Retro Reviews

Sunday, 21 August 2016

Everyone’s Favourite Pathway: Kynurenine, Neurotransmitters and BH4 in ME/CFS

Summary:
- L-tryptophan is metabolised along two pathways: kynurenine (KY) and 5-HTP.
- 5-HTP preceeds neurotransmitters and inflammatory modulators serotonin and melatonin.
- Kynurenine pathway upregulation is a hallmark of inflammatory pathology.
- ROS and pro-inflammatory cytokines activate the KY-pathway via enzymes TDO and IDO.
- Neurotoxic compounds of the KY-pathway (QUIN) may exacerbate neuroinflammation.

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Scientific breakthroughs come in two flavours. The first is the bolt from the blue type, a radical advancement out of left field that takes the world of science by surprise. Einstein’s theory of general relativity comes to mind, or in medicine, the fortuitous discovery of penicillin. Yet such sudden revelations carry the risk of whipping the scientific community into a frenzy, only for excitement to deflate as the initial connection proves spurious and unreplicable. The abortive XMRV coup in ME/CFS is a prime example. Its un-discovery was a psychological blow to both researchers and patients, whom briefly thought salvation at hand.

The second type of breakthrough has a traceably longer genesis, the product of steady knowledge accumulation along one research vector. Distinctly less eye-catching but certainly not lower on revolutionary potential, it resembles a swelling tide that slowly accumulates critical mass until it finally washes away our previous ignorance.

If that analogy sounds pseudo-biblical, it is because the medical community is approaching something called the ‘kynurenine pathway’ with ever-increasing reverence.

In a sentence: the kynurenine pathway is a branching system of L-tryptophan metabolism - an essential amino acid - that may be sheltering answers relevant to this blog’s usual suspects: the closely-related spectrum of (suspected) immune neuro-inflammatory conditions of ME/CFS, Alzheimer’s, autism, and depression, but also ALS and schizophrenia.

I’ll give you the caveats up front: actual treatments could be years away. The excitement is a research buzz, not a pharmaceutical buzz. Cracking tryptophan metabolism alone is not going to ‘solve’ ME/CFS. And while the kynurenine pathway (KY-pathway) currently seems slated to take after Breaking Bad, the slow starter that ended up redefining its genre, it may still peter out like The Hobbit, the movie trilogy destined for greatness but delivering only deep disappointment and sour aftertaste.

But where there’s smoke, there’s fire. Interest in the KY-pathway in chronic illness has been building for a few years. Academics converge on it from different fields. Big names in ME/CFS, such as Columbia University’s Mady Hornig and UAB’s Jarred Younger, are hitting the KY trail. It commands the attentive eye of any ME researcher, and of late, seems to be everyone’s ‘favourite pathway’. As for myself - to borrow from The Monkees - I saw their case, and now I’m a believer.

Why? First, aberrant L-tryptophan metabolism connects seamlessly with known pieces of the ME jigsaw, in particular immune dysregulation and oxidative stress. Second, it goes on to connect a few ME pathology dots of its own. Disturbed sleep, unstable mood, cognitive problems - tryptophan metabolites are a likely culprit in all.

Hitting the KY Trail
By way of general introduction to tryptophan metabolism, it is worth noting the strangely divergent lives that tryptophan (Trp) molecules may lead. A lucky Trp (1-5%) may enter the privileged 5-HTP pathway to undergo conversion into the well-known neurotransmitters serotonin or melatonin. The vast majority, however, go down the kynurenine pathway. A proportion then ends up entering the Krebs Cycle as acetyl-CoA (thus helping to generate ATP) through a series of mind-bogglingly complex transformations. The chief contingent, however, hightails it out of body in your urine after passing through a series of fascinating but still poorly understood intermediates.

These intermediates’ implication in chronic illness pathology has led us to grasp the nasty aspects of their personalities much better than their presumed routine tasks. Obligatory name-drop: these include 3-OH-Kynurenine (3-OH-K), a free-radical generator, and the neuro-excitatory quinolinic acid (QUIN), We’ll get to know these more intimately later. When circumstances are unfavourable, they don their Sith robes with great panache.

