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Thursday 6 February 2020

Monty in Mardin – on the Silk Road with Autism



Mardin is a city in Southeastern Turkey on the overland Silk Road (in red)


Today’s post does not have any science.

Just after the new year Monty went to southeast Turkey to visit Mardin, an old city that was on the Silk Road linking Asia to Europe and then to see more of Istanbul.

Travel can be difficult for people with autism, even fully verbal people with above average IQ.  Some people with Asperger’s refuse to fly because they fear the plane is going to crash.  Exaggerations and distortions of normal perception are actually examples of psychotic behavior, in the extreme these are the "positive" symptoms of schizophrenia.  This loss of touch with reality does feature to varying degrees in some Aspies.

Monty started to fly while still a baby and, in spite of a few glitches along the way, has become quite a seasoned traveller for a 16 year old.  

The latest trip was a little off the beaten track for western tourists, in fact we did not see any other western tourists, just a few people visiting from Istanbul.

Mardin is an old town built up the side of a small mountain giving panoramic views across Mesopotamia.  All the old merchant’s houses have large terraces facing the same direction, so they could see incoming traffic from the Silk Road.



Mardin


            A merchant’s house in Mardin



The Front Door of the Deyrilzafaran Monastery




Midyat, a three-thousand-year-old city



Passing for Normal

The Holy Grail for treating someone born with severe autism is just passing for normal.

Passing for normal may not be politically correct, but it is practically correct in actual daily life.

Two recent events stand out in my mind. Walking around the narrow streets of Mardin we entered one of the most impressive old houses.  In the courtyard was a security office; it was clearly was some kind of official building. On the large upper terrace, with the stunning view over Mesopotamia, was a well-dressed man and we started a conversation in English.  He beckoned us to come up past the security guard and see the old house.  It is now used as a school for tourism and hotel/catering.  He offered us Turkish tea and was telling us all about the history of the building and the city.

At first, he thought Monty was a local boy acting as our guide, then later on he turned to Monty and said “what are you going to study later at University?”.  Instead of me having to explain that Monty has autism and so probably will not be going to University, Monty just answered “science”.   Bravo Monty.

The other recent event was the school’s poetry night, when they combine music recitals with poetry readings.  Monty went up on stage for his piano recital, bowed at the end and, as the school principal pointed out to me later, what you saw was just another quiet teenager with some musical talent, not the token one with a disability.  She has known him for 13 years and did not expect him to cope in high school, even with his assistant. Bravo again, Monty.

Unlike how autism is now portrayed in media, people with autism often do look very different and act very differently to other people.  The looking different part could be used to identify the underlying biological cause, but usually is not.  The acting different part is often the result of behaviors learned from their autistic peers and then reinforced by them.  This is a big disadvantage of special schools and group homes, “odd” behaviors get copied until everyone has them.

In a somewhat bizarre twist of nature, sometimes the unusual facial features show up instead on the sibling of the child with autism.  This was recently highlighted again in the research.


Back to Istanbul

Istanbul is one of the world’s great cities, with an amazing location on the Bosporus which connects the Black Sea to the Mediterranean.  It is full of interesting things to see and is great value.  The overland Silk Road ended in Constantinople, now called Istanbul; goods then moved west by sea to Venice.

Istanbul is a good place for people, like Monty, who like to eat fish and also all kinds of cakes and desserts.  Monty even acquired a taste for drinking Turkish tea in small glasses. There are some good, very child-friendly, museums, with parent-friendly admission prices.




Seagulls with the Dolmabahçe Palace on the European side of the Bosphorus in the background




The Rumeli Fortress in Europe and the third Bosphorus Bridge, leading to Asia



Life before the PolyPill

The summer before Monty started his autism Polypill in 2012, we went to Portugal. I still clearly remember having to pin him down in his seat with my arms and legs, as he went berserk in the plane. We had rented a car, but we could not leave Monty in the back with his older brother, because he would attack him.  That was travel with a 9-year-old with medically untreated severe autism.


Expect the unexpected

As our reader Tanya, recently commented, life with autism is always full of surprises, to be overcome.  It is perhaps an adventure you would rather not be on, like a trip with Bear Grylls.  

