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Tuesday 23 April 2024

Maternal Agmatine or Choline to prevent autism? International brain pH project. Androgen levels in autism spectrum disorders. Apigenin works for BTBR mice. Auditory hypersensitivity, myelin and Nav1.2 channels. Dopamine transporter binding abnormalities and self-injury

 


Shutting the stable door after the horse has bolted


Today’s post is a summary of what I found interesting in the latest research.  Many items have been touched on previously.

The topic of maternal treatment to prevent future autism did come up in some recent comments on this blog. Two of the recent papers cover this very subject. One uses agmatine, from my autism PolyPill therapy, while the other used choline.

Auditory sound sensitivity is a complex subject and today we see the potential role impaired myelination and Nav1.2 ion channels can play.

A Chinese study reconfirms the elevated level of androgen hormones in autism.  

Apigenin which was covered in an earlier post is shown to help “autistic” mice in the popular BTBR model. This is a model where the corpus callosum is entirely absent.

Self-injury is a recuring nightmare for many with severe autism and today we look at a possible correlation with dopamine transporter binding abnormalities.

We start with easier subject matter and leave the hard parts for later in the post.


Preventing future autism

It may seem like too late to be talking about preventing autism, but it is a recurring subject. Today we have two new ideas that have appeared in the literature, and both are very simple. One is choline and other agmatine; both are used in the treatment of already existing autism.

 

Maternal choline to prevent autism

“maternal choline supplementation may be sufficient to blunt some of the behavioral and neurobiological impacts of inflammatory exposures in utero, indicating that it may be a cheap, safe, and effective intervention for neurodevelopmental disorders.” 

 

Maternal choline supplementation modulates cognition and induces anti-inflammatory signaling in the prefrontal cortex of adolescent rats exposed to maternal immune activation


Maternal infection has long been described as a risk factor for neurodevelopmental disorders, especially autism spectrum disorders (ASD) and schizophrenia. Although many pathogens do not cross the placenta and infect the developing fetus directly, the maternal immune response to them is sufficient to alter fetal neurodevelopment, a phenomenon termed maternal immune activation (MIA). Low maternal choline is also a risk factor for neurodevelopmental disorders, and most pregnant people do not receive enough of it. In addition to its role in neurodevelopment, choline is capable of inducing anti-inflammatory signaling through a nicotinic pathway. Therefore, it was hypothesized that maternal choline supplementation would blunt the neurodevelopmental impact of MIA in offspring through long- term instigation of cholinergic anti-inflammatory signaling.

To model MIA in rats, the viral mimetic polyinosinic:polycytidylic acid (poly(I:C)) was used to elicit a maternal antiviral innate immune response in dams both with and without choline supplementation. Offspring were reared to both early and late adolescent stages (postnatal days 28 and 50, respectively), where cognition and anxiety-related behaviors were examined. After behavioral testing, animals were euthanized, and their prefrontal cortices (PFCs) were collected for analysis. MIA offspring demonstrated sex-specific patterns of altered cognition and repetitive behaviors, which were modulated by maternal choline supplementation. Choline supplementation also bolstered anti-inflammatory signaling in the PFCs of MIA animals at both early and late adolescent stages. These findings suggest that maternal choline supplementation may be sufficient to blunt some of the behavioral and neurobiological impacts of inflammatory exposures in utero, indicating that it may be a cheap, safe, and effective intervention for neurodevelopmental disorders.

 

Prenatal Agmatine to prevent autism

Agmatine is a cheap bodybuilder supplement also used in psychiatry that has been extensively covered in this blog. Here we see how in a popular mouse model it can prevent autism.


The prenatal use of agmatine prevents social behavior deficits in VPA-exposed mice by activating the ERK/CREB/BDNF signaling pathway


Background: According to reports, prenatal exposure to valproic acid can induce autism spectrum disorder (ASD)-like symptoms in both humans and rodents. However, the exact cause and therapeutic method of ASD is not fully understood. Agmatine (AGM) is known for its neuroprotective effects, and this study aims to explore whether giving agmatine hydrochloride before birth can prevent autism-like behaviors in mouse offspring exposed prenatally to valproic acid.

Methods: In this study, we investigated the effects of AGM prenatally on valproate (VPA)-exposed mice. We established a mouse model of ASD by prenatally administering VPA. From birth to weaning, we evaluated mouse behavior using the marble burying test, open-field test, and three-chamber social interaction test on male offspring.

Results: The results showed prenatal use of AGM relieved anxiety and hyperactivity behaviors as well as ameliorated sociability of VPA-exposed mice in the marble burying test, open-field test, and three-chamber social interaction test, and this protective effect might be attributed to the activation of the ERK/CREB/BDNF signaling pathway.

Conclusion: Therefore, AGM can effectively reduce the likelihood of offspring developing autism to a certain extent when exposed to VPA during pregnancy, serving as a potential therapeutic drug.


This builds on an earlier paper that first identified the benefit.

