WASF3 disrupts mitochondrial respiration and may mediate exercise intolerance in myalgic encephalomyelitis/chronic fatigue syndrome

[datadragon]

Am I stupid or can you easily put together this relyvrio yourself using freely available nutritional supplements? Is it sodium phenylbutyrate?

It looks like 4-phenylbutyrate (sodium phenylbutyrate) was used to lower copper induced ER STRESS in this study (human) https://pubmed.ncbi.nlm.nih.gov/27502587/ Sodium phenylbutyrate is a sodium salt of an aromatic fatty acid, made up of an aromatic ring and butyric acid (butyrate). I had mentioned zinc and sodium butyrate earlier in studies also have known effects to lower ER Stress so they may work well to try together and are needed for numerous other functions related to ME/CFS that are additionally dysregulated. HDAC inhibitors (Butyrate is one) blocked GRP78 release by inducing its aggregation in the ER. GRP78 increases WASF3 levels. https://www.nature.com/articles/srep30406 Butyrate inhibits histone deacetylase activity (HDAC) https://www.ncbi.nlm.nih.gov/pubmed/12840228 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6333934/ TUDCA has also been mentioned perhaps in tandem with zinc.

A drug approach to target a pathway hopefully will work great in some cases as they would have an effect on all downstream areas of that specific pathway, but sometimes they cant replace all the functions of the missing nutrient as well as other pathways that may require that nutrient. A recent work previously highlighted the ability of butyrate to enhance the expression of AhR-dependent genes through its histone deacetylase inhibitor (HDACi) properties. We confirmed the butyrate-activating role at the transcriptional level on AhR-dependent genes in Caco-2 and HT-29 cell lines. Interestingly, other HDAC inhibitors that could enhance AhR-dependent gene expression were unable to mimic the butyrate dependent-activation of the AhR reporter system so butyrate worked beyond just its hdac inhibition in that case where the HDAC inhibition alone could be replaced with a drug for example. https://www.nature.com/articles/s41598-018-37019-2

 

[SlamDancin]

@datadragon I’ve been discussing HDACi’s because there’s a group buy happening for Vorinostat as an anxiety treatment. Someone crunched the math and was saying that in order to inhibit HDAC to the same degree as 50mg Vorinostat you’d need to take over 10 grams of Sodium Butyrate. Not sure about Tributynin but do you know if 4-NaSB is a more potent HDACi than standard SB?

 

[SlamDancin]

Ive been taking 450 mg of Tributynin but it might be severely underdosed relative to HDAC inhibition. Someone just bought a kg to try for anxiety

 

[SlamDancin]

So the second component is just the taurine conjugation of the UDCA bile acid, abbreviated TURSO, found in trace amounts in humans and a prominent component of bear bile @mariovitali check this out. According to Wikipedia;

“Apoptosis is largely influenced by the mitochondria. If the mitochondria are distressed, they release cytochrome C (cyC) and calcium which activate caspases to propagate a cascade of cellular mechanisms to cause apoptosis. Tauroursodeoxycholic acid prevents apoptosis with its role in the BAXpathway.[7] Tauroursodeoxycholic acid prevents BAX from being transported to the mitochondria which protects the mitochondria from perturbation and the activation of caspases.[8] Many effects of tauroursodeoxycholic acid appear to be dependent on the activation of the cell membrane receptors TGR5, S1PR2 and α5β1-Integrin.[8]

Tauroursodeoxycholic acid also acts as a chemical chaperone to help maintain the stability and correct folding of proteins.[9]”

Remains to be seen in my research whether this is superior to TUDCA

 

[datadragon]

Ive been taking 450 mg of Tributynin but it might be severely underdosed relative to HDAC inhibition. Someone just bought a kg to try for anxiety

Butyrate is a strong HDAC inhibitor, as is 4-phenylbutyric acid. https://www.nature.com/articles/s41598-018-36941-9 and https://onlinelibrary.wiley.com/doi/full/10.1002/kjm2.12376 Many HDACs are zinc-dependent enzymes which require the zinc ion for the catalytic reaction, yet another reason I keep mentioning zinc in tandem. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6009916/ Remember under inflammation/infection zinc uptake is lowered at minimum and further made less available when inflammatory cytokines are present. See this point about halfway down for a small part. https://forums.phoenixrising.me/thr...-b6-production-utilization.57030/post-2440317

Vorinostat and two other HDAC inhibitors, Belinostat and Panobinostat may help with HDAC inhibition but may not fix the other functions of butyrate or zinc is my point.
https://www.pnas.org/doi/10.1073/pnas.1801156115

 

[Murph]

WASF3 was found to be high in these patients. 9/14 had high levels.