We still encounter some travel issues, but they are now trivial.  Monty’s ears do not pop when the plane lands and then they hurt and this affects behavior.  The first solution was his idea, jump deep down into a swimming pool.  Ears pop and the problem is solved.

Recently this developed into not swallowing after the plane has landed.  Mouth fills up with saliva, so you cannot talk and also your face bulges and the automatic face recognition at passport control does not work.

I did earlier have Monty’s ears checked and the ENT doctor found nothing, but suggested using the Dymista allergy nose spray containing Azelastine and the steroid Fluticasone. We already have this and it does not help ears pop.

The solution I came up with is to bring a sports-type water bottle on the plane, the type you have to suck to get the water.  During the last 10 minutes of the flight, suck the water, swallow, then squeeze your nose tightly and try to swallow several times.  It works, one problem solved and now we await the next one.

Autism flare-ups, sudden onset vocal or motor tics have all come, been figured out and then been overcome.


Autism and vacations in the literature

Researchers even publish papers about holidays/vacations taken by people with autism.  In short, autism families rarely have holidays and when they do it can be quite stressful for everyone.

The key to happy holidays is medically treating severe autism. 



This work aims at identifying the types of holiday experienced by families of children with autism spectrum disorder (ASD). 35 families of ASD children and 25 control families of children with Down’s Syndrome (DS) living in Scotland took part in a small scale semi-qualitative study to explore their experiences on holiday. In both groups, a significant proportion of the families had not taken any holiday away from home more than once in the last 3 years and there was limited use of children holiday centers. Families of ASD children who had been on holiday expressed overall less positive impressions of their experiences and showed limited use of public places such as restaurants, cafes, cinemas and hotels normally accessed by typical families whilst on vacation. Five areas were identified as influencing the quality of their experiences: 1-child’s disability, particularly with regard to behavior, 2-lack of suitable holiday structures, 3-financial limitation of the family, 4-lack of empathy from surrounding communities towards the disabled child and his or her family, 5- general state of exhaustion of the parents. The paper further describes two pilot holiday community experiences organised with 10 families with ASD children in an attempt to address some of the issues hereby identified. A retrospective analysis of these experiences and surveys suggests that amongst all five identified barriers, the issue of the child’s behavior is the most significant difficulty encountered by these families. Supporting families in understanding and improving their child’s’ behavior is needed to enable families to maximise their experience on holiday. Increasing the understanding of the condition, improving access to leisure activities and some financial aid would equally be beneficial

  

There is an evolving tourism literature around psychological wellbeing, social exclusion and disability. This paper advances tourism knowledge into the terrain of psychological health and developmental complexities, and psychological distress. It draws on a phenomenological position to understand the lived experiences of mothers of children with developmental difficulties, in this case diagnosed with autism spectrum disorder (ASD). It discusses the emotional and everyday challenges of caring for a child diagnosed with ASD on holiday, discusses the perceived benefits holidays offer and outlines care-giving strategies adopted by mothers to manage their children’s tourism experiences. The paper discusses the uniqueness of the context of autism and problematizes popular discourses, which predominantly frame tourism as pleasurable settings of escape, stimulation, novelty and relaxation.