 

Agmatine rescues autistic behaviors in the valproic acid-induced animal model of autism

  

Highlights

                  Single treatment of agmatine rescues social impairment in the VPA-induced animal model of autism.

                  Effect of agmatine in social improvement in the VPA model is induced from agmatine itself, not its metabolite.

                  Agmatine rescues repetitive and hyperactive behavior, and seizure susceptibility in the VPA model.

                  Overly activated ERK1/2 in the brain of the VPA model is relieved by agmatine.

 

Apigenin


50mg of Apigenin

1g of dried parsley
15-20g of dried chamomile flowers

 

I have previously written about Apigenin, which is an OTC supplement. There has been another paper recently published about it. There is a logical connection with the maternal choline therapy from above.

 

What does Apigenin have in common with Choline?  α7-nAChRs

Choline is interesting because it acts as both a precursor for acetylcholine synthesis and it is a neuromodulator itself.

Choline is activates α7-nAChRs, alpha-7 nicotinic acetylcholine receptors.

These receptors are extremely important in learning and sensory processing.  They also play a key role in inflammation and signaling via the vagus nerve.

Apigenin is a flavonoid found in many plants, fruits, and vegetables. It has been shown to have a number of health benefits, including anti-inflammatory and antioxidant effects. Apigenin has also been shown to interact with α7-nAChRs.

Studies have shown that apigenin can:

Enhance α7-nAChR function: Apigenin has been shown to increase the activity of α7-nAChRs. This may be due to its ability to bind to a specific site on the receptor.

Protect α7-nAChRs from damage: Apigenin may also help to protect α7-nAChRs from damage caused by oxidative stress.

 

Apigenin Alleviates Autistic-like Stereotyped Repetitive Behaviors and Mitigates Brain Oxidative Stress in Mice


Studying the involvement of nicotinic acetylcholine receptors (nAChRs), specifically α7-nAChRs, in neuropsychiatric brain disorders such as autism spectrum disorder (ASD) has gained a growing interest. The flavonoid apigenin (APG) has been confirmed in its pharmacological action as a positive allosteric modulator of α7-nAChRs. However, there is no research describing the pharmacological potential of APG in ASD. The aim of this study was to evaluate the effects of the subchronic systemic treatment of APG (10–30 mg/kg) on ASD-like repetitive and compulsive-like behaviors and oxidative stress status in the hippocampus and cerebellum in BTBR mice, utilizing the reference drug aripiprazole (ARP, 1 mg/kg, i.p.). BTBR mice pretreated with APG (20 mg/kg) or ARP (1 mg/g, i.p.) displayed significant improvements in the marble-burying test (MBT), cotton-shredding test (CST), and self-grooming test (SGT) (all p < 0.05). However, a lower dose of APG (10 mg/kg, i.p.) failed to modulate behaviors in the MBT or SGT, but significantly attenuated the increased shredding behaviors in the CST of tested mice. Moreover, APG (10–30 mg/kg, i.p.) and ARP (1 mg/kg) moderated the disturbed levels of oxidative stress by mitigating the levels of catalase (CAT) and superoxide dismutase (SOD) in the hippocampus and cerebellum of treated BTBR mice. In patch clamp studies in hippocampal slices, the potency of choline (a selective agonist of α7-nAChRs) in activating fast inward currents was significantly potentiated following incubation with APG. Moreover, APG markedly potentiated the choline-induced enhancement of spontaneous inhibitory postsynaptic currents. The observed results propose the potential therapeutic use of APG in the management of ASD. However, further preclinical investigations in additional models and different rodent species are still needed to confirm the potential relevance of the therapeutic use of APG in ASD.

  

Altered acidity (pH) levels inside the brain

I found it intriguing that a large study has examined the altered acidity (pH) levels inside the brain of those with neurological disorders.

For all the disorders other than autism there was a clear pattern of low pH, which means increased acidity.

For autism certain autism models exhibited decreased pH and increased lactate levels, but others showed the opposite pattern, reflecting subpopulations within autism.

Altered brain energy metabolism is an acknowledged feature of autism, so we should not be surprised to find altered levels of acidity.

The easy reading version:

 

Brain Acidity Linked With Multiple Neurological Disorders

 

The study itself:

Large-scale animal model study uncovers altered brain pH and lactate levels as a transdiagnostic endophenotype of neuropsychiatric disorders involving cognitive impairment

