Stanford Community Symposium 2018: Phair, Metabolic traps, Tryptophan trap

[FMMM1]

In theory I agree with you, but what if the only diagnostic test available at the time the samples are collected turns out to only identify a subgroup of pwME. Does that mean the rest should not be investigated?

It would be wonderful to have a diagnostic test that clearly differentiated everyone who fits the definitions of ME that include PEM from every other illness but we are a way off that yet, with only small samples tested with the different methods.

Edit: But I do agree that genetic testing using subgroups that show positive on a biomedical test would be ideal.

I think there'll be a bunch of people who have altered metabolism caused by exosome signalling and others who have another disease (potentially known but undiagnosed - NIH study etc.). If this is correct then the data will clean up once a diagnostic test is applied. Yes everyone is important - including those incorrectly diagnosed with ME.
The potential test published by Karl Morten/Cara Thomas (and others) i.e. Raman spectroscopy only seems to lack funding - as does the nano-needle. I.e. the barriers to a biomedical diagnosis appear to be funding (particularly public/government funding) rather than technology.

 

[sb4]

The fact that those with a defective IDO2 [or tyrosine] gene

Could you tell me more on the Tyrosine gene/trap?

 

[Milo]

Just wanted to drop in a note that I made from the recent IiME conference when Ron was talking about the metabolic trap.

Thank you @Andy, it is an important warning and at this point until they feel confident about their instruments, their data, and whatever is needed for good science, we need to be cautious about hypothesis even if things look good on paper.

 

[FMMM1]

Could you tell me more on the Tyrosine gene/trap?

I understand that it's the same as the tryptophan trap. I.e.
1) there's a substrate limited enzyme which functions at lower (tryptophan) levels; and
2) a second enzyme which operates at higher (tryptophan) levels.

So if you have a non-functioning copy of the second enzyme then you can't metabolise high intracellular levels of tyrosine - tyrosine trap.

Note @Milo 's words of caution - above.

 

[Ron]

in Ron's (wise) selection of very ill people.

The patients were picked by Dr. Kogelnik. I don't think Dr. Kogelnik is involved in the study anymore though.

 

[RDP]

Are all the patients tested so far infectious onset cases?

This is an interesting and important question. I'll look into it. I will, however, be greatly surprised if all our CFS patients (now n=66) acquired ME/CFS following infection because our criterion is diagnosis by a physician who sees a large number of ME/CFS patients, and uses the appropriate diagnostic criteria. These patients also cover the entire range of illness severity.

 

[RDP]

Summary: UK Biobank data does not support the idea that IDO2 DNA variation is causal of ME/CFS. Hope this helps.

As Chris points out there are many SNVs in IDO2. The IDO metabolic trap hypothesis, however, is not based on a GWAS approach to the data. The hypothesis originates from a need to explain both genetic predisposition and the existence of ME/CFS epidemics or outbreaks. I'm thinking one cannot explain epidemics with rare mutations. It seems unlikely to me that a rare combination of triggers could settle on a geographic region or a school or a hospital and encounter a large number of people with the required rare predisposing mutation(s). In this context, it's important to remember that attack rates of up to 26% are reported in such epidemics.

So the pivotal feature of IDO2 SNPs is not that there are so many of them, but that a few of them are both damaging (missense/nonsense, the gene product is non-functional) and extremely common. Indeed four of these variants have allele fractions > 0.01. It's because of this that one cannot just look at the UK Biobank Manhattan plot for IDO2 even if we set aside the thorny question of correct diagnosis. We are not asking if a particular SNP is significantly more common in the ME/CFS population than in the population as a whole. We are asking if at least one copy of the IDO2 gene in each ME/CFS patient is broken. So far, the answer is yes. (n=66 and counting)

Finally, I am not proposing that mutations in IDO2 are causal, so on this point I completely agree with Chris. The hypothesis is that damaging mutations in IDO2 predispose to ME/CFS.

 

[FMMM1]

The patients were picked by Dr. Kogelnik. I don't think Dr. Kogelnik is involved in the study anymore though.

I think Ron has spoken on a number of occasions about the decision to study the most severely ill patients (study design) - the logic is that this will provided a clearer insight into the disease compared with moderately ill people (my words).

I wouldn't be entirely surprised to find that the approach of studying the most severely ill people has been used in other diseases.

I think I'm commenting on study design rather than patient selection.

 

[Barry]

I think Ron has spoken on a number of occasions about the decision to study the most severely ill patients (study design) - the logic is that this will provided a clearer insight into the disease compared with moderately ill people (my words).

I wouldn't be entirely surprised to find that the approach of studying the most severely ill people has been used in other diseases.

I think I'm commenting on study design rather than patient selection.

Bearing in mind I'm neither scientist nor medically trained, something I have wondered is if there could also be a benefit to studying mild/moderate patients, given they are much less likely to suffer from secondary conditions, especially deconditioning. In which case would it be much more likely that the discrepancies-from-normal being observed might perhaps be purely due to the ME?

 

[Mij]

Hopefully they will study long term 'moderate' patients, 20+ years into illness so that PEM will not confused other comorbitity symptoms.