We know very little about WASF3, there' s only 90 or so papers that even *mention* it in existence. Almost all of them are cancer studies. But I was trawling through them and lo and behold. An MECFS paper.

Bioinformation. 2011; 6(3): 120–124.
Published online 2011 Apr 22. doi: 10.6026/97320630006120
PMCID: PMC3089886
PMID: 21584188

Meta analysis of Chronic Fatigue Syndrome through integration of clinical, gene expression, SNP and proteomic data​

Vasyl Pihur,1 Somnath Datta,2 and Susmita Datta2,*


We start by constructing gene-gene association networks based on about 300 genes whose expression values vary between the groups of CFS patients (plus control). Connected components (modules) from these networks are further inspected for their predictive ability for symptom severity, genotypes of two single nucleotide polymorphisms (SNP) known to be associated with symptom severity, and intensity of the ten most discriminative protein features. We use two different network construction methods and choose the common genes identified in both for added validation. Our analysis identified eleven genes which may play important roles in certain aspects of CFS or related symptoms. In particular, the gene WASF3 (aka WAVE3) possibly regulates brain cytokines involved in the mechanism of fatigue through the p38 MAPK regulatory pathway.

I dn't see a smoknig gun here interms of connections, but it is interesting thse two small research areas intersect, and may show they are really onto something.

 

[Murph]

The top left chart here is the key one:

full paper here:

https://drive.google.com/file/d/1AFlhpoZ-kvmJDSrqSjR377EbFuiuY8AK/view

 

[Judee]

fix the other functions of butyrate or zinc is my point

So are you taking a butyrate product or just prebiotics to try to increase this? Plus zinc supplementation?

Edit: https://forums.phoenixrising.me/thr...lation-dosage-suggestions.90630/#post-2441884

 

[Violeta]

Butyrate is a strong HDAC inhibitor, as is 4-phenylbutyric acid. https://www.nature.com/articles/s41598-018-36941-9 Many HDACs are zinc-dependent enzymes which require the zinc ion for the catalytic reaction, yet another reason I keep mentioning zinc in tandem. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6009916/ Remember under inflammation/infection zinc uptake is lowered at minimum and further made less available when inflammatory cytokines are present. See this point about halfway down for a small part. https://forums.phoenixrising.me/thr...-b6-production-utilization.57030/post-2440317

Vorinostat and two other HDAC inhibitors, Belinostat and Panobinostat may help with HDAC inhibition but may not fix the other functions of butyrate or zinc is my point.
https://www.pnas.org/doi/10.1073/pnas.1801156115

I am glad you keep mentioning zinc, and along with that copper bioavailability. And now a B6 connection. Very helpful.

 

[SlamDancin]

https://pubmed.ncbi.nlm.nih.gov/21597935/

Quick pubmed search and found this…The B6 and Butyrate synergy has been shown before

 

[Dude]

https://twitter.com/i/web/status/1703732927924711698

 

[SlamDancin]

So the drug used at NIH was Salubrinal as someone mentioned and it’s quoted in the Washington Post article re: the work on WASF3.

Wiki for Salubrinal

So, how can we acquire some of this stuff? It’s experimental still

 

[datadragon]

So the drug used at NIH was Salubrinal as someone mentioned and it’s quoted in the Washington Post article re: the work I’m WASF3.

To add to the discussion, this mentions: synthetic ER stress inhibitors like 4-PhenylButyric acid (4-PBA), Sephin 1, Salubrinal and natural ER stress inhibitors like Tauroursodeoxycholic acid (TUDCA), Cordycepin, Proanthocyanidins, Crocin, Purple Rice extract and cyanidin and Caffeic Acid Phenethyl Ester (CAPE) in coffee and essentially present in honey, is an active component of propolis produced by honey bees.
https://pubmed.ncbi.nlm.nih.gov/37289403/

Sodium Butyrate https://www.sciencedirect.com/science/article/pii/S2666566222000077 and Zinc https://pubmed.ncbi.nlm.nih.gov/32549180/ can be added and were my first thought. Proanthocyanidins are in ready available supplements Pycnogenol, grape seed extract and many other natural sources. https://www.sciencedirect.com/science/article/pii/S0753332219305359

Curcumin. https://www.sciencedirect.com/science/article/abs/pii/S1043661819302981?via=ihub

Resveratrol low dose. https://onlinelibrary.wiley.com/doi/epdf/10.1002/ptr.7192 Resveratrol has dose dependent effects generally https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2990065/

Drug/synthetic inhibitor availability will have to be looked into as well when that also looks promising as mentioned in the articles so far.