Increased levels of lactate, an end-product of glycolysis, have been proposed as a potential surrogate marker for metabolic changes during neuronal excitation. These changes in lactate levels can result in decreased brain pH, which has been implicated in patients with various neuropsychiatric disorders. We previously demonstrated that such alterations are commonly observed in five mouse models of schizophrenia, bipolar disorder, and autism, suggesting a shared endophenotype among these disorders rather than mere artifacts due to medications or agonal state. However, there is still limited research on this phenomenon in animal models, leaving its generality across other disease animal models uncertain. Moreover, the association between changes in brain lactate levels and specific behavioral abnormalities remains unclear. To address these gaps, the International Brain pH Project Consortium investigated brain pH and lactate levels in 109 strains/conditions of 2,294 animals with genetic and other experimental manipulations relevant to neuropsychiatric disorders. Systematic analysis revealed that decreased brain pH and increased lactate levels were common features observed in multiple models of depression, epilepsy, Alzheimer’s disease, and some additional schizophrenia models. While certain autism models also exhibited decreased pH and increased lactate levels, others showed the opposite pattern, potentially reflecting subpopulations within the autism spectrum. Furthermore, utilizing large-scale behavioral test battery, a multivariate cross-validated prediction analysis demonstrated that poor working memory performance was predominantly associated with increased brain lactate levels. Importantly, this association was confirmed in an independent cohort of animal models. Collectively, these findings suggest that altered brain pH and lactate levels, which could be attributed to dysregulated excitation/inhibition balance, may serve as transdiagnostic endophenotypes of debilitating neuropsychiatric disorders characterized by cognitive impairment, irrespective of their beneficial or detrimental nature.

In conclusion, the present study demonstrated that altered brain pH and lactate levels are commonly observed in animal models of SZ, BD, ID, ASD, AD, and other neuropsychiatric disorders. These findings provide further evidence supporting the hypothesis that altered brain pH and lactate levels are not mere artifacts, such as those resulting from medication confounding, but are rather involved in the underlying pathophysiology of some patients with neuropsychiatric disorders. Altered brain energy metabolism or neural hyper- or hypoactivity leading to abnormal lactate levels and pH may serve as a potential therapeutic targets for neuropsychiatric disorders

 

Why would the brain be acidic (reduced pH)?

To function optimally mitochondria need adequate oxygen and glucose. When performance is impaired, for example due to the lack of Complex 1, mitochondria switch from OXPHOS (oxidative phosphorylation) to fermentation to produce energy (ATP). Lactic acid is the byproduct and this will lower pH.

 

Does brain pH matter?

It does matter and is linked to cognitive impairments, headaches, seizures etc.

Many enzymes in the brain rely on a specific pH range to function properly. Deviations from the ideal pH can hinder their activity, impacting various neurochemical processes essential for brain function.

Some ion channels are pH sensitive.

 

Chemical buffers in the brain aim to regulate pH in the brain

·       Carbonic Acid/Bicarbonate Buffer System: Similar to the blood, the brain utilizes this system to regulate pH.

·   Organic Phosphates: These molecules, like creatine phosphate, can act as buffers in the brain by binding or releasing hydrogen ions.

These buffering systems work together to maintain a tightly controlled pH range in both the blood (around 7.35-7.45) and the brain (slightly more acidic than blood, around 7.0-7.3). Even slight deviations from this ideal range can have significant consequences for cellular function.

  

Androgen Levels in Autism

Androgens are male hormones like testosterone, DHEA and DHT, but females have them too, just at lower levels.

Drugs that reduce the level of these hormones are called antiandrogens.

Finasteride reduces DHT and is used to treat hair loss in men as Propecia. This drug was trialed in women, but failed to show a benefit over the placebo.

The main use of Finasteride is for the treatment of benign prostatic hyperplasia (BPH) in older men.

Women sometimes take antiandrogens like Spironolactone to control acne.

Numerous studies have show elevated levels of males hormones in both males and females with autism.

A recent paper was published on this very subject: 


Androgen levels in autism spectrum disorders: A systematic review and meta-analysis

Background:

Accumulating evidence suggests that the autism spectrum disorder (ASD) population exhibits altered hormone levels, including androgens. However, studies on the regulation of androgens, such as testosterone and dehydroepiandrosterone (DHEA), in relation to sex differences in individuals with ASD are limited and inconsistent. We conducted the systematic review with meta-analysis to quantitatively summarise the blood, urine, or saliva androgen data between individuals with ASD and controls.

Methods:

A systematic search was conducted for eligible studies published before 16 January 2023 in six international and two Chinese databases. We computed summary statistics with a random-effects model. Publication bias was assessed using funnel plots and heterogeneity using I 2 statistics. Subgroup analysis was performed by age, sex, sample source, and measurement method to explain the heterogeneity.

Results:

17 case-control studies (individuals with ASD, 825; controls, 669) were assessed. Androgen levels were significantly higher in individuals with ASD than that in controls (SMD: 0.27, 95% CI: 0.06-0.48, P=0.01). Subgroup analysis showed significantly elevated levels of urinary total testosterone, urinary DHEA, and free testosterone in individuals with ASD. DHEA level was also significantly elevated in males with ASD. Androgen levels, especially free testosterone, may be elevated in individuals with ASD and DHEA levels may be specifically elevated in males.

 

By coincidence I was just sent the paper below, showing the benefit of Finasteride in one model of autism. 

Therapeutic effect of finasteride through its antiandrogenic and antioxidant role in a propionic acid-induced autism model: Demonstrated by behavioral tests, histological findings and MR spectroscopy

 

I do recall I think it was Tyler, long ago, writing a comment about the potential to use Finasteride in autism.

