Breaking the Serpent's Back

In the first months of 2020, nearly every eye was on one part of the virus. The spike protein — and within it the receptor-binding domain, the outward face that latches onto ACE2 — became the most studied structure in biology. It is the obvious target. Neutralise the part that grabs the cell and the virus cannot get in. Almost every vaccine and monoclonal antibody programme in the world went for it.

It is also the part the virus is most willing to change. The outward face is exactly where selection pressure lands hardest, and where each new variant does its most inventive work.

I kept being pulled somewhere else. Not to the fangs, but to the hinge behind them.

The search for silver bullets

What follows are collated notes on possible pharmacological interventions, written as the science was still forming — deliberately cautious hunches about mechanism, and a few bolder predictions about which compounds might matter, and why.

16 April 2020

Having floated, back on 6 February, the idea that ivermectin might prove active against SARS-CoV-2, I feel moved to share a further set of predictions about why a certain class of drugs might matter.

There is some connection between anti-parasitic mechanisms and blunting COVID-19 that isn’t clear, but seems to be stacking up: (hydroxy)chloroquine, ivermectin, methylene blue. Metronidazole, another broad-spectrum antibiotic, antifungal and antiprotozoal, has also been suggested.

Ivermectin blocks importin, which cells use to move proteins inside themselves. That makes it harder for RNA viruses to infiltrate cells, so it makes mechanistic sense that it would have antiviral properties in a dish. However, (hydroxy)chloroquine and methylene blue — both anti-protozoans — don’t appear to share this importin-related mechanism. Neither does metronidazole. Something else must be at play.

Here is what I think that something is. Cellular microtubules play a role in viral infection. SARS-CoV-2 spike proteins have been shown to interact with cytoskeleton filaments — microtubules and actin — for internalisation into host cells, a critical step in pathogenesis. I suspect that, like rabies and mouse noroviruses, SARS-CoV-2 disrupts cellular microtubules by depolymerising them.

I also suspect a link between viral depolymerisation of microtubules and the selective, preferential polymerisation of parasitic tubulin or ezrin under anthelmintic drugs. Such drugs act on non-parasitic cells too, just to a lesser degree. I posit that this modest alteration of microtubule polymerisation within host cells could change how easily the virus attacks those cells — perhaps through a hormetic eustress process.

If that is right, then the interesting drugs are not the ones that stop the virus touching the cell. They are the ones that stiffen the machinery it needs once it has arrived.

Fenbendazole is one drug well documented in altering microtubule inhibition. I predict that fenbendazole and its sister compounds (phenothiazine, thiabendazole, parbendazole, mebendazole, albendazole, febantel, cambendazole, oxibendazole, flubendazole, oxfendazole, cyclobendazole, thiophanate and triclabendazole) are worth investigating. So too the veterinary deworming agents moxidectin and selamectin, agonists of glutamate-gated chloride channels, which belong in the same enquiry as ivermectin — as do chlorinated salicylanilides such as niclosamide, oxyclozanide and rafoxanide.

Biscoclaurine alkaloids — cepharanthine, coclobine, berbamine, pendulin — are worth a look. I further wonder whether routine consumption of the alkaloids in bitter gourd, or of bitter plants bearing oleanolic acid and other bitter glycosides (marigold, gentian), or of saponin- and phenol-bearing plants such as ginseng root and eucalyptus leaf, might carry some limited value — particularly non-steroidal saponin drugs (triterpene glycosides) with oleanane ring systems. These are hypotheses about mechanism, not treatment recommendations.

It has been shown for some coronaviruses that targeting the cytoskeleton with tubulin or actin inhibitors reduces viral load in cell culture. I have a hunch that parasite tubulin or ezrin may be a useful antigen for provoking the body’s defences. And I wonder whether parts of the world with greater parasite loads might, all else equal, see less symptomatic infection — a testable ecological hypothesis, to be held loosely.

23 April 2020

Some early cohort reports suggested, counter-intuitively, that smokers might be under-represented among hospitalised patients. If there were anything to it, one candidate explanation is hormesis — a small, chronic disruption leaving a system more resilient — and another is that nicotine is itself a historical anthelmintic, used against parasites before modern drugs, which fits the pattern I keep circling. Nicotine also has neuroprotective properties.