Conclusion The mothers in this study navigated the emotional dimensions of their family holidays through a range of strategies. Firstly, they chose destinations and accommodation, which provided detailed information on all aspects of the holiday to prepare their child in advance. Secondly, they selected quiet destinations and accommodation and thirdly, they packed familiar possessions that are important to the child’s daily routine and ensured that the accommodation could provide for their child’s needs (e.g. Wi-Fi and television). Some mothers also carried an identification card or official document from health or psychology services for their child to deal with public censure if the child’s behaviour became too extreme or to confirm his entitlement to preferential attention, such as in theme park queues. Such strategies could be employed by other parents and, if we can appreciate the coping approaches of parents of children with a disability, these may ‘‘provide valuable insight for other families and parents” (Dodd et al., 2009, p.266). Tourism researchers have much to do to provide such insight and to adequately connect disability, psychological health, distress and the family. This paper has contributed to the unfolding work on this area and, in focusing on developmental difficulties and psychological distress, has widened the scope of an evolving tourism literature exploring wellbeing and psychological health. The area of psychological distress in general, and specifically in tourism, requires much more investigation since studies on disability too often assume that the ‘‘obstacles, barriers and constraints which face disabled people are identical across all people with a disability” (Blichfeldt & Nicolaisen, 2011, p.83). Our research has highlighted the unique context of autism and the challenges faced by mothers of children diagnosed with ASD, illuminating their complex care-giving responsibilities and the impact these have on family holiday experiences, including those of siblings. Whilst all the mothers we spoke with identified benefits to holiday-taking, they balance these against the disruption to routines and exposure to sensory-stimulating environments (Bellini, 2004), which make holidays a challenging experience for both them and their child or children. In addition, expectations to conform to certain behaviours and public reactions to their child’s behaviour can exacerbate the difficulties of holidaying and make them an isolating experience. In this, families with children diagnosed with ASD share common ground with other groups who experience stressful travel encounters in an unaccommodating world (Small & Harris, 2012). Embodiment, attitude, behaviours and stigma are common across disability groups and tourism’s gaze of discipline and surveillance, which causes such stress to many people with disabilities (Eichhorn et al., 2013) and to parents of young children in general, bears further scrutiny. This leads us to perhaps the most thought-provoking contribution of our study. It illustrates the ways in which tourism experiences are journeys of mixed emotions: far from the binary of pleasure and apprehension, they are indeed multifaceted, complex, interlinked and intersubjective (Williams & Aaker, 2002). We need to know more in this area. We also require further explorations of the multiple, complex and nuanced meanings of ‘the holiday’; especially ones that investigate what constitutes a holiday for individuals and families who fall outside of our world’s increasingly unforgiving expectations and norms. Conceptualisations of the holiday have focused on its benefits and on its opportunities for positive transformation and renewal (e.g. Reisinger, 2013). Holidays also entail considerable ‘work’, anxiety and emotional labour (Dieffendorff et al., 2005), especially for care-givers, and particularly as here, for mothers of children diagnosed with ASD. At each stage of the holiday and planning process, the women invested significant time, energy and emotional labour to facilitate trips for their families, in which they themselves experienced both stress and escapism. Indeed, different holiday practices (e.g. air transport, visiting resorts) evoke a series of different emotions and emotional labour depending on the life-world of our participants. With the consequences of emotional labour often resulting in psychological distress (Strazdins & Broom, 2004), we suggest more attention should be given to this area in future tourism research, alongside a more comprehensive account of gender and the lived experience of care-givers in the tourism context.



Conclusion

The target for people with autism should be to live independently.  You may not achieve this, but the closer you get, the better life will be.

Traveling is a good way to learn some of the important life skills you are going to need later.

You can strive to make accommodations for the disability of autism with special autism-friendly periods in shops, theatre, cinemas, etc. There can even be disabled parking privileges for parents, although most people with autism can walk just fine.

The other extreme is tough love, where you always push for being “normal” and fitting in, over acceptance of being different and then just excluding yourself.

What works best for you is determined by how severely disabled the person really is.  Only 15-20% of modern autism is severely disabling; those people really need a lot of help, and indeed personalized medicine.  The majority of those with autism in 2020 are starting from a point (level 1 and 2 in DSM5), not so far away from normal.

Away from the bubble of social media, the real world has changed little.  Strangers on the street did not get more tolerant; kids at school did not stop bullying the odd one out; best to be as "normal" as you can.

Whether you chose travel, sport, music or something else, better to broaden horizons and ambitions with actions, rather than tweet about all the problems.





Tuesday 28 January 2020

Piperine/Resveratrol/Sunitinib for Rett’s and indeed much Autism? Or, R-Baclofen to raise KCC2 expression in Bumetanide-responsive autism.



Piperine/Pepper             Resveratrol/Red wine          Sunitinib/Sutent
  

This post is all about lowering chloride within neurons, by increasing the expression of the transporter that lets it leave, called KCC2.


Today’s post is one I never finished writing from last year; I looked up the price of Sutent/Sunitinib and then I remembered why. It does again highlight how cancer drugs, when they become cheap generics, will provide interesting options for autism treatment. It also shows again how Rett Syndrome is getting attention from researchers.