 

[Violeta]

WASF3 was found to be high in these patients. 9/14 had high levels.

We know very little about WASF3, there' s only 90 or so papers that even *mention* it in existence. Almost all of them are cancer studies. But I was trawling through them and lo and behold. An MECFS paper.

Bioinformation. 2011; 6(3): 120–124.
Published online 2011 Apr 22. doi: 10.6026/97320630006120
PMCID: PMC3089886
PMID: 21584188

Meta analysis of Chronic Fatigue Syndrome through integration of clinical, gene expression, SNP and proteomic data​

Vasyl Pihur,1 Somnath Datta,2 and Susmita Datta2,*


We start by constructing gene-gene association networks based on about 300 genes whose expression values vary between the groups of CFS patients (plus control). Connected components (modules) from these networks are further inspected for their predictive ability for symptom severity, genotypes of two single nucleotide polymorphisms (SNP) known to be associated with symptom severity, and intensity of the ten most discriminative protein features. We use two different network construction methods and choose the common genes identified in both for added validation. Our analysis identified eleven genes which may play important roles in certain aspects of CFS or related symptoms. In particular, the gene WASF3 (aka WAVE3) possibly regulates brain cytokines involved in the mechanism of fatigue through the p38 MAPK regulatory pathway.

I dn't see a smoknig gun here interms of connections, but it is interesting thse two small research areas intersect, and may show they are really onto something.

@Murph, would/could this cause mental fatigue/brain fog/lack of motivation? I am trying to figure out if WASF3 causes other ME/CFS symptoms besides PEM. Because if something is the cause of ME/CFS it would most likely cause those symptoms, too.

 

[SlamDancin]

@datadragon can 4-PBA be acquired in a group buy?

 

[junkcrap50]

IV sodium phenylbutyrate or IV argnine butyrate plus valganciclovir cured EBV positive nasopharyngeal carcinoma in a majority of patients. Problem was immunopathology from tumor lysis syndrome so they abandoned the protocol >10 years ago rather than titration. Outrageous.

https://twitter.com/i/web/status/1691627741991280845

What's the risk profiles of sodium phenylburtyrate and arginine butyrate? Seems they're just providing butyrate and no different to supplemental butyric acid except in likely differences in absorbed doses. Seems like butyrate (from these two drugs, tributyrin, or other butyrate sources) + TUDCA would be easy to try.

The tweet references this study: https://ashpublications.org/blood/a...96/A-phase-1-2-trial-of-arginine-butyrate-and

A phase 1/2 trial of arginine butyrate and ganciclovir in patients with Epstein-Barr virus–associated lymphoid malignancies
Blood (2007) 109 (6): 2571–2578.
https://doi.org/10.1182/blood-2006-01-024703

Malignancies associated with latent Epstein-Barr virus (EBV) are resistant to nucleoside-type antiviral agents because the viral enzyme target of these antiviral drugs, thymidine kinase (TK), is not expressed. Short-chain fatty acids, such as butyrate, induce EBV-TK expression in latently infected B cells. As butyrate has been shown to sensitize EBV+ lymphoma cells in vitro to apoptosis induced by ganciclovir, arginine butyrate in combination with ganciclovir was administered in 15 patients with refractory EBV+ lymphoid malignancies to evaluate the drug combination for toxicity, pharmacokinetics, and clinical responses. Ganciclovir was administered twice daily at standard doses, and arginine butyrate was administered by continuous infusion in an intrapatient dose escalation, from 500 mg/(kg/day) escalating to 2000 mg/(kg/day), as tolerated, for a 21-day cycle. The MTD for arginine butyrate in combination with ganciclovir was established as 1000 mg/(kg/day). Ten of 15 patients showed significant antitumor responses, with 4 CRs and 6 PRs within one treatment cycle. Complications from rapid tumor lysis occurred in 3 patients. Reversible somnolence or stupor occurred in 3 patients at arginine butyrate doses of greater than 1000 mg/(kg/day). The combination of arginine butyrate and ganciclovir was reasonably well-tolerated and appears to have significant biologic activity in vivo in EBV+ lymphoid malignancies which are refractory to other regimens.