Scientists turned their attention to Trp metabolism because something is throwing off its normal flow in a subclass of chronic illnesses. To be sure, between 95-99% of all Trp walks the KY-path even in healthy individuals. The remaining 1-5% still suffices for neurotransmitter synthesis.

But in ME/CFS, serotonin levels can be so low as to be undetectable. This is worrying, as serotonin controls a large variety of functions, including food intake, memory and learning, sexual behaviour, cardiovascular function, mood and, of special importance to ME, regulation of the HPA axis. Further, melatonin, which regulates circadian rhythm a.k.a. sleep cycles, has antioxidative effects, and is also a ‘very potent immuno-active agent’ (to quote Hornig) is synthesised from serotonin. No serotonin, no melatonin, simply put. Indeed, it is one of ME’s fundamental ironies that PWME just cannot fall asleep no matter how exhausted they are.

On the other side of the scales, 3-OH-K and QUIN levels are sharply upregulated in ME/CFS. All evidence points to lopsided, and very harmful, patterns of Trp metabolism. What could be causing this?

Happy Tryp
We’ll turn our attention first to neurotransmitter synthesis. Before any feel-good compounds can be made, free Trp must cross the blood-brain barrier into the central nervous system. The basic biochemistry that ensues is relatively simple: Trp is tied linearly to serotonin (5-HT) and melatonin (see image). This implies the initial step in the chain is critical; a deficiency in the conversion of the master compound reverberates throughout. This places the spotlight on the enzyme tryptophan hydroxylase (TPH) that converts Trp into 5-HTP.

Or does it?


Image Credit: Sengupta and Tosini, ‘The Pineal Gland’, http://photobiology.info/Tosini.html

While it may seem like TPH is the gatekeeper to the so-called 5-HTP pathway, the covert rate-limiter is actually a cofactor catchily known as tetrahydrobiopterin, or BH4 for short. The quantity of available BH4 prevents us from either falling short on neurotransmitters or experiencing an excess. Homeostasis as always, is key. BH4 is a really, really important co-enzyme central to the tale of Trp metabolism, and it pops up again in many other contexts, including metabolism of the amino acid phenylalanine and dopamine synthesis.

We’ll examine BH4 more closely later. For now, the key point is that the first transformation must come off clean. Low BH4 (or low iron, another cofactor) will impair 5-HTP synthesis. Of course, the ubiquitous SNP’s may derail 5-HTP operations (e.g. here), but for the moment, we needn’t assume that PWME uniformly carry these.

Bad Tryp
Painting in broad brushstrokes of ‘good and evil’ metabolistic pathways is an admittedly unfair characterisation. To reiterate: what I’ll go on to describe as a ‘bad Tryp’ is only so in illness. The KY pathway - whatever the exact purpose of each intermediate - has presumably no pernicious effects in healthy individuals or, at a minimum, consequences that are balanced, or countered, by other compounds.

In ME/CFS, the intermediates have gained the upper hand. This is - to current thinking - the direct consequence of immune dysregulation. Certain chemokines, in particular interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), and probably interferon-alpha (IFN-α), strongly induce an enzyme called IDO. IDO is no less than the rate-limiting step to the KY-pathway and its potential nasties. (As a side note, ROS activate an enzyme called TDO, which also triggers the KY-pathway.) From here, the adverse effects escalate rapidly.

The theory, as I understand it, is as follows. As the KY-pathway sucks in more free Trp, synthesis of 3-OH-K (free radical generator) and QUIN (neuro-excitatory) increases. Trp is funnelled away from the 5-HTP pathway, reducing production of serotonin and melatonin. By way of 3-OH-K, a further impulse is given to oxidative stress, already high as a result of the body’s pro-inflammatory state. QUIN, in turn, fuels microglial hypersensitisation, amping up the inflammatory response and creating more oxidative stress. And to top it off, BH4 supply comes under pressure. Remember: BH4 is the key to the 5-HTP chain - and it is extremely sensitive to oxidation.