It also highlights that really clever Americans are looking for bumetanide alternatives, in the false belief that bumetanide has troubling side effects that cannot be managed/mitigated.

The study is by some clever guys in Cambridge Massachusetts.

Another group of clever guys from MIT burned through $40 million dollars a few years ago trying to develop R-Baclofen for Fragile-X and autism.  After that Roche-funded clinical trial failed, R-Baclofen has now been resurrected and a new trial is planned, with different end points (measures of success).

Today we see why many people should indeed respond positively to R-Baclofen, but the mode of action is entirely different to the one originally targeted by the clever guys from MIT.

Tucked away in the supplementary material of today’s paper we see that R-Baclofen increases the expression of the transporter (KCC2) that takes chloride out of neurons. So, R-Baclofen is doing the same thing as Bumetanide, just to a lesser extent and in a different way.  Both lower intracellular chloride.

That means that people responsive to bumetanide should get a further boost from R baclofen, but you might need a lot of it.

Clever they may be, but these researchers do not know how to communicate their findings.  I had to dig through the supplementary tables to extract the good stuff, which is a list of what substances increase KCC2 in regular brains (Table S1) and specifically in Rett Syndrome brains (Table S2).

This blog does rather bang on about blocking/inhibiting NKCC1 that lets chloride into neurons, you can of course alternatively open up KCC2 to let the chloride flood out. This latter strategy is proposed by the MIT researchers.

What really matters is the ratio KCC2/NKCC1.  In people with bumetanide-responsive autism, which pretty clearly will include girls with Rett Syndrome, you want to increase KCC2/NKCC1. So, block/down-regulate NKCC1 and/or up-regulate KCC2.

·        NKCC1

·        KCC2


The researchers identified 14 compounds.  To be useful as drugs these compounds have to be able to cross the blood brain barrier to be of much use, many do not.

In the paper they call KCC2 expression-enhancing compounds KEECs.

We have five approved drugs to add to the list that are functionally the same to primary hit compounds. 

·        Sunitinib
·        Crenolanib
·        Indirubin Monoxiome
·        Cabozantinib
·        TWS-119


The researchers went on to test just two compounds in Rett syndrome mice; they picked piperine (from black pepper) and KW 2449 (a leukemia drug)


Even R-baclofen pops up, with a “B score” of 6.65 (needs to be >3 to increase KCC2 expression).



Abstract
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. To develop neuron-based high-throughput screening (HTS) assays to identify chemical compounds that enhance the expression of the KCC2 gene, we report the generation of a robust high-throughput drug screening platform that allows for the rapid assessment of KCC2 gene expression in genome-edited human reporter neurons. From an unbiased screen of more than 900 small-molecule chemicals, we have identified a group of compounds that enhance KCC2 expression termed KCC2 expression-enhancing compounds (KEECs). The identified KEECs include U.S. Food and Drug Administration-approved drugs that are inhibitors of the fms-like tyrosine kinase 3 (FLT3) or glycogen synthase kinase 3β (GSK3β) pathways and activators of the sirtuin 1 (SIRT1) and transient receptor potential cation channel subfamily V member 1 (TRPV1) pathways. Treatment with hit compounds increased KCC2 expression in human wild-type (WT) and isogenic MECP2 mutant RTT neurons, and rescued electrophysiological and morphological abnormalities of RTT neurons. 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.





Table S1. KEECs identified from screening with WT human KCC2 reporter neurons.






Table S2. KEECs identified from screening with RTT human KCC2 reporter neurons


Note Baclofen, Quercetin, Luteolin etc

















Fig. 3. Identification of KEECs that increase KCC2 expression in human RTT neurons
B score >3 indicates compounds potentially increasing KCC2 expression

In cultured RTT neurons, treatment with KEECs KW-2449 and BIO restored the impaired KCC2 expression and rescued deficits in both GABAergic and glutamatergic neurotransmissions, as well as abnormal neuronal morphology. Previous data suggested that disrupted Cl− homeostasis in the brainstem causes abnormalities in breathing pattern (64), consistent with breathing abnormalities seen in mice carrying a conditional Mecp2 deletion in GABAergic neurons (67). The reduction in locomotion activity observed in the Mecp2 mutant mice has also been attributed to abnormalities in the GABAergic system (65). Therefore, treatment with the KEEC KW-2449 or piperine may ameliorate disease phenotypes in MeCP2 mutant mice through restoration of the impaired KCC2 expression and GABAergic inhibition.