 

[Hoopoe]

As Chris points out there are many SNVs in IDO2. The IDO metabolic trap hypothesis, however, is not based on a GWAS approach to the data. The hypothesis originates from a need to explain both genetic predisposition and the existence of ME/CFS epidemics or outbreaks. I'm thinking one cannot explain epidemics with rare mutations. It seems unlikely to me that a rare combination of triggers could settle on a geographic region or a school or a hospital and encounter a large number of people with the required rare predisposing mutation(s). In this context, it's important to remember that attack rates of up to 26% are reported in such epidemics.

I have wondered whether epidemic ME is the same illness as today's ME that is mainly not epidemic.

 

[FMMM1]

Bearing in mind I'm neither scientist nor medically trained, something I have wondered is if there could also be a benefit to studying mild/moderate patients, given they are much less likely to suffer from secondary conditions, especially deconditioning. In which case would it be much more likely that the discrepancies-from-normal being observed might perhaps be purely due to the ME?

Your post reminds me of something I'd picked up on recently - not 100% related to your question!

I have a limited science training (as a technician - haven't worked in science for 15+ years). I think I've picked up that you need to ensure that your diagnostic method correctly identifies people who are severely ill and those who are moderately ill. I assume that the same applies to treatments i.e. does it work for people who are severely ill and/or those who are moderately ill/ or mild ---?

If you have a signal to noise ratio of 1 unit then the smallest true measurement you can rely on is a difference in 3 units i.e. between healthy controls and people with ME (from memory). So if your test is struggling to separate people who are moderately/mild ill from the background noise then your method is not ideal. Ron Davis has covered this on his talk on intracellular tryptophan measurement using mass spectrometry (metabolic trap) - they're struggling to get reproducible results since they are right at the limit of detection (3:1 signal:noise ratio).

Think of a toxin like lead/arsenic. If you've eaten a very large amount, i.e. in a single dose, then it's easy to diagnose lead/arsenic poisoning. If you eat a small, but still toxic, amount i.e. every day, then diagnosis will be more difficult - since you are closer to "normal". The closer you are to normal the more difficult it will be to separate you from normal.

I think the nano-needle/Raman spectroscopy potential diagnostic tests will be able to separate mild/moderately ill people from healthy controls.

Luckily those working on this are (to my mind) leading researchers: Ron Davis, Karl Morten ---.

They need funding ---

ME Action are lobbying for funding for the development of a diagnostic test and treatments https://www.meaction.net/…/announcing-millionsmissing-2019…/

 

[jamari]

Could you tell me more on the Tyrosine gene/trap?

All patients that Robert Phair examined had either a damaging mutiation in IDO2 (typtophan pathway) or tyrosine hydroxylase (tyrosine pathway).

Tyrosine hydroxylase is enzyme that catabolizes L-tyronise into L-DOPA, which then goes on to become Dopamine etc.

The facinating discovery behind this is that they are both self inhibiting enzymes which play a role in the midbrain. They both fit the trap model.

ref:

 

[Mithriel]

I have wondered whether epidemic ME is the same illness as today's ME that is mainly not epidemic.

I have exactly the disease that is described for the epidemics. Studies nearer that time showed there were also sporadic cases. The Incline Village outbreak where the invented CFS and the Lyndonville outbreak showed that epidemic ME carried on.

When forums started it was common for people to say that a few people became ill at the same time. For myself, my uncle took ill at the same time as me but his illness took a different course that did not have PEM and burnt out after about 10 years.

The change is most likely due to polio vaccination. All the enteroviruses are very closely related so it would protect against all the others as well. Having said that there have been outbreaks of other enteroviral epidemics in places like India. I do not know if this is ME as by then CFS was muddy everything up.

It is possible that everyone told they have ME nowadays does not have the same illness as epidemic ME or that may be enteroviral ME but it does exist.

Then, of course, viruses shift their makeup and disease expression becomes different, like scarlet fever which became rare for decades but has now returned.

 

[mariovitali]

cc : @wigglethemouse

I found an interesting connection of Quinolinc acid which is a product of kynurenine pathway with excitotoxicity but also impaired cholinergic signalling which is related to the vagus nerve and the work of Petter Brodin at Karolinska. Excitotoxicity and the NMDA becomes a major target for further Research

Quinolinic acid was identified by Machine Learning (see snapshots). Dr Phair and Janet Dafoe have been e-mailed with more information :

https://www.ncbi.nlm.nih.gov/pubmed/2939936

 

[Robert 1973]

I’ve been very slow to catch up with this thread but I have some questions for @RDP or anyone else who might be able to answer.

My understanding is that the Metabolic Trap Theory predicts a stable (rather than progressive) illness state, which appears to be consistent with most (but not all) people’s experience of ME. The theory seems to predict that once the trap is triggered and IDO 1 is inhibited, the levels of tryptophan will get higher and higher (due to a vicious cycle). Given that increasing levels of tryptophan are predicted to lead to a stable rather than progressive illness state, am I correct in thinking that the levels of tryptophan themselves are not assumed to have any effect on symptoms once the IDO 1 inhibiting threshold has been crossed?

