A new way to look at ALS (thank God)

It’s always good when a new way to look at a basically untreatable disease comes along.  We’ll know soon if looking at filamin A will be useful for Alzheimer’s disease.  Here’s another:  something we’ve known about for years (polyphosphate) may be important in Amyotrophic Lateral Sclerosis (ALS).   I used riluzole for ALS, but never saw any benefit.  It may have slowed the decline, but riluzole never stopped disease progression.

It is stated that 10% of ALS is familial, but I think this is an overstatement.  Even so mutations in a variety of proteins(superoxide dismutase 1 (SOD1) TDP43, C9orf72) do cause ALS, and studying them has taught us a lot about ALS.  There is plenty of work to do.  In 2016 a mere 160 mutations in the 153 amino acids of SOD1 had been found, but we still don’t know how they cause ALS despite hundreds of papers on the subject.  The proteins have allowed us to make mouse models of ALS, by putting in one or the other of mutated SOD1, TDP43, C9orf72 in motor neurons (or in whole animals)

Some real gumshoe work led to polyphosphate [ Neuron vol. 110 pp. 1603 – 1605 ’22 ].  Obviously in ALS, the motor neurons die, but recent work has shown that motor neurons are killed by neighboring astrocytes (containing any of the 3 the mutant proteins), when they are cultured together.   Normal astrocytes don’t do this.

So a lot of hard work found that it was polyphosphate in the supernatant fluid that was the killer.

So what is polyphosphate?  It’s been known for years, and is found in ALL cells — bacterial, plant, animal.  It also produced abiotically in volcanic exudates and deep sea steam vents.  No one knows what it does, so it has been called a molecular fossil.  Again teleology should inform biologic research (but it doesn’t).  Polyphosphate must be doing something useful or it wouldn’t be present in all living cells.

Chemically, polyphosphate is a chain of HUNDREDS to THOUSANDS of phosphate residues linked by high energy phosphoanhydride bonds.

Like this —

HO – PO2 – OH  + HO -PO2 -OH –>  HO – PO2 – 0 – PO2 – OH + H20

— the – O – in the middle is the phosphoanhydride bond

The authors treated motor neurons in culture with polyphosphate and found that it killed 40% of them.  So what?  Schmidt’s law of pharmacology, says that enough of anything will do anything,  So they looked at the spinal cords of patients dying of ALS and found that polyphosphate levels were higher than in neurologically normal controls.

So it’s open season on polyphosphate. Finding out what it does in normal cells, finding out how it kills motor neurons, finding out if decreasing its levels will help ALS (it does in cultures of motor neurons but that’s a long way from a living patient).  It’s an entirely new angle on an awful disease, with no useful treatment.  There is simply an enormous amount of work to be done.

Watch this space.

 

 

A pile of spent bullets — take II

I can tell you after being in neurology for 50 years that back in the day every microscopic inclusion found in neurologic disease was thought to be causative.  This was certainly true for the senile plaque of Alzheimer’s disease and the Lewy body of Parkinsonism.  Interestingly, the protein inclusions in ALS weren’t noticed for decades.

However there are 3 possible explanations for any microscopic change seen in any disease.  The first is that they are causative (the initial assumption).  The second is that they are a pile of spent bullets, which the neuron uses to defend itself against the real killer.  The third is they are tombstones, the final emanations of a dying cell, a marker for the cause of death rather than the cause itself.

An earlier post concerned work that implied that the visible aggregates of alpha-synuclein in Parkinson’s disease were protective rather than destructive — https://luysii.wordpress.com/2018/01/07/are-the-inclusions-found-in-neurologic-disease-attempts-at-defense-rather-then-the-cause/.