The consequence is the creation of a self-reinforcing feedback loop connecting Trp metabolism to an elevated immune response and rising oxidative stress. Trp may be depleted entirely, solidifying imbalances in its metabolites. A look at the biochemistry will help flesh out this causal chain.



Image Credit: Yamada et. al, ‘Proinflammatory cytokine interferon-gamma increases induction of indoleamine 2,3-dioxygenase in monocytic cells primed with amyloid beta peptide 1-42: implications for the pathogenesis of Alzheimer's disease’, Journal of Neurochemistry, 110:3 (2009), pp. 791-800.

Failsafe Failure
Not all of these metabolites are understood to equal extent. In this blog, I am also less interested in each metabolite’s deepest secrets than the overall stabilising effect that overproduction is intended to initiate. Paradoxically, the KY-pathway harbours a failsafe mechanism that should slow down KY-entry and protect Trp from permanent depletion by actually dampening down the immune response. Yet this balancing act is clearly not working in ME/CFS, as the immune system remains chronically dysregulated in favour of pro-inflammatory cytokines, with consequent rock-bottom serotonin levels. If anything, this is a definite indication that the sheer scope of ME havoc exceeds tryptophan metabolism alone.

Here’s how the KY ‘handbrake’ is supposed to work (we think). It starts with IDO, the gatekeeper enzyme to the KY-pathway. IDO responds to both cytokines and certain reactive oxygen species (ROS) - remember oxidative stress! - to begin Trp transformation into ‘nasty’ metabolites, but only so long as Trp stocks exist. Once Trp levels fall low, IDO changes its spots to an immunosuppressant, now beginning to also exercise an anti-proliferative and apoptotic effect on T-cells. (T-cells, you may be aware, secrete cytokines such as IFN-a, IFN-y and TNF-a that co-induce the KY-pathway in the first place.) IDO is assisted in this task by, paradoxically, QUIN and 3-OH-K. Together, these ‘act in concert’ to produce an ‘additive effect’. The immune system should calm down, slowing down KY-pathway Trp metabolism, and perhaps permitting greater use of the 5-HTP pathway.

Yet two problems in ME/CFS keep the handbrake from pulling. The first is oxidative stress. Even if, theoretically, the failsafe mechanism were to succeed at slowing the immune response, rampaging ROS are still able to induce IDO. Thus, oxidants continue as a source of KY-pathway stimulation. The second problem stems from pesky, two-faced QUIN. Yes, it can seemingly act as an immunosuppressant, but its existence is also neuro-excitatory since it helps to ‘prime’ the microglia. (As you may remember from a previous post, microglia are immune system components resident in the brain. They both respond to and produce cytokines, and when called upon, they add to a pro-inflammatory immune response.) Which QUIN-effect dominates, the contra or pro-stimulatory effect?

QUINner Chaos
Medicine suspects the latter outweighs the former, to the point that QUIN is labelled an ‘excitotoxin’. QUIN is always around, but its levels are sharply upregulated during the inflammation associated with an immune response, making its effects felt more strongly. As one paper notes, elevated QUIN concentrations have been found in the CSF in several neurodegenerative diseases, and QUIN injections can cause neurodegeneration in mice. Yet despite these revelations, we know precious little about the enzymes involved in QUIN formation, nor how they, or QUIN itself, are controlled. IDO inhibitors to - presumably - block the KY-pathway and bring down QUIN in that manner are in Phase II clinical trials (source), but these seem a radical solution - akin to cutting off the nose to spite the face.

Back to BH4
So what are we to do about aberrant Trp metabolism in the absence of pharmaceutical drugs? I think this a good opportunity to reroute the discussion back to BH4. But before continuing, I would like to once more caution not to construe anything that follows as treatment advice. The below is merely informed speculation, not medical advice.