Most KEECs that enhanced KCC2 expression in WT neurons, including KW-2449, BIO, and resveratrol, also induced a robust increase of KCC2 reporter activity in RTT neurons (Fig. 3, A and B; a complete list of hit compounds is provided in table S2). The increase in KCC2 signal induced by KEECs was higher in RTT neurons than in WT neurons,


Our results establish a causal relationship between reduced FLT3 or GSK3 signaling activity and increased KCC2 expression.

Two hit compounds, resveratrol and piperine, act on different pathways than the kinase inhibitors, activating the SIRT1 signaling pathway (50) and the TRPV1 (51), respectively

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


The group of KEECs reported here may help to elucidate the molecular mechanisms that regulate KCC2 gene expression in neurons. A previous study conducted with a glioma cell line showed that resveratrol activates the SIRT1 pathway and reduces the expression of NRSF/REST (50), a transcription factor that suppresses KCC2 expression (52). Our results demonstrate that resveratrol increases KCC2 expression by a similar mechanism, which could contribute to the therapeutic benefit of resveratrol on a number of brain disease conditions (68, 69). We also identified a group of GSK3 pathway inhibitors as KEECs. Overactivation of the GSK3 pathway has been reported in a number of brain diseases (70). Thus, our results suggest that GSK3 pathway inhibitors could exert beneficial effects on brain function through stimulating KCC2 expression. Another major KEEC target pathway, the FLT3 kinase signaling, has been investigated as a cancer therapy target (71, 72). Although FLT3 is expressed in the brain (73), drugs that target FLT3 pathway have not been extensively studied as potential treatments for brain diseases. Our results provide the first evidence that FLT3 signaling in the brain is critical for the regulation of key neuronal genes such as KCC2. Therefore, this work lays the foundation for further research to repurpose a number of clinically approved FLT3 inhibitors as novel brain disease therapies

Our results are valuable for the development of novel therapeutic strategies to treat neurodevelopmental diseases through rectification of dysfunctional neuronal chloride homeostasis. Because of the lack of pharmaceutical reagents that enhance KCC2 expression, bumetanide, a blocker of the inward chloride transporter NKCC1 that counteracts KCC2, has been used as an alternative (74). Bumetanide treatment has shown benefits in treating symptoms in mouse models of fragile X syndrome (75) and Down’s syndrome (76) and was shown to confer symptomatic benefit to human patients with autism or fragile X syndrome (77, 78). These findings strongly suggest that pharmacological restoration of disrupted chloride homeostasis may provide symptomatic treatment for various neurodevelopmental and neuropsychiatric disorders. However, NKCC1 lacks the neuron- restricted expression pattern of KCC2 and is also expressed in nonbrain tissue including kidney and inner ears (79), consistent with knockout of Nkcc1 in mouse model leading to deafness and imbalance (30). Therefore, bumetanide treatment may trigger undesirable side effects, thus severely limiting its therapeutic application. In contrast, the expression of KCC2 is restricted to neurons, and a number of the KEECs identified in this study that enhance KCC2 expression in neurons are Food and Drug Administration–approved and have not elicited any severe adverse effects in clinical trials (80–83). The promising efficacy of KEECs demonstrated in this study and the known safety of the KCC2 target warrant further preclinical and clinical studies to investigate these drugs and their derivatives as potential therapies for neurodevelopmental diseases.