Some very expensive antiandrogens have been used in autism and this became rather controversial.

We saw in earlier posts that RORα/RORalpha/RORA is a key mechanism where the balance between male and female hormones controls some key autism gene.

 


The schematic illustrates a mechanism through which the observed reduction in RORA in autistic brain may lead to increased testosterone levels through downregulation of aromatase. Through AR, testosterone negatively modulates RORA, whereas estrogen upregulates RORA through ER.

 androgen receptor = AR             estrogen receptor = ER


Cerebellum and neurodevelopmental disorders: RORα is a unifying force

Errors of cerebellar development are increasingly acknowledged as risk factors for neuro-developmental disorders (NDDs), such as attention deficit hyperactivity disorder (ADHD), autism spectrum disorder (ASD), and schizophrenia. Evidence has been assembled from cerebellar abnormalities in autistic patients, as well as a range of genetic mutations identified in human patients that affect the cerebellar circuit, particularly Purkinje cells, and are associated with deficits of motor function, learning and social behavior; traits that are commonly associated with autism and schizophrenia. However, NDDs, such as ASD and schizophrenia, also include systemic abnormalities, e.g., chronic inflammation, abnormal circadian rhythms etc., which cannot be explained by lesions that only affect the cerebellum. Here we bring together phenotypic, circuit and structural evidence supporting the contribution of cerebellar dysfunction in NDDs and propose that the transcription factor Retinoid-related Orphan Receptor alpha (RORα) provides the missing link underlying both cerebellar and systemic abnormalities observed in NDDs. We present the role of RORα in cerebellar development and how the abnormalities that occur due to RORα deficiency could explain NDD symptoms. We then focus on how RORα is linked to NDDs, particularly ASD and schizophrenia, and how its diverse extra-cerebral actions can explain the systemic components of these diseases. Finally, we discuss how RORα-deficiency is likely a driving force for NDDs through its induction of cerebellar developmental defects, which in turn affect downstream targets, and its regulation of extracerebral systems, such as inflammation, circadian rhythms, and sexual dimorphism.

  



Figure 2. RORα regulates multiple genes and plays extensive roles in cerebellar development. (A) Key stages of PC development which are regulated by RORα. These are at all stages from embryonic development to adult maintenance. (B) A schema showing the central role of RORα in multiple cellular processes, that are modified in NDDs. When RORα is reduced (central red circle), its regulation of gene transcription is altered. Here we include the known RORα target genes that are also involved in NDDs. The effects in red illustrate the induced abnormalities according to the direction of change: estrogen and PC development are reduced, circadian rhythms are perturbed, but inflammation and ROS are increased.

 

Sound sensitivity in autism and Nav1.2

At this point today’s post does get complicated.

Researchers have learnt that the sodium ion channel Nav1.2 (expressed by the SCN2A gene) can play a key role in hypersensitivity to sound in autism.

Lack of these ion channels in the cells that produce myelin produces “faulty auditory circuits”, with too much sound sensitivity.

An impairment in myelin structure can trigger cascading effects on neuronal excitability. Sound sensitivity is just one example.

There is a great deal of evidence that genes involved in myelination are miss-expressed in many models of autism. Imaging studies have shown variations in myelination.

 

Scn2a deletion disrupts oligodendroglia function: Implication for myelination, neural circuitry, and auditory hypersensitivity in ASD

Autism spectrum disorder (ASD) is characterized by a complex etiology, with genetic determinants significantly influencing its manifestation. Among these, the Scn2a gene emerges as a pivotal player, crucially involved in both glial and neuronal functionality. This study elucidates the underexplored roles of Scn2a in oligodendrocytes, and its subsequent impact on myelination and auditory neural processes. The results reveal a nuanced interplay between oligodendrocytes and axons, where Scn2a deletion causes alterations in the intricate process of myelination. This disruption, in turn, instigates changes in axonal properties and neuronal activities at the single cell level. Furthermore, oligodendrocyte-specific Scn2a deletion compromises the integrity of neural circuitry within auditory pathways, leading to auditory hypersensitivity—a common sensory abnormality observed in ASD. Through transcriptional profiling, we identified alterations in the expression of myelin-associated genes, highlighting the cellular consequences engendered by Scn2a deletion. In summary, the findings provide unprecedented insights into the pathway from Scn2a deletion in oligodendrocytes to sensory abnormalities in ASD, underscoring the integral role of Scn2a-mediated myelination in auditory responses. This research thereby provides novel insights into the intricate tapestry of genetic and cellular interactions inherent in ASD.

Therefore, our study underscores the region-specific relationship between myelin integrity and ion channel distribution in the developing brain. We emphasize that any disturbances in myelin structure can trigger cascading effects on neuronal excitability and synaptic function in the CNS, especially at nerve terminals in the auditory nervous system. 