With that out of the way: BH4. As we hinted at earlier, BH4 is a true multi-tasker. It converts Trp to 5-HTP, thus kickstarting the ‘happy’ Trp-pathway. As side-jobs it transforms phenylalanine to tyrosine and then to dopa - precursor to neurotransmitter dopamine. It also helps regulate the oxidative load through its function as also cofactor to an enzyme called eNOS. eNOS creates nitric oxide (NO). If BH4 is depleted, eNOS becomes ‘uncoupled’, producing ROS rather than NO. And interestingly, BH4 itself is highly sensitive to oxidation, which transforms it from highly active BH4 to inactive BH3 and/or BH2 which, as far as I’m aware, don’t help with anything.


Busy BH4.
Image Credit: Miller, Alan L., ‘The Methylation, Neurotransmitter, and Antioxidant Connections Between Folate and Depression’, Alternative Medicine Review 13: 3 (2008), pp. 216-226.

As a true multi-tasker, BH4 is continuously in high demand even when oxidative stress is sharply reducing supply. If it is medicine’s goal is to promote the 5HTP-pathway over the KY-pathway, it makes intuitive sense to replenish BH4 stocks. BH4 supplementation is a known effective treatment for phenylketonuria and reportedly also has benefits in cardiovascular disease. Its potential in ME/CFS, however, is largely untested.

Alternatively (or perhaps concurrently), we can strive to protect BH4 against oxidation. An intriguing tie-in with Vitamin B9 (folate) exists, which has the capacity to - in the form of 5-MTHF - regenerate oxidised BH4. That’s right, 5-MTHF can convert BH2/BH3 back into BH4, replenishing supply. But there’s more. Because the chemical structures of 5-MTHF and BH4 are very similar, the eNOS enzyme accepts 5-MTHF as a substitute cofactor. 5-MTHF can thus stand in for BH4 in the event of the latter’s depletion to prevent the undesirable generation of more ROS. The biochemical applications of folate seem truly limitless, which to some extent vindicates the attention recently lavished on the MTHFR-gene in chronic illness.

Simple Vitamin C (ascorbic acid) may aid in boosting BH4, too. First, Vitamin C is a known antioxidant, and thus combats the ROS that disables BH4. Second, it is a cofactor to BH4 in the enzymic reactions that transform tryptophan, phenylalanine and tyrosine into neurotransmitter precursors. Third, and most fascinatingly, ascorbic acid directly deters BH4 oxidation. It is still uncertain whether Vit C accomplishes this by shielding BH4 or regenerating it from BH2/BH3 much like 5-MHTF does. Regardless, it appears that ‘intracellular BH4 levels are critically dependent on cellular levels of ascorbate’. This triple threat - in addition to its anti-inflammatory properties - could be why high doses of Vit C are sometimes deployed in a subclass of chronic illnesses. Vit C certainly seems to help repress Epstein-Barr Virus (EBV) - a known trigger, and possible perpetuator, of ME/CFS.

[Further, if we are to believe Prof. Martin Pall, Vit C can also revert ‘spent’ (oxidised) 5-MTDF back into 5-MTHF. I’m not sure I buy this last step - it sounds too good to be true. The evidence in the academic literature is certainly scant. An article in top-tier journal Nature did find a synergistic effect between 5-MTHF and ascorbic acid supplementation, but fell short of proposing a concrete mechanism. Pall’s theory could fit the bill.]

Finally, of course, direct intervention in the 5-HTP pathway is possible. 5-HTP itself is supplementable with established efficacy in depression and fibromyalgia. Melatonin tablets may aid sleep but not much else - and even that assertion is contested. Both amount to the most proximal form of fire-fighting in Trp metabolism, stepping in to provide the body with its immediate needs. I’m not a fan of this method , and as a matter of principle prefer intervention higher up in the causal chain, so the body can return subordinate processes to homeostasis by itself. Yet at the end of the day, so long as we feel better and don’t inflict new and different damage - does it matter?

Note 05/2021: New insights of the "metabolic trap hypothesis" take a much more informed and nuanced view of kynurenine pathway problems in MECFS. Actual science is happening!

Find the full academic text by Kashi, Phair and Davis in Diagnostics here: https://www.mdpi.com/2075-4418/9/3/82
OMF commentary is here: https://www.omf.ngo/the-ido-metabolic-trap/