molecule called RNA (ribonucleic acid) in the cell nucleus.

Both RNA and DNA are made up of a chain of nucleotide bases, but they have slightly different chemical properties. The type of RNA that contains the information for making a protein is called messenger RNA (mRNA) because it carries the information, or message, from the DNA out of the nucleus into the cytoplasm.

Translation, the second step in getting from a gene to a protein, takes place in the cytoplasm. The mRNA interacts with a specialized complex called a ribosome, which "reads" the sequence of mRNA bases. Each sequence of three bases, called a codon, usually codes for one particular amino acid. (Amino acids are the building blocks of proteins.) A type of RNA called transfer RNA (tRNA) assembles the protein, one amino acid at a time. Protein assembly continues until the ribosome encounters a “stop” codon (a sequence of three bases that does not code for an amino acid).
The flow of information from DNA to RNA to proteins is one of the fundamental principles of molecular biology. It is so important that it is sometimes called the "central dogma.”


Genes turn on and off in cells

Each cell expresses, or turns on, only a fraction of its genes. The rest of the genes are repressed, or turned off. The process of turning genes on and off is known as gene regulation. Gene regulation is an important part of normal development. Genes are turned on and off in different patterns during development to make a brain cell look and act different from a liver cell or a muscle cell, for example. Gene regulation also allows cells to react quickly to changes in their environments. Although we know that the regulation of genes is critical for life, this complex process is not yet fully understood.

Gene regulation can occur at any point during gene expression, but most commonly occurs at the level of transcription (when the information in a gene’s DNA is transferred to mRNA). Signals from the environment or from other cells activate proteins called transcription factors. These proteins bind to regulatory regions of a gene and increase or decrease the level of transcription. By controlling the level of transcription, this process can determine the amount of protein product that is made by a gene at any given time.


The epigenome

DNA modifications that do not change the DNA sequence can affect gene activity. Chemical compounds that are added to single genes can regulate their activity; these modifications are known as epigenetic changes. The epigenome comprises all of the chemical compounds that have been added to the entirety of one’s DNA (genome) as a way to regulate the activity (expression) of all the genes within the genome. The chemical compounds of the epigenome are not part of the DNA sequence, but are on or attached to DNA (“epi-“ means above in Greek). Epigenomic modifications remain as cells divide and in some cases can be inherited through the generations. Environmental influences, such as a person’s diet and exposure to pollutants, can also impact the epigenome.

Epigenetic changes can help determine whether genes are turned on or off and can influence the production of proteins in certain cells, ensuring that only necessary proteins are produced. For example, proteins that promote bone growth are not produced in muscle cells. Patterns of epigenome modification vary among individuals, different tissues within an individual, and even different cells.

A common type of epigenomic modification is called methylation. Methylation involves attaching small molecules called methyl groups, each consisting of one
carbon atom and three hydrogen atoms, to segments of DNA. When methyl groups are added to a particular gene, that gene is turned off or silenced, and no protein is produced from that gene.

Because errors in the epigenetic process, such as modifying the wrong gene or failing to add a compound to a gene, can lead to abnormal gene activity or inactivity, they can cause genetic disorders. Conditions including cancers, metabolic disorders, and degenerative disorders have all been found to be related to epigenetic errors.

Scientists continue to explore the relationship between the genome and the
chemical compounds that modify it. In particular, they are studying what effect
the modifications have on gene function, protein production, and human health.




How geneticists indicate the location of a gene
Geneticists use maps to describe the location of a particular gene on a chromosome. One type of map uses the cytogenetic location to describe a gene’s position. The cytogenetic location is based on a distinctive pattern of bands created when chromosomes are stained with certain chemicals. Another type of map uses the molecular location, a precise description of a gene's position on a chromosome. The molecular location is based on the sequence of DNA building blocks (base pairs) that make up the chromosome.


Cytogenetic location

Geneticists use a standardized way of describing a gene's cytogenetic location.
In most cases, the location describes the position of a particular band on a stained chromosome:

        17q12

It can also be written as a range of bands, if less is known about the exact location:

        17q12-q21

The combination of numbers and letters provide a gene's “address” on a chromosome. This address is made up of several parts:


  • The chromosome on which the gene can be found. The first number or letter used to describe a gene's location represents the chromosome. Chromosomes 1 through 22 (the autosomes) are designated by their chromosome number. The sex chromosomes are designated by X or Y.
  • The arm of the chromosome. Each chromosome is divided into two sections (arms) based on the location of a narrowing (constriction) called the centromere. By convention, the shorter arm is called p, and the longer arm is called q. The chromosome arm is the second part of the gene's address. For example, 5q is the long arm of chromosome 5, and Xp is the short arm of the X chromosome.
  • The position of the gene on the p or q arm. The position of a gene is based on a distinctive pattern of light and dark bands that appear when the chromosome is stained in a certain way. The position is usually designated by two digits (representing a region and a band), which are sometimes followed by a decimal point and one or more additional digits (representing sub-bands within a light or dark area). The number indicating the gene position increases with distance from  the centromere. For example: 14q21 represents position 21 on the long arm of chromosome 14. 14q21 is closer to the centromere than 14q22. Sometimes, the abbreviations “cen” or “ter” are also used to describe a gene's cytogenetic location. “Cen” indicates that the gene is very close to the  centromere. For example, 16pcen refers to the short arm of chromosome 16 near the centromere. “Ter” stands for terminus, which indicates that the gene is very close to the end of the p or q arm. For example, 14qter refers to the tip of the long arm of chromosome 14. (“Tel” is also sometimes used to describe a gene's location. “Tel” stands for telomeres, which are at the ends of each chromosome. The abbreviations “tel” and “ter” refer to the same location.)