How are Nav1.2  channels, encoded by Scn2a, involved in OL maturation and myelination? One possible explanation is that the activation of Nav1.2 may be pivotal for triggering Cav channel activation, leading to a Ca2+ flux within OLs, which is involved in OL proliferation, migration, and differentiation. Specifically, Ca2+ signaling facilitated by R-type Cav in myelin sheaths at paranodal regions, might influence the growth of myelin sheaths. To activate high-voltage activated calcium channels such as L- and R-Type efficiently, the activation of Nav1.2 channels should be required for depolarizing OL membrane to around -30 mV. Consequently, the synergic interplay between Nav1.2 and Cav channels could amplify calcium signaling in OLs, initiating the differentiation and maturation processes. 

Defects in myelination can create a spectrum of auditory dysfunctions, including hypersensitivity. Our results demonstrated how OL-Scn2a is involved in the relationship between myelin defects, neuronal excitability, and auditory pathology in ASD, potentially paving the way for targeted therapeutic interventions.

 

One subject that some people write to me repeatedly about is self-injurious behavior, so I took note of the paper below.  

Dopamine Transporter Binding Abnormalities Are Associated with Self-injurious Behavior in Autism Spectrum Disorder 

Utilizing single-photon emission computed tomography dopamine transporter scans (DaTscan) we examined whether imaging markers of the dopaminergic system are related to repetitive behaviors as assessed by the Repetitive Behavior Scale-Revised in ASD.

Background: 

Autism spectrum disorder (ASD) is characterized by impairments in social communication, and restricted repetitive behaviors. Self-injurious behaviors are often observed in individuals with ASD. Dopamine is critical in reward, memory, and motor control. Some propose the nigrostriatal motor pathway may be altered in ASD, and alterations in dopamine are reported in some rodent models based on specific ASD genes. Additionally, repetitive behaviors may to be related to reward systems. Therefore, we examined the dopaminergic system, using DaTscans, to explore its relationship with measures of repetitive behavior in a clinical ASD population.

Design/Methods: 

Twelve participants (aged 18–27) with ASD were recruited from the Thompson Center for Autism and Neurodevelopment and completed the Repetitive Behaviors Scale - Revised (RBS-R). Of the 12 participants, 10 underwent a 45-minute DaTscan. ANOVA was used to compare the dopamine imaging findings with the overall total RB scores on the RBS-R. while other domains of the RBS-R were also investigated in an exploratory manner.

Results: 

Five of the participants had regional deficits in dopamine transporter binding in the striatum on DaTscan. Individuals with deficits on the DaTscan had significantly higher Self-Injurious Endorsed Scores than those with normal scans.

Conclusions: 

Half of the DaTscans obtained were determined abnormal, and abnormal scans were associated with greater endorsing of self-injurious behavior. Larger samples are needed to confirm this, and determine the impact of laterality of abnormalities, but this preliminary work suggests a potential role the dopaminergic system in self-injurious RBs. Elucidation of this relationship may be important for future interventional outcomes, with potential impact on targeted treatment, as the only currently approved medications for ASD are atypical neuroleptics.

 

Dopamine transporter binding abnormalities refer to deviations from the normal levels of dopamine transporter (DAT) in the brain. DAT is a protein on the surface of cells that reabsorbs dopamine from the synapse, regulating its availability.

Imaging techniques like DAT scans (dopamine transporter scans) are used to assess DAT levels. These scans measure the binding of radiotracers to DAT, with lower binding indicating reduced DAT levels.

Dopamine transporter binding abnormalities have been linked to various neurological and psychiatric conditions, including:

                 Parkinson's disease: Degeneration of dopamine-producing neurons in the substantia nigra, a hallmark of Parkinson's disease, leads to a significant decrease in dopamine levels and DAT binding in the striatum.

                 Attention deficit hyperactivity disorder (ADHD): Some studies suggest that individuals with ADHD may have abnormal DAT function, though the nature of the abnormality (increased or decreased DAT) is debated.

                 Autism spectrum disorder (ASD): Research suggests that a subgroup of individuals with ASD may have DAT abnormalities, potentially linked to repetitive behaviors and social difficulties.

                 Addiction: Dopamine plays a central role in reward and motivation. Drugs like cocaine and methamphetamine can cause long-term changes in DAT function, potentially contributing to addiction.

DAT binding abnormalities may not always translate to functional impairments.

 

Treatment options for DAT binding abnormalities

Unfortunately, medications that directly target Dopamine Transporter (DAT) binding abnormalities do not exist.

In Parkinson's disease the goal is to increase dopamine levels in the brain. Medications like levodopa, a dopamine precursor, or dopamine agonists (drugs that mimic dopamine) are used.

  

Conclusion

It certainly is not easy to figure out how to treat autism and its troubling symptoms like self-injury. Our reader currently trying to make sure his second child does not have severe autism is wise to invest his time now.

Today we added agmatine and choline to our list of preventative strategies to consider.

As regards strategies to treat autism in children and adults, we see that the research very often is repeating what has already been published over the past two decades.

Ion channels do seem to be central to understanding and treating autism.