Comes now Proc. Natl. Acad. Sci. vol. 115 pp. 4661 – 4665 ’18 on Superoxide Dismutase 1 (SOD1) and ALS. Familial ALS is fortunately less common than the sporadic form (under 10% in my experience).  Mutations in SOD1 are found in the familial form.  The protein contains 153 amino acids, and as 6/16 160 different mutations in SOD1 have been found.  Since each codon can contain only 3 mutations from the wild type, this implies that, at a minimum,  53/153 codons of the protein have been mutated causing the disease.  Sadly, there is no general agreement on what the mutations actually do — impair SOD1 function, produce a new SOD1 function, cause SOD1 to bind to something else modifying that function etc. etc.  A search on Google Scholar for SOD1 and ALS produced 28,000 hits.

SOD1 exists as a soluble trimer of proteins or the fibrillar aggregate.   Knowing the structure of the trimer, the authors produced mutants which stabilized the trimer (Glycine 147 –> Proline) making aggregate formation less likely and two mutations (Asparagine 53 –> Isoleucine, and Aspartic acid 101 –> Isoleucine) which destabilized the trimer making aggregate formation more likely.  Then they threw the various mutant proteins at neuroblastoma cells and looked for toxicity.

The trimer stabilizing mutant  (Glycine 147 –> Proline) was toxic and the destabilizing mutants  (Asparagine 53 –> Isoleucine, and Aspartic acid 101 –> Isoleucine)  actually improved survival of the cells.  The trimer stabilizing mutant was actually more toxic to the cells than two naturally occurring SOD1 mutants which cause ALS in people (Alanine 4 –> Valine, Glycine 93 –> Alanine).  Clearly with these two something steric is going on.

So, in this experimental system at least, the aggregate is protective and what you can’t see (microscopically) is what kills cells.

Stephen Hawking R. I. P.

Stephen Hawking, brilliant mathematician and physicist has died.  Forget all that. He did something for my patients with motor neuron disease that I, as a neurologist, could not do.  He gave them hope.

What has chemistry done for them?  Quite a bit, but there’s so much left.

Chemistry, when successful, just becomes part of the wallpaper and ignored. All genome sequencing depends on what some chemist did.

For one spectacular example of what, without chemistry, would be impossible is Infantile Spinal Muscular Atrophy (Werdnig Hoffmann disease).  For the actual molecular biology behind it — please see — https://luysii.wordpress.com/2016/12/25/tidings-of-great-joy/.   Knowing the cause has led to not one but two specific therapies — an antisense oligonucleotide and a virus which infects neurons and actually changes the gene.

So knowing what the cause of a disease is should lead to a treatment, shouldn’t it?  Hold that thought.  Sometimes one form of motor neuron disease (amyotrophic lateral sclerosis or ALS) can be hereditary.  Find out what is being inherited to find how ALS is caused.

Well, the first protein in which a mutation is associated with familial ALS (FALS) was found exactly 25 years ago.  It is called superoxide dismutase (SOD1).  Over 150 mutations have been found in the protein associated with FALS, and yet despite literally thousands of papers on the subject we don’t know if the mutations cause a loss of function, a gain of function (and if so what that function is), an increased tendency to fold incorrectly, and on and on and on.  It’s a fascinating puzzle for the protein chemist and over the years my notes on the papers I’ve read about SOD1 have ballooned to some 25,000 words.

If you’re tired of working on SOD1, try a few of the other proteins in which mutations have been associated with FALS — Alsin, TAF15, Ubiquilin, Optineurin, TBK1 etc. etc.  The list is long.

Now it’s biology’s turn.  Motor neurons go from the spinal cord (mostly) and brain to produce muscle contraction.  Why should only this tiny (but crucial) minority of cells be affected.  The nerve fibers leave the spinal cord and travel to muscle in nerves which contain sensory nerve fibers making the same long trip, yet somehow these nerves are spared.

More than that, why should these mutations affect only these neurons, and that often after decades.  Also why should great athletes (Lou Gehrig, Ezzard Charles, etc. etc. ) get the disease.

One closing point.  Hawking shows why, in any disease median survival (when 50% of those afflicted die) is much a more meaningful statistic than average duration of survival.  Although he gave my patients great hope, they all died within a few years even as he mightily extended average survival.