The different ways in which a genetic condition can be inherited?

Some genetic conditions are caused by mutations in a single gene. These conditions are usually inherited in one of several patterns, depending on the gene involved:



Patterns of inheritance



Inheritance                   Description                                                                       Examples
pattern



Autosomal                     One mutated copy of the gene in each                      Huntington disease
Dominant                       cell is sufficient for a person to be affected               Marfan syndrome
by an autosomal dominant disorder. In
some cases, an affected person inherits the
condition from an affected parent.
                                   In others, the condition may result from a new
                                      mutation in the geneand occur in people with
                                      no history of the disorder in their family



Autosomal                In autosomal recessive inheritance,                           Both cystic fibrosis
Recessive                copies of the gene in each cell have                          sickle
mutations. The parents of an individual with
an autosomal recessive condition each carry
one copy of the mutated gene, but they typically
do not show signs and symptoms of the condition.

Autosomal recessive disorders are typically
not seen in every generation of an affected
family.



X-linked                    X-linked dominant disorders are caused by               Fragile X
Dominant                  mutations in genes on the X chromosome,                syndrome
one of the two sex chromosomes in
each cell. In females (who have two X
chromosomes), a mutation in one of the two
copies of the gene in each cell is sufficient
to cause the disorder. In males (who have
only one X chromosome), a mutation in the
only copy of the gene in each cell causes
the disorder. In most cases, males experience
more severe symptoms of the disorder than
females. A characteristic of X-linked inheritance
is that fathers cannot pass X-linked traits to their
sons (no male-to-male transmission).



X-linked                    X-linked recessive disorders are also                         Hemophilia,
Recessive                caused by mutations in genes on the X                      Fabry disease
chromosome. In males (who have only
one X chromosome), one altered copy of
the gene in each cell is sufficient to cause
the condition. In females (who have two
X chromosomes), a mutation would have
to occur in both copies of the gene to
cause the disorder.
Because it is unlikely that females will have
two altered copies of this gene, males are
affected by X-linked recessive disorders
much more frequently than females. A
characteristic of X-linked inheritance is that
fathers cannot pass X-linked traits to their
sons (no male-to-male transmission).



Y-linked                   A condition is considered Y-linked if the                     Y chromosome
mutated gene that causes the disorder                      infertility, someis             located on the Y chromosome, one of                         cases of Swyer
the two sex chromosomes in each of a                     syndrome
male's cells. Because only males have a
Y chromosome, in Y-linked inheritance, a
mutation can only be passed from father to
son.

Codominant             In codominant inheritance, two different                    ABO blood group
versions (alleles) of a gene are expressed,
and each version makes a slightly different
protein. Both alleles influence the genetic trait
or determine the characteristics of the genetic
condition.



Mitochondrial            Mitochondrial inheritance, also known as                  Leber hereditary
maternal inheritance, applies to genes in                   optic neuropathy
mitochondrial DNA. Mitochondria, which                   (LHON)
are structures in each cell that convert
molecules into energy, each contain a
small amount of DNA. Because only
egg cells contribute mitochondria to the
developing embryo, only females can pass
on mitochondrial mutations to their children
Conditions resulting from mutations in
mitochondrial DNA can appear in every
generation of a family and can affect both
males and females, but fathers do not pass
these disorders to their daughters or sons.


Many health conditions are caused by the combined effects of multiple genes or by interactions between genes and the environment. Such disorders usually do not follow the patterns of inheritance described above. Examples of conditions caused by multiple genes or gene/environment interactions include heart disease, diabetes, schizophrenia, and certain types of cancer.
Disorders caused by changes in the number or structure of chromosomes also do not follow the straightforward patterns of inheritance listed above.
Other genetic factors sometimes influence how a disorder is inherited.

Reduced penetrance and variable expressivity
Reduced penetrance and variable expressivity are factors that influence the effects of particular genetic changes. These factors usually affect disorders that have an autosomal dominant pattern of inheritance, although they are occasionally seen in disorders with an autosomal recessive inheritance pattern.

Reduced penetrance

Penetrance refers to the proportion of people with a particular genetic change (such as a mutation in a specific gene) who exhibit signs and symptoms of a genetic disorder. If some people with the mutation do not develop features of the disorder, the condition is said to have reduced (or incomplete) penetrance.