Is there known/assumed to be any upper limit on levels of intracellular tryptophan? If not, does the theory predict that there will be a positive correlation between the levels of intracellular tryptophan and illness duration? Has any such correlation been found?

Finally, does the metabolic trap theory offer any explanation as to why there is such diversity of severity of illness among ME patients? If so, what is the explanation? Is the suggestion that illness severity could be determined by the degree of IDO 1 inhibition? If so, what would determine the degree of inhibition? Or is the assumption that illness severity may be determined by other factors once the trap has been triggered?

 

[Barry]

My understanding is that the Metabolic Trap Theory predicts a stable (rather than progressive) illness state, which appears to be consistent with most (but not all) people’s experience of ME.

...

Finally, does the metabolic trap theory offer any explanation as to why there is such diversity of severity of illness among ME patients?

Is that necessarily the case? Are you sure any "stability" does not just mean firmly locked into the trapped unhealthy state? Which would be distinct from the degree of illness when locked into that state. Please note I'm not speaking from a position of deep knowledge or understanding here, but only that I can appreciate that stability of a state is not the same as the stability of an illness that might be provoke by being in that state. (ETA: If that comes across as sarcasm I promise it is not).

Edited for clarity.

 

[Jim001]

I dont have any technical knowledge about this, but I have wondered if the severity of ones illness could in part have to do with the number of entrapped cells, and not just the degree of entrapment at the cellular level, since the metabolic trap seems to be a cellular phenomenon rather than a systems phenomenon.

 

[RDP]

My understanding is that the Metabolic Trap Theory predicts a stable (rather than progressive) illness state, which appears to be consistent with most (but not all) people’s experience of ME. The theory seems to predict that once the trap is triggered and IDO 1 is inhibited, the levels of tryptophan will get higher and higher (due to a vicious cycle). Given that increasing levels of tryptophan are predicted to lead to a stable rather than progressive illness state, am I correct in thinking that the levels of tryptophan themselves are not assumed to have any effect on symptoms once the IDO 1 inhibiting threshold has been crossed?

To answer @Robert1973, your post includes many questions. I'll do my best. The IDO metabolic trap hypothesis does predict a stable steady state after cellular tryptophan (Trp) has increased to the point where cellular uptake of Trp equals the sum of degradation (by the kynurenine pathway) and conversion (to 5-hydroxytryptophan and serotonin). But we don't know how long that approach to the new (pathological) steady state will take. This will depend on the exact characteristics of each patient's membrane transporters and enzymes. Moreover, as @Jim001 notes above, there is always the possibility that more and more cells of a given cell type are becoming trapped. This too would yield a progressive phenotype. There are thought to be on the order of 10,000 serotonergic neurons in the mid-brain Raphe. You can imagine how a binary untrapped/trapped state for these cells could translate to a continuum of disease severity having 10,000 different levels. I think this is the same as what @Jim001 has in mind.

To your second point, increasing cellular Trp might well have disease consequences because it provides a huge mass-action drive toward serotonin over-production in cells expressing the brain isoform of TPH2, and a substrate-inhibition-mediated under-production of serotonin in the gut and the pineal where the "peripheral" isoform, TPH1, is expressed.

 

[RDP]

Is there known/assumed to be any upper limit on levels of intracellular tryptophan? If not, does the theory predict that there will be a positive correlation between the levels of intracellular tryptophan and illness duration? Has any such correlation been found?

An upper limit on cellular Trp? Certainly there is the limit of solubility, but below that the level achieved in any cell will depend on the kinetic properties of the LAT1 transporter bringing Trp into the cell and the kinetic properties of the enzymes that consume Trp. An amazing feature of LAT1 is that is exhibits kinetic asymmetry - the Km for Trp uptake is 1000-fold less than the Km for Trp export. I don't know for sure yet, but I think this plays a role in allowing very high intracellular Trp. Measuring intracellular Trp is not as easy as you might think, especially when you want to measure it in a sub population of cells making up only 1% of the total number of cells. ME/CFS research, especially metabolomics, has been so focused on measurements in plasma that we tend to forget the problems you encounter when you want to know what's happening in the cytosol of a given cell type. If you've followed my talks, you've probably sensed my chagrin that it's taken us so long to make this measurement. I promise you I am at least as frustrated by this as you are. Remember we have to measure kynurenine too and that's 10-fold lower in concentration even in normal cells. I think we are getting close. What we need is biochemical evidence of the trap being sprung. It's always possible that plasmacytoid dendritic cells are not in the trap even though serotonergic neurons ARE in the trap. This is why some critics think we are wasting precious time on dendritic cells and we should instead be measuring these metabolites in serotonergic nerves in the midbrain. Unfortunately, I don't know how to do that either - yet. There's a common story in physics research describing theoretical physicists flying around the world trying to get an experimental physicist to make a particular measurement. I've been on both sides of the experimental/theoretical fence in biology. Both are hard - sometimes, really hard. Once we can measure these things with accuracy and precision, correlations will become easy to plot.