In summary, in this work, we investigated the efficacy of KEECs to rescue a number of well-documented cellular and behavior phenotypes of RTT, including impaired GABA functional switch, reductions in excitatory synapse number and strength, immature neuronal morphology (53, 54), as well as an increase in breathing pauses and a decrease in locomotion (84). It is possible, however, that KEECs may also be effective in treatment of conditions other than RTT, as impairment in KCC2 expression has been linked to many brain diseases (17, 85) including epilepsy (86–88), schizophrenia (19, 20, 89), brain and spinal cord injury (21, 90), stroke and ammonia toxicity conditions (91–93), as well as the impairments in learning and memory observed in the senile brain (23). Thus, a phenotypically diverse array of brain diseases may benefit from enhancing the expression of KCC2. The newly identified KEECs are potential therapeutic agents for otherwise elusive neurological disorders



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. To develop neuron-based high-throughput screening (HTS) assays to identify chemical compounds that enhance the expression of the KCC2 gene, we report the generation of a robust high-throughput drug screening platform that allows for the rapid assessment of KCC2 gene expression in genome-edited human reporter neurons. From an unbiased screen of more than 900 small-molecule chemicals, we have identified a group of compounds that enhance KCC2 expression termed KCC2 expression– enhancing compounds (KEECs). The identified KEECs include U.S. Food and Drug Administration–approved drugs that are inhibitors of the fms-like tyrosine kinase 3 (FLT3) or glycogen synthase kinase 3 (GSK3) pathways and activators of the sirtuin 1 (SIRT1) and transient receptor potential cation channel subfamily V member 1 (TRPV1) pathways. Treatment with hit compounds increased KCC2 expression in human wild-type (WT) and isogenic MECP2 mutant RTT neurons, and rescued electrophysiological and morphological abnormalities of RTT neurons. 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.


By screening these KCC2 reporter human neurons, we identified a number of hits KCC2 expression–enhancing compounds (KEECs) from ~900 small-molecule compounds. Identified KEECs were validated by Western blot and quantitative reverse transcription polymerase chain reaction (RT-PCR) experiments on cultured human wild-type (WT) and isogenic RTT neurons, as well as on organotypic mouse brain slices. Pharmacological and molecular biology experiments showed that identified KEECs act through inhibition of the fms-like tyrosine kinase 3 (FLT3) or glycogen synthase kinase 3b (GSK3b) kinases, or activation of the sirtuin 1 (SIRT1) or transient receptor potential cation channel subfamily V member 1 (TRPV1) pathways. Treatment of RTT neurons with KEECs rescued disease-related deficits in GABA functional switch, excitatory synapses, and neuronal morphological development. Last, injection of the identified KEEC KW-2449 or piperine into a Mecp2 mutant mice ameliorated behavioral phenotypes including breathing pauses and reduced locomotion, which represent important preclinical data, suggesting that the KEECs identified in this study may be effective in restoring impaired E/I balance in the RTT brain and provide symptomatic treatment for patients with RTT.





Fig. 2. KEEC treatment–induced enhancement of KCC2 protein and mRNA expression in cultured organotypic mouse brain slices and a hyperpolarizing EGABA shift in cultured immature neurons.

(E to G) KCC2 and NKCC1 mRNA expression induced by FLT3 inhibitors including sunitinib (n = 4), XL-184 (n = 6), crenolanib (n = 4), or a structural analog of BIO termed indirubin monoxime (n = 6). The calculated ratios of KCC2/NKCC1 mRNA expression are shown in (G). A.U., arbitrary units




Our results are valuable for the development of novel therapeutic strategies to treat neurodevelopmental diseases through rectification of dysfunctional neuronal chloride homeostasis. Because of the lack of pharmaceutical reagents that enhance KCC2 expression, bumetanide, a blocker of the inward chloride transporter NKCC1 that counteracts KCC2, has been used as an alternative (74). Bumetanide treatment has shown benefits in treating symptoms in mouse models of fragile X syndrome (75) and Down’s syndrome (76) and was shown to confer symptomatic benefit to human patients with autism or fragile X syndrome (77, 78). These findings strongly suggest that pharmacological restoration of disrupted chloride homeostasis may provide symptomatic treatment for various neurodevelopmental and neuropsychiatric disorders. However, NKCC1 lacks the neuron restricted expression pattern of KCC2 and is also expressed in nonbrain tissue including kidney and inner ears (79), consistent with knockout of Nkcc1 in mouse model leading to deafness and imbalance (30). Therefore, bumetanide treatment may trigger undesirable side effects, thus severely limiting its therapeutic application. In contrast, the expression of KCC2 is restricted to neurons, and a number of the KEECs identified in this study that enhance KCC2 expression in neurons are Food and Drug Administration–approved and have not elicited any severe adverse effects in clinical trials (80–83). The promising efficacy of KEECs demonstrated in this study and the known safety of the KCC2 target warrant further preclinical and clinical studies to investigate these drugs and their derivatives as potential therapies for neurodevelopmental diseases.