Adding histone deacetylase inhibitors can also possibly make latently infected EBV cells vulnerable to certain anti-virals:

For symptomatic patients (without HLH), we hypothesize that 1 potentially effective strategy includes combining ganciclovir, an antiviral agent, with drugs that can induce lytic gene expression in EBV-infected T cells (Figure 2). The latently infected EBV+ T cells are resistant to nucleoside-type antiviral agents because the drugs are not activated in these cells as the EBV-protein kinase (BGLF4) is not expressed.26 Histone deacetylase inhibitors such as vorinostat or romidepsin, proteasome inhibitors such as bortezomib, and short-chain fatty acids such as butyrate induce EBV protein kinase expression in latently infected cells, which phosphorylates ganciclovir, resulting in cell death.

source: https://ashpublications.org/blood/a...ow-I-treat-T-cell-chronic-active-Epstein-Barr

 

[Murph]

Like a lot of findings in me/cfs, we don't know for sure if the high production of wasf3 is

1. upstream and a cause of a lot of other dysregulations;
2. downstream and a negative effect of upstream problems or dysregulations
3. downstream and a useful compensation for upstream problems or dysregulations.

But the mouse studies where the high wasf3 makes them tired suggest strongly the first explanation. That's why this study is exciting.

Still, there has to be a reason why the cells are pumping out wasf3. Is it high levels of endoplasmic reticulum stress? And could that be caused by lingering infection? These papers, shared by @Forummember9922 hint that enteroviruses could cause Er stress.

https://pubmed.ncbi.nlm.nih.gov/36366584/

Enteroviruses Manipulate the Unfolded Protein Response through Multifaceted Deregulation of the Ire1-Xbp1 Pathway​

https://pubmed.ncbi.nlm.nih.gov/20070307/

Endoplasmic reticulum stress is induced and modulated by enterovirus 71​

https://www.nature.com/articles/4401833

Viruses, endoplasmic reticulum stress, and interferon responses​

So a plausible model of the disease that looks more likely now than it did before this research is this: lingering enteroviruses create chronic ER stress that creates exercise limitations. when we exercise our ERs start the "unfolded protein reponse" (an emergency repsonse when the endoplasmic retticulum starts failing) and our cells pump out wasf3. the wasf3 gets in the mitochondria and prevents us from doing glycolysis properly. we conk out.

At this point, I have a couple of questions :
1. What is the timsecale of the unfolded protein response. Does it sometimes take a while to kick in? How long does it stay activated for? i.e. could it be the cause of PEM?
2. What is the way scientists test for wasf3? How cheap will it be for labs to start looking for it? Is this a highly specialised NIH thing or can any old schmo with a pipette access a test for it?

 

[Murph]

@Murph, would/could this cause mental fatigue/brain fog/lack of motivation? I am trying to figure out if WASF3 causes other ME/CFS symptoms besides PEM. Because if something is the cause of ME/CFS it would most likely cause those symptoms, too.

Short answer is yes, definitely. If wasf3 is screwing up energy production the list of things it could make worse is very long. And if wasf3 is being caused by the unfolded protein response (an emergency state your cell goes into when the endoplasmic reticulum is under stress), then the list of possible effects in your body is even longer.

What we really need now is an exercise provocation study to look at these. IS there any sign of ER stress being elevated during or after exercise? (Or, even better, is there an existing metabolomic study that has the answer? the recent Hanson study with the 4 measurements before and after 2 bouts of exercise measured thousands of things, surely one of them would be a good proxy for ER stress and we can check that !)

 

[Murph]

wasf3 production is part of the "unfolded protein response"


Nobody seems to have looked at the link from the "unfolded protein response" and mecfs before. the two search terms combined create no hits on pubmed. I can't decide if that's because this is an exciting new avenue of enquiry or an obvious dead end.

But I did find this:

"The UPR has been most extensively studied as it relates to protein quality control in the ER, but membrane lipid metabolism appears to be equally important."

It seems like you can trigger the UPR by stuffing up lipid metabolism. And we do know that we have bad lipid metabolism; that's been a consistent finding that we've never quite known what to do with.