Reduced penetrance often occurs with familial cancer syndromes. For example, many people with a mutation in the BRCA1 or BRCA2 gene will develop cancer during their lifetime, but some people will not. Doctors cannot predict which people with these mutations will develop cancer or when the tumors will develop.

Reduced penetrance probably results from a combination of genetic, environmental, and lifestyle factors, many of which are unknown. This phenomenon can make it challenging for genetics professionals to interpret a person’s family medical history and predict the risk of passing a genetic condition to future generations.


Variable expressivity

Although some genetic disorders exhibit little variation, most have signs and symptoms that differ among affected individuals. Variable expressivity refers to the range of signs and symptoms that can occur in different people with the same genetic condition. For example, the features of Marfan syndrome vary widely— some people have only mild symptoms (such as being tall and thin with long, slender fingers), while others also experience life-threatening complications involving the heart and blood vessels. Although the features are highly variable, most people with this disorder have a mutation in the same gene (FBN1).

As with reduced penetrance, variable expressivity is probably caused by a combination of genetic, environmental, and lifestyle factors, most of which have not been identified. If a genetic condition has highly variable signs and symptoms, it may be challenging to diagnose.


Gene therapy

Gene therapy is an experimental technique that uses genes to treat or prevent disease. In the future, this technique may allow doctors to treat a disorder by inserting a gene into a patient’s cells instead of using drugs or surgery. Researchers are testing several approaches to gene therapy, including:

  • Replacing a mutated gene that causes disease with a healthy copy of the gene.
  • Inactivating, or “knocking out,” a mutated gene that is functioning improperly.
  • Introducing a new gene into the body to help fight a disease.

Although gene therapy is a promising treatment option for a number of diseases (including inherited disorders, some types of cancer, and certain viral infections), the technique remains risky and is still under study to make sure that it will be safe and effective. Gene therapy is currently only being tested for the treatment of diseases that have no other cures.


How does gene therapy work?

Gene therapy is designed to introduce genetic material into cells to compensate for abnormal genes or to make a beneficial protein. If a mutated gene causes a necessary protein to be faulty or missing, gene therapy may be able to introduce a normal copy of the gene to restore the function of the protein.

A gene that is inserted directly into a cell usually does not function. Instead, a carrier called a vector is genetically engineered to deliver the gene. Certain viruses are often used as vectors because they can deliver the new gene by infecting the cell. The viruses are modified so they can't cause disease when used in people. Some types of virus, such as retroviruses, integrate their genetic material (including the new gene) into a chromosome in the human cell. Other viruses, such as adenoviruses, introduce their DNA into the nucleus of the cell, but the DNA is not integrated into a chromosome.

The vector can be injected or given intravenously (by IV) directly into a specific tissue in the body, where it is taken up by individual cells. Alternately, a sample of the patient's cells can be removed and exposed to the vector in a laboratory setting. The cells containing the vector are then returned to the patient. If the treatment is successful, the new gene delivered by the vector will make a functioning protein.

Researchers must overcome many technical challenges before gene therapy will be a practical approach to treating disease. For example, scientists must find better ways to deliver genes and target them to particular cells. They must also ensure that new genes are precisely controlled by the body













Genetic Testing Methods

If you made it through the above information you may be interested in how it is possible to detect these genetic variances.
We are looking for two distinct types of variances, SNPs (Single-Nucleotide Polymorphism) and CNVs (Copy Number variants).


Microarray-based genotyping vs Whole Exome/Genome Sequencing
There are three general types of testing that you might encounter in trying to achieve a biological diagnosis of a person’s autism.

The simplest and cheapest is microarray genotyping, also known as chromosomal microarray CMA .  This tests the prevalence of a very large number of known suspect DNA sequences that have been programed into the machine.  CMA is often suggested as the first tier test for individuals with developmental disabilities, intellectual disabilities, autism spectrum disorders, or multiple congenital anomalies.  This is a good method if you know what variances you are looking for.

Whole Exome Sequencing (WES) established the exact nucleotide sequences of DNA at the thousands of exon loci tested.  This test used to be very expensive, but is now quite widely available in the United States, both for research and commercially.

Whole Genome Sequencing (WGS) looks at the entire genome, rather than the much smaller exome.  This test is available, but normally with the caveat “for research purposes”. This should be the holly grail of genetic testing.

The cost of all types of genetic testing continues to fall.

The tests are only as good as the interpretation of their results.

If you live in the US, you can join SPARK for free. This is another initiative of the Simons Foundation, this time to collect genetic material from people affected by autism. They aim to collect saliva samples from 50,000 people with autism, and the their families.  They will then conduct Whole Exome Sequencing (WES).

                          https://sparkforautism.org/portal/page/about-spark/



A more ambitious project is MSSNG, which Google backed project to analyse the entire genome in 10,000 people with autism.

https://www.mss.ng/#