In summary, in this work, we investigated the efficacy of KEECs to rescue a number of well-documented cellular and behavior phenotypes of RTT, including impaired GABA functional switch, reductions in excitatory synapse number and strength, immature neuronal morphology (53, 54), as well as an increase in breathing pauses and a decrease in locomotion (84). It is possible, however, that KEECs may also be effective in treatment of conditions other than RTT, as impairment in KCC2 expression has been linked to many brain diseases (17, 85) including epilepsy (86–88), schizophrenia (19, 20, 89), brain and spinal cord injury (21, 90), stroke and ammonia toxicity conditions (91–93), as well as the impairments in learning and memory observed in the senile brain (23). Thus, a phenotypically diverse array of brain diseases may benefit from enhancing the expression of KCC2. The newly identified KEECs are potential therapeutic agents for otherwise elusive neurological disorders.




The science-light version:-

Drug screen reveals potential treatments for Rett syndrome

An experimental leukemia drug and a chemical in black pepper ease breathing and movement problems in a mouse model of Rett syndrome, according to a new study.

Rett syndrome is a rare brain condition related to autism, caused by mutations in the MECP2 gene. Because the gene is located on the X chromosome, the syndrome occurs almost exclusively in girls. No drugs are available to treat Rett.
The team screened 929 compounds from three large drug libraries, including one focused on Rett therapies. They found 30 compounds that boost KCC2’s expression in the MECP2 neurons; 14 of these also increased the protein’s expression in control neurons.

The team tested two of the identified compounds in mice with mutations in MECP2: KW-2449, which is a small molecule in clinical trials for leukemia, and piperine, an herbal supplement and component of black pepper. These mice have several traits reminiscent of Rett. They are prone to seizures, breathing problems, movement difficulties and disrupted social behavior.
Injecting the mice with either drug daily for two weeks improved the animals’ mobility relative to untreated mice. The drugs also eased the mice’s breathing problems, decreasing the frequency of pauses in breathing (apnea). The findings appeared in July in Science Translational Medicine.


 

Piperine, Resveratrol and analogs thereof

Piperine and Resveratrol are commercially available supplements.

Resveratrol has been mentioned many times in this blog.  It has numerous beneficial properties, to which we can now add increasing KCC2 expression, but it is held back by its poor ability to cross the blood barrier.

The other natural substance highlighted in the study is piperine. Piperine is the substance that gets added to curcumin to increases its bioavailability and hopefully get its health benefits.

Piperine has been recently been found to be a positive allosteric modulator of GABAA receptors.

It may be that piperine has 2 different effects on GABA, or maybe it is just the same one?

The result is that people are trying to develop modified versions of piperine that could be patentable commercial drugs.

Piperine also activated TRPV1 receptors.

You might wonder what is the effect in humans of plain old piperine in bumetanide-responsive autism.

Invitro blood–brain-barrier permeability predictions for GABAA receptor modulating piperine analogs

The alkaloid piperine from black pepper (Piper nigrum L.) and several synthetic piperine analogs were recently identified as positive allosteric modulators of γ-aminobutyric acid type A (GABAA) receptors. In order to reach their target sites of action, these compounds need to enter the brain by crossing the blood–brain barrier (BBB). We here evaluated piperine and five selected analogs (SCT-66, SCT-64, SCT-29, LAU397, and LAU399) regarding their BBB permeability. Data were obtained in three in vitro BBB models, namely a recently established human model with immortalized hBMEC cells, a human brain-like endothelial cells (BLEC) model, and a primary animal (bovine endothelial/rat astrocytes co-culture) model. For each compound, quantitative UHPLC-MS/MS methods in the range of 5.00–500 ng/mL in the corresponding matrix were developed, and permeability coefficients in the three BBB models were determined. In vitro predictions from the two human BBB models were in good agreement, while permeability data from the animal model differed to some extent, possibly due to protein binding of the screened compounds. In all three BBB models, piperine and SCT-64 displayed the highest BBB permeation potential. This was corroborated by data from in silico prediction. For the other piperine analogs (SCT-66, SCT-29, LAU397, and LAU399), BBB permeability was low to moderate in the two human BBB models, and moderate to high in the animal BBB model. Efflux ratios (ER) calculated from bidirectional permeability experiments indicated that the compounds were likely not substrates of active efflux transporters.