Thursday 4 April 2024

Advances in personalized medicine to treat Autism/IDD – Rett syndrome as an example. Also, Piperine to upregulate KCC2, but what about its direct effect on GABAa receptors?

 

Source:  https://www.cell.com/neuron/pdf/S0896-6273(21)00466-9.pdf


Today’s post is drawn from a workshop I am invited to present at an autism conference in Abu Dhabi.

I decided to talk about advances in personalized medicine – no surprise there.  Since I have 2 ½ hours, I thought I will need some interesting examples to maintain the audiences interest.  One such topic is going to be Rett syndrome.

I regard Rett syndrome and all the other such syndromes in this blog as “single gene autisms” (monogenic autism).  If you apply the American DSM classification, from 2013 onwards Rett syndrome is no longer part of autism.  Hopefully there are no such purists attending in Abu Dhabi. 

Two gene therapies for Rett syndrome are currently undergoing human trials and one drug therapy has been FDA approved.  This looks very encouraging, so let’s dig a little deeper.



Rett syndrome can present with a wide range of disability ranging from mild to severe. 

Rett syndrome is the second most common cause of severe intellectual disability after Down syndrome.

Other symptoms may include:

      Loss of speech

      Loss of purposeful use of hands

      Loss of mobility or gait disturbances

      Loss of muscle tone

      Seizures or Rett “episodes”

      Scoliosis

      Breathing issues

      Sleep disturbances

      Slowed rate of growth for head, feet and hands

Here are the new therapies: 


TSHA-102: This gene therapy, developed by Taysha Therapeutics, is a gene replacement therapy that aims to deliver a functional copy of the MECP2 gene to brain cells.  It utilizes an AAV-9 virus to carry the miniMECP2 gene product into cells for the body to produce more MeCP2 protein, which is deficient in Rett syndrome. As of February 2024, Taysha completed dosing for the first cohort (low dose) in their REVEAL Phase 1/2 adolescent and adult trial in Canada, with positive interim data on safety. They are also conducting trials in the US for both pediatric and adolescent/adult populations.

NGN-401: This gene therapy, by Neurogene Inc., employs a different approach. It uses an AAV9 vector to deliver a regulated version of the MECP2 gene called EXACT. This technology aims to control the amount of MECP2 protein produced by the gene, mitigating the risk of overproduction. NGN-401 is currently in a Phase 1/2 trial for girls with Rett syndrome aged 4 to 10 years old.


Daybue (trofinetide)

Daybue is the first and only FDA-approved treatment specifically for Rett syndrome in adults and children two years of age and older. It is not a gene therapy, but rather a medication taken orally.

The optimistic AI generated view:

Here's a breakdown of Daybue for Rett syndrome:

  • Mechanism: The exact way Daybue works in Rett syndrome isn't fully understood, but it's believed to target neuroinflammation and support synaptic function.
  • Dosage: The recommended dose is based on the patient's weight and is taken twice daily, morning and evening, with or without food.
  • Administration: Daybue comes as an oral solution and can be taken directly or through a gastrostomy tube if swallowing is difficult.
  • Efficacy: Studies have shown that Daybue can improve symptoms of Rett syndrome, including reducing scores on the Rett Syndrome Behavior Questionnaire (RSBQ) and showing improvement on the Clinical Global Impression-Improvement (CGI-I) scale.
  • Side Effects: The most common side effects of Daybue are diarrhea and vomiting. Weight loss can also occur in some patients. It's important to consult with a healthcare professional for monitoring and managing any potential side effects.

Daybue is an expensive medication. Here's what we know about the cost:

  • List Price: The list price of Daybue is around $21.10 per milliliter.
  • Annual Cost: This translates to an estimated average annual cost of around $375,000 for patients.
  • Dosage Variability: It's important to note that the dosage of Daybue is based on a patient's weight, so the annual cost can vary depending on the individual.

Insurance and Assistance Programs:

  • The high cost of Daybue highlights the importance of insurance coverage. Whether insurance covers Daybue and to what extent will depend on your specific plan.
  • The manufacturer, Acadia Pharmaceuticals, offers a copay program called Daybue Acadia Connect. This program may help eligible commercially insured patients pay $0 for their monthly prescription.

What are the parents' groups saying? 

Not as good as you might be expecting for $375,000 a year.




Affordable potential alternatives to Daybue/Trofinetide

Daybue/Trofinetide is the product of decades of research into a growth factor called IGF-1.

It is a complicated subject and as usual the abbreviations can be confusing.

As you will see below there already is an OTC product commercialized by one of the original researchers, Dr Jian Guan.

One Rett syndrome parent, who reads this blog, has trialed cGP and sees a benefit. You rather wonder why the Phelan-McDermid, Pitt Hopkins, Angelman and Prader-Willi parents don’t follow him and splash out 50 USD and make a trial.


 


 



Gene-therapy

Gene therapy is undoubtedly very clever and ultimately will likely be the best therapy.  It still may not be that silver bullet.

