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Alzheimer's Disease

Sulfur, Sulfur, Sulfur

Does anyone know of any phosphatase inhibitors that aren’t hideous? I ask this because someone sent along a question about this paper, from last August, that I’d missed at the time (press release here, but the paper’s open-access as well). Here’s a commentary in the journal itself. It’s work from Yale on an enzyme called STtriatal-Enriched tyrosine Phosphatase (STEP), a brain-specific enzyme which has been thought to be involved in memory and other CNS processes. The group was looking for an inhibitor, and screening for phosphatase inhibitors has been, historically, No Fun Whatsoever.
I’d say that this is still the case, given that the best molecule that turned up was TC-2153, shown at left. I’ve seen several molecules in this polysulfide class over the years, but I’ve never seen one that went very far. It’s not necessarily an impossible molecule, but it’s going to need some special care in development. You’d want to look out for oxidation (both on storage and metabolic), exchange reactions with other thiols, nucleophilic ring opening, and so on. The compound appears to work, in fact, by reacting with the thiol on a catalytic Cys residue in the enzyme’s active site.
This structure was arrived at by an unusual route: during the initial screen, several compounds were apparent hits. Resynthesis of them, though, gave no activity (a common experience when working with difficult targets!) Checking the original samples turned up an impurity that actually showed potent activity. This was fortunate – you often go back to these things looking for such a clue, but finding something identifiable isn’t easy. Often it turns out to be some sort of polar polymeric gunk (like this), colorful stuff that sticks to silica on a quick filtration, but in this case, it was a yellow crystalline substance that eluted late. X-ray crystallography of it showed it to be elemental sulfur, S8, which must have been an interesting surprise. Its IC50 was 17 nM! Potency aside, that’s really going to have trouble as a CNS-targeted drug, as the team well realized, so they deliberately had a look at the benzopentathiepins as surrogates. (As a side note, the authors point out that they didn’t include DTT as a reducing agent in their assay conditions – had it been there, they very likely would never have picked up on this compound class at all). It’s also worth noting that when you do find the Amazing Active Impurity, the chances are increased that it’s working through a covalent mechanism.
TC-1253 itself showed good activity in cells and in mice. But it also shows evidence of off-target effects at higher doses, as well it might, and shows some activity against related protein tyrosine phosphatases in vitro. (It looks better in cells and in the mice than it does against the straight enzymes, which might be the same sort of effect that the Cravatt group saw with covalent inhibitors as well, moving from proteins to cells).
Given these results, I’d suggest going full-on covalent for the discovery of further STEP inhibitors. Since that catalytic Cys is important, and indeed seems to be more nucleophilic than usual, why not go after it with a big array of acrylates, weak leaving groups, and so on? The tolerance for such mechanisms has increased over the years in med-chem. There are a lot of interesting Cys groups out there, and sufficiently diverse compounds should be able to target them selectively. It’s tricky work – the line between a lead compound and an assay-interfering compound is not a clean one – but what other success has anyone had against phosphatases, anyway?

16 comments on “Sulfur, Sulfur, Sulfur”

  1. Kazoochemist says:

    I recall an analogous result in the infectious disease area. Screening yielded a hit that showed reasonable antibacterial activity. Detective work by the medicinal chemist also revealed that the active substance was an S8 impurity in the screening sample. It is easy to be led down the garden path by an organic soluble substance that does no show up on many analytical methods such as NMR.

  2. Anonymous says:

    Would the inhibition of STEP lead to more pTau?

  3. Anonymous says:

    Renal tissue also expresses STEP.

  4. Anonymous says:

    looks like a cysteine oxidant

  5. julien says:

    Makes me think of anetholetrithione. Already a drug, used for hyposialy in France. It is obtained by reacting anethole with sulfur. And is used as a H2S donor in biological assay as far as I know.

  6. David Borhani says:

    Elemental sulfur (S8) inhibits STEP with a reported IC50 of 17 nM; S8 solubility in water is ~19 nM (Boulegue (1978) Phophorus and Sulfur 5:127-128; value given is 1.9(+/-0.6)x10^-8 mole S8/kg). A bit close for comfort. Any possibility for colloidal (PAINs/Shoichet) inhibition here?
    So the authors turned to TC-2153, apparently as the first and only compound other than S8 selected for testing. Amazing luck, because this compound works exceedingly well, IC50 25 nM! But, it inhibited only weakly, IC50 8 uM, when the assay was run in the presence of reducing agents, to replicate the intracellular environment.
    Nonetheless, again encouragingly, TC-2153 is apparently active in vivo, despite no provided evidence that the compound has the pharmacokinetic properties to reach the target in the brain.
    Am I just a bit too suspicious?
    P.S. – The authors apparently didn’t obtain a MS of their resynthesized TC-2153. Their reported melting point (HCl salt), 147 oC, doesn’t match the 90 oC reported by Kulikov (Mol Biol. (2011) 45(2):251-257; they did obtain satisfactory HRMS & elemental analysis for their sample).

  7. David Borhani says:

    P.P.S. – Additional data are reported here: Baguley et al. (2015) BMCL 25:1044. Not much of an SAR (maybe not surprisingly). “This study establishes that the electron deficient trifluoromethyl group contributes to potency while the amino group does not”, referring to a 3-fold decrease in the second-order inactivation rate constant when the CF3 is replaced by H.

  8. Anonymous BMS Researcher says:

    I’ll bet anetholetrithione would have a rather strong taste and smell, though I’ve never worked with the stuff…

  9. Ted says:

    “No Fun Whatsoever” indeed!
    Back when I slogged away on this front, our lead structure carried 5 negative charges at pH 7. The team leader explained that this was just fine, “we’ll optimize it as a pro-drug later.”
    It actually made the quinoline sulfonic acids look good, in retrospect.
    Continual griping guaranteed that I was in the first round of cutbacks, which left me feeling like I had grown wings.
    Sometimes I miss med. chem., but other times I remember the phosphatases…

  10. Jim says:

    Why are phosphatase inhibitors typically “hideous”?

  11. Michael says:

    Point well taken about the polysulfides generally not going far, but lets not forget about Calicheamicin!

  12. milkshaken says:

    they should test crude reaction mixtures from Willgerodt reaction in a combichem fashion…
    by the way there are some marine-derived natural compounds with this kind of insane cyclic aromatic polysulfide

  13. julien says:

    #8 : I have used anetholetrithione tablets. They do not smell. And nobody complained about it around me.

  14. Anonymous says:

    “Does anyone know a phosphatase inhibitor that isn’t hideou?”
    Why yes ! It’s called Lithium chloride

  15. Anonymous says:

    1,2,3,5,6-pentathiepanes are found in nature in shiitake mushrooms, also called lenthionine.
    If its good enough for nature its good enough as a drug candidate, so don’t let structures fool you.
    But bias is rampant in drug discovery, isnt it?

  16. Mfernflower says:

    Was this by any means based off of this anti-tumor agent found in what I believe was a sea sponge?

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