The alkaloid piperine, the major pungent component of black pepper (Piper nigrum L.), was recently identified as a positive allosteric γ-aminobutyric acid type A (GABAA) receptor modulator. The compound showed anxiolytic-like activity in behavioral mouse models, and was found to interact with the GABAA receptors at a binding site that was independent of the benzodiazepine binding site [1,2]. Given that the compound complied with Lipinski’s “rule of five” [1], it represented a new scaffold for the development of novel GABAA receptor modulators [1–3]. Given that piperine also activates the transient receptor potential vanilloid 1 (TRPV1) receptors [4] which are involved in pain signaling and regulation of the body temperature [5,6], structural modification of the parent compound was required to dissect GABAA and TRPV1 activating properties

For drugs acting on the central nervous system (CNS), brain penetration is required. This process is controlled by the blood-brain barrier (BBB), a tight layer of endothelial cells lining the brain capillaries that limits the passage of molecules from the blood circulation into the brain [10]. Since low BBB permeability can reduce CNS exposure [11], lead compounds should be evaluated at an early stage of the drug development process for their ability to permeate the BBB [12].

Conclusions

Piperine and five selected piperine analogs with positive GABAA receptor modulatory activity were screened in three in vitro cell-based human and animal BBB models for their ability to cross the BBB. Data from the three models differed to some extent, possibly due to protein binding of the piperine analogs. In all three models, piperine and SCT-64 displayed the highest BBB permeation potential, which could be corroborated by in silico prediction data. For the other piperine analogs (SCT-66, SCT-29, LAU397, and LAU399), BBB permeability was low to moderate in the two human models, and moderate to high in the animal model. ER calculated from bidirectional permeability experiments indicated that the compounds were likely not substrates of active efflux. In addition to the early in vitro BBB permeability assessment of the compounds, further studies (such as PK and drug metabolism studies) are currently in progress in our laboratory. Taken together, these data will serve for selecting the most promising candidate molecule for the next cycle of medicinal chemistry optimization




Conclusion

My conclusions are a little different to the MIT researchers

“The newly identified KEECs are potential therapeutic agents for otherwise elusive neurological disorders.”

This assumes that you cannot safely use bumetanide/azosemide, which you can.  Open your eyes and look at France, where several hundred children with autism are safely taking bumetanide.

”It is possible, however, that KEECs may also be effective in treatment of conditions other than RTT, as impairment in KCC2 expression has been linked to many brain diseases”

We have copious evidence that elevated chloride is a feature of many conditions, not just Rett’s and an effective cheap therapy has been sitting in the pharmacy for decades.

In the clinical trial of R-Baclofen that failed, there were some positive effects on some subjects.  Were the positive effects just caused by the effect of Baclofen in increasing KCC2 expression?

Should R-Baclofen become a cheap generic, it might indeed become a useful add-on for those with bumetanide-responsive. Regular Baclofen (Lioresal) is an approved drug, but it does have some side effects, so most likely R-baclofen will have side effects in some.

Baclofen itself in modest doses has little effect on bumetanide-responsive autism.



A cheap side-effect free KCC2 enhancer would be a good drug for autism, although cheap, safe NKCC1 blockers already exist. 

I have no idea if piperine benefits bumetanide-responsive autism.  Piperine has long been used in traditional medicine.

The TRPV1 receptor also affected by piperine plays a role in pain and anxiety.

We saw in the post below that TRPV1 controls cortical microglia activation and that GABARAP modulates TRPV1 expression.

So, TRPV1 and GABAA receptors are deeply intertwined.

  

GABAa receptor trafficking, Migraine, Pain, Light Sensitivity, Autophagy, Jacobsen Syndrome,Angelman Syndrome, GABARAP, TRPV1, PX-RICS, CaMKII and CGRP ... Oh and the"fever effect"



Is Piperine going to make autism better, or worse?