To be effective gene therapy needs to be given at a very young age, ideally as a fetal therapy prior to birth. Note that we saw that in the Rett mouse model they gave bumetanide to the pregnant mother just before birth.

Fetal therapy is not a crazy idea and much is already written about it; many pregnancies are terminated because genetic anomalies are detected prior to birth. Down syndrome is the best-known example. Fetal therapy is realistic for some disorders.

Girls with Rett syndrome are often diagnosed first with idiopathic autism and then years later with a more precise diagnosis of Rett syndrome. This is a common experience among readers of this blog.


Classic Rett syndrome 

The average age of diagnosis for this form is around 2.5 years old in the US and 5 years old in the UK.  Why do you think that is?

Research in mouse models has shown that the effect of gene therapy ranges from curative when given extremely young to more limited the later it is given.


Off-target effects

Gene therapy has the potential for off-target effects. This is a significant concern in the field and researchers are actively working on ways to minimize these risks. Here is a breakdown of what off-target effects are and why they matter:

During gene therapy, a modified gene is delivered to target cells with the aim of correcting a genetic defect.

Ideally, the modified gene integrates into the intended location in the genome.

However, there's a chance it might insert itself into unintended locations (off-target sites).


Potential Consequences of Off-Target Effects

Disrupting normal genes at off-target sites could lead to unpredictable and potentially harmful consequences. This could include triggering uncontrolled cell growth, which is a risk factor for cancer.

It can also cause unexpected side effects depending on which genes are accidentally disrupted.


Minimizing Off-Target Effects

Researchers are developing various strategies to improve the accuracy and specificity of gene therapy techniques.

This includes using more precise gene editing tools like CRISPR-Cas9 with optimized guide RNAs to reduce off-target edits.

Additionally, researchers are working on methods to detect and potentially repair any off-target modifications that might occur.


Over-expression of the target gene

Yes, there is a possibility that the replaced gene in gene therapy could overproduce the expressed protein. This can be a potential complication and researchers are working on ways to control the level of protein expression. Here's a breakdown of the concern:

  • Gene Dosing: Ideally, gene therapy aims to deliver a functional copy of the gene at the right amount to compensate for the deficiency.
  • Overproduction Risks: However, if the delivered gene is too active or multiple copies are inserted, it can lead to overproduction of the protein.

Consequences of Protein Overproduction:

  • Overproduction of a protein can disrupt the delicate balance in the cell, potentially leading to cell dysfunction or even cell death.
  • In some cases, the protein itself might have harmful effects if present in excessive amounts.


Controlling Protein Expression:

Researchers are developing several strategies to control protein expression in gene therapy:
    • Promoter selection: Using promoters that have a weaker switch can help regulate protein production.
    • Viral vectors: Engineering viral vectors to control the number of gene copies delivered to cells.
    • Inducible systems: Developing gene therapy methods where the expression of the introduced gene can be turned on and off as needed.


The cost of gene therapy

      Despite the high cost, gene therapy can be a cost-effective treatment for some diseases. This is because it can eliminate the need for lifelong treatment with other medications.

      Here are some examples of the cost of currently available pediatric gene therapies:

      Luxturna (gene therapy for Leber congenital amaurosis type 10): $425,000

      Zolgensma (gene therapy for spinal muscular atrophy type 1): $2.1 million

      Skysona (gene therapy for adrenoleukodystrophy): $3 million


Piperine to correct KCC2 expression in Rett syndrome?

One key feature of Rett syndrome is impaired cognition.

As regular readers are aware, there are many types of treatable intellectual disability (ID).

One type of treatable ID is caused when the GABA developmental switch fails to occur shortly after birth.  This creates an excitatory/inhibitory imbalance in neurons which impairs cognition and lowers IQ.

The faulty GABA switch is a feature of many types of autism, but far from all of them.

By using pharmaceuticals to lower chloride within neurons, you can compensate for the failure of the GABA switch.

This treatment can be achieved by:

1.     Blocking or down regulating NKCC1

2.     Up regulating KCC2

In the paper below they look at up regulating KCC2

Pharmacological enhancement of KCC2 gene expression exerts therapeutic effects on human Rett syndrome neurons and Mecp2 mutant mice

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the methyl CpG binding protein 2 (MECP2) gene. There are currently no approved treatments for RTT. The expression of K+/Cl cotransporter 2 (KCC2), a neuron-specific protein, has been found to be reduced in human RTT neurons and in RTT mouse models, suggesting that KCC2 might play a role in the pathophysiology of RTT.

Injection of KEEC KW-2449 or piperine in Mecp2 mutant mice ameliorated disease-associated respiratory and locomotion phenotypes. The small-molecule compounds described in our study may have therapeutic effects not only in RTT but also in other neurological disorders involving dysregulation of KCC2.

Thus, our data demonstrate that activation of the SIRT1 pathway or the TRPV1 channel enhances KCC2 expression in RTT human neurons.