My great weakness is perpetual optimism but this looks like a wonderful avenue of enquiry to me. I can see it all coming together in the endoplasmic reticulum. A virus and a genetic inclination sits upstream; disrupted energy production is downstream. Maybe we'll all be relabelled as having ER-overload syndrome.

 

[datadragon]

Zinc deficiency evokes the endoplasmic reticulum (ER)-stress response https://pubmed.ncbi.nlm.nih.gov/23748779/

SOD1 as a molecular switch for initiating the homeostatic ER stress response under zinc deficiency https://pubmed.ncbi.nlm.nih.gov/24076220/

Zinc is needed for A20 aka TNF alpha-induced protein 3 TNFAIP3 that negatively regulates the itaconate shunt. The zinc finger protein A20 is a tumor necrosis factor (TNF)- and interleukin 1 (IL-1)-inducible protein that negatively regulates nuclear factor-kappa B (NF-kappaB)-dependent gene expression. The mitochondrial enzyme aconitate decarboxylase 1 (ACOD1, best known as immunoresponsive gene 1 [IRG1]) is upregulated under various inflammatory conditions and serves as a pivotal regulator of immunometabolism involved in itaconate production, macrophage polarization, inflammasome activation, and oxidative stress. Under stress conditions, especially inflammatory stimulation, the expression of ACOD1 is upregulated by macrophages, monocytes, and DCs in the innate immunity system. The expression of ACOD1 is also upregulated in the tissue under infection, The upregulated ACOD1 also acts as a feedback mechanism to regulate the activation of transcription factors. For example, lipid A induces Acod1 mRNA upregulation by activating two transcription factors, namely nuclear factor kappa B subunit 1 (NFKB1) and interferon regulatory factor 3 (IRF3), while the increased expression of ACOD1 inhibits NFKB1 and IRF3, leading to LPS tolerance. This negative feedback mechanism between ACOD1 and NFKB1 can be mediated by the deubiquitinase TNF alpha-induced protein 3 (TNFAIP3, also known as A20), which inhibits NFKB1 activation and subsequent ACOD1 expression in myeloid cells in response to LPS, TNF, or carbon monoxide (CO). Increased ACOD1 expression limits NFKB1 activation by sustaining the expression of TNFAIP3. In addition, ACOD1-mediated itaconate production leads to the expression of activating transcription factor 3 (ATF3), thereby inhibiting the translation of the NFKB inhibitor zeta (NFKBIZ) and subsequent interleukin-6 (IL6) expression https://www.sciencedirect.com/science/article/pii/S2667100X22000147 https://pubmed.ncbi.nlm.nih.gov/10385526/

In vitro studies have shown that zinc decreases NF-κB activation and its target genes, such as TNF-α and IL-1β, and increases the gene expression of A20 and PPAR-α, the two zinc finger proteins with anti-inflammatory properties. Studies have demonstrated that physiological reconstitution of zinc restrains immune activation, whereas zinc deficiency, in the setting of severe infection, provokes a systemic increase in NF-κB activation. https://pubmed.ncbi.nlm.nih.gov/28083748/

Zinc inhibits the transactivation activity of NF-κB. Zinc inhibits iNOS promoter activity. https://www.sciencedirect.com/science/article/pii/S2213231714000834

Mechanistically, several immune receptors (e.g., TLRs and IFNAR), adapter proteins (e.g., MYD88), ubiquitin ligases (e.g., A20), and transcription factors (e.g., NF-κB, IRFs, and STATs) form complex signal transduction networks to control ACOD1 expression in a context-dependent manner. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395145/

HIF1a is downstream of ACOD1, and HIF1a increases WASF3 also (see pattysmith post image on first page). So I just linked the itaconate shunt to increase in WASF3.

Also can look at this post regarding some of the many downstream effects of inflammation/infection and NLRP3 over activation related to WASF3/ER Stress. ER Stress activates the NLRP3 inflammasome. the protein encoded by the SHANK3 gene which regulates the gut barrier is regulated by zinc... and is lowered via NLRP3. Shank3 also regulates zinc uptake in the gut. https://forums.phoenixrising.me/thr...se-protein-high-dose-zinc.90738/#post-2442998 ER Stress leads to NLRP3 activation as mentioned before which also sets off those effects.

@HTester