Treatment with piperine (10 μM), an activator of the TRPV1 channel (51), induced a significant rise in KCC2 expression in cultured human neurons 

We already knew this was likely from earlier research from Ben Ari, see below for a reminder.  Is Piperine an interesting option for those restricted to OTC interventions?

Early alterations in a mouse model of Rett syndrome: the GABA developmental shift is abolished at birth

Genetic mutations of the Methyl-CpG-binding protein-2 (MECP2) gene underlie Rett syndrome (RTT). Developmental processes are often considered to be irrelevant in RTT pathogenesis but neuronal activity at birth has not been recorded. We report that the GABA developmental shift at birth is abolished in CA3 pyramidal neurons of Mecp2-/y mice and the glutamatergic/GABAergic postsynaptic currents (PSCs) ratio is increased. Two weeks later, GABA exerts strong excitatory actions, the glutamatergic/GABAergic PSCs ratio is enhanced, hyper-synchronized activity is present and metabotropic long-term depression (LTD) is impacted. One day before delivery, maternal administration of the NKCC1 chloride importer antagonist bumetanide restored these parameters but not respiratory or weight deficits, nor the onset of mortality. Results suggest that birth is a critical period in RTT with important alterations that can be attenuated by bumetanide raising the possibility of early treatment of the disorder.

One day before delivery, maternal administration of the NKCC1 chloride importer antagonist bumetanide restored these parameters but not respiratory or weight deficits, nor the onset of mortality. Results suggest that birth is a critical period in RTT with important alterations that can be attenuated by bumetanide raising the possibility of early treatment of the disorder.

Treating the mother prior to delivery with bumetanide was a partially effective therapy in the mouse model of Rett syndrome.


Piperine

Bumetanide is cheap and very possibly effective in human Rett syndrome, but it is a prescription drug.

Piperine is an OTC supplement and a compound found in black pepper. By activating the TRPV1 channel it causes an increase in expression of the KCC2 transporter that allows flow of chloride out of neurons. So piperine should lower chloride inside neurons.  Piperine can cross the blood brain barrier, so when taken orally it should have some effect on intracellular chloride.


Piperine is also a positive allosteric modulator of GABAA receptors

This means that piperine multiplies the effect of whatever GABA is around. This means that in typical people piperine should have anti-anxiety effects.

Piperine was recently found to interact with a previously unknown  benzodiazepine-independent binding site.

Researchers are currently toying with the piperine molecule to try and separate the effect on TRPV1 from the effects on  GABAA.  They want to create 2 new drugs.

1.     a selective TRPV1 activator

2.     a selective GABAA modulator (PAM)


Piperine as an alternative or complement to Bumetanide?

One effect of piperine would be great to have (TRPV1 activator) but the second effect would not be helpful (positive allosteric modulator of GABAA).

The question is what is the net effect. Nobody will be able to answer that without a human trial.

I was advised long ago by one drug developer than it is best to focus on reducing flow into neurons via NKCC1, rather than increase its exit by KCC2, because nobody had yet been successful with KCC2; many have tried.  KCC2 plays a key role in neuropathic pain and that is why it has been researched.


Conclusion

We did see years ago that taking coffee with your bumetanide made sense. Coffee contains compounds that are OAT3 inhibitors and slow down the excretion of bumetanide from the body; coffee increases the effect of bumetanide. You can achieve something very similar by just increasing the dose of bumetanide.

Taking black pepper (piperine) with your bumetanide might be good, or might not be. It certainly would be easy to find out. As with Daybue/Trofinetide, the result is likely to vary from person to person. If GABA function, post- bumetanide, is still a bit excitatory amplifying GABA signaling will make autism worse. If GABA function has been shifted to inhibitory then amplifying GABA signaling will be calming.

Gene therapy will require much earlier diagnosis of single gene autisms.

“Precision medicine” therapies like Daybue/Trofinetide may not be that precise after all and large variations exist in the response, even among children with the same affected gene.

The huge expense means that for most of the world they will see no benefit from gene therapy or indeed “precision medicine.”

The low hanging fruit is to repurpose affordable existing drugs and get the benefit from their secondary effects.  This is what I term personalized medicine.

The research clearly indicates that some girls with Rett syndrome likely will benefit from Bumetanide therapy. For a young child this therapy would cost 50 US dollars/euros a year, if you pay the actual price for generics.

Why are they trialing genetic therapies for Rett instead of first doing the obvious thing and trialing cheap bumetanide? They will likely be able to sell the gene therapy for $2 million a shot.  There is little interest in trialing a $50 a year therapy.

Our new reader from Turkey, MÜCADELECI ANNE DENIZ ( = FIGHTING MOTHER DENIZ), likely does not have $2 million to spend, but seems to be on the way to creating her own personalized medicine therapy for her son. Good luck to her.

As to the cGP Max supplement, it seems to work for some and have no effect in others. Nobody has reported any side effects. It looks worth a try for Rett syndrome.  As a supplement it is not cheap, that is until you see what they charge for Daybue.