Nicotine is highly addictive and its traditional delivery mechanisms are dangerous, so any test would need a patch; a French group was investigating exactly that. My hypothesis was that nicotine might prove useful via microtubule effects. The apparent smoker effect did not hold up; it was confounded, and some of the studies suggesting it were later discredited.

30 April 2020

Early reports were circulating that claimed large mortality reductions in hospitalised patients treated with ivermectin. The most dramatic of them were later retracted for fraud or fabricated data, so their figures are not reproduced here.

19 May 2020

A combination of ivermectin with doxycycline was being discussed on the basis of very preliminary, essentially anecdotal reports — close to hearsay even then. The interesting part is the mechanism. Doxycycline has inhibitory effects on calcium channels, and calcium transport is a significant regulator of microtubule dynamics, which is precisely the lever I had been arguing for. Doxycycline and its sister minocycline — though not tetracycline — have also been implicated in blunting hypoxic effects, hypoxia being a feature of severe COVID. Both drugs are common and cheap.

In the same period I fielded questions about traditional botanicals. Nigella sativa (black cumin) has traditional uses for dyspnoea and for clearing intestinal parasites; thymoquinone, its principal bioactive compound, acts on calcium channels. Artemisia annua is another anti-parasitic acting on ion channels. I would not have been surprised to see some in-vitro signal from either — a signal in a dish being a very different thing from a benefit in a patient.

15 July 2020

More studies on ivermectin appeared through the summer. The studies that looked most impressive proved the least sound; with the fraudulent work stripped out, the pooled clinical picture was null.

19 August 2020

Clinicians were publicly advocating ivermectin-based combinations, with doxycycline or azithromycin and zinc, sometimes in extraordinarily strong terms. The mechanistic rationale — that certain antibiotics can inhibit viral entry — is real at the bench. The extraordinary clinical claims were not borne out.

26 August 2020

The supposition that eucalyptus leaf might carry antiviral value drew interest from work at UK defence laboratories: a molecule-level finding, of the kind worth chasing in the lab and dangerous to over-read at the pharmacy.

7 December 2020

The ecological hypothesis — that regions with routine anti-parasite mass drug administration might show lower incidence — had produced some suggestive correlational papers. Correlations of this kind are notoriously confounded by climate, demography, testing capacity and age structure, and this one did not translate into clinical benefit when tested directly.

20 December 2020

Glycyrrhizin, a liquorice-plant saponin, and lycorine, a bitter alkaloid, showed activity in vitro — consistent with the saponin and alkaloid strands of the April predictions.

30 December 2020

By the year’s end the more transmissible variants had arrived, and the mood was grim. Much of this entry originally argued for mass prophylactic ivermectin as a firebreak; the clinical evidence that would have justified it never arrived. What holds up from that period is narrower: that vitamin D status is worth attending to, and that the epistemics of the moment were genuinely bad, with serious, referenced discussion of repurposed drugs often flattened by platform moderation into the same bin as quackery. That flattening was its own problem, and a separate one.

Breaking the serpent’s back

The thread running through all of the above is not really about any one drug. It is about a target.

The spike protein has an outward face and an inward one. The world went after the outward face — the fangs. The inward part, the endodomain, is the short tail that sits inside the cell membrane, and it is how the virus gets a grip on the host’s own scaffolding. The protein it reaches for is ezrin, which tethers the cell membrane to the actin cytoskeleton. Interrupt that handshake and the virus cannot leverage the machinery it needs to complete its entry. Not drawing the serpent’s fangs. Breaking its back, so that it cannot strike at all.

Ezrin appears by name in the April 2020 entry above, two years before I could do anything about it.

What I could eventually do, with my co-author Dr Selvaakumar Chellasamy, was model it. No experimental structure for the SARS-CoV-2 endodomain existed, so we built one — the first simulation of that structure — alongside its SARS-CoV-1 counterpart, and ran protein–protein docking and molecular dynamics to characterise how it binds human ezrin. A single amino-acid substitution between the two viruses turned out to change the binding pattern entirely. We then screened compounds that might occupy the pocket and hold ezrin stable against the viral tail: quercetin, minocycline, calcifediol, calcitriol, selamectin, ivermectin and ergocalciferol all docked favourably. Several of them — selamectin, the minocycline family, the vitamin D metabolites — were named in these notes before the work began.

The paper is Docking and molecular dynamics studies of human ezrin protein with a modelled SARS-CoV-2 endodomain and their interaction with potential invasion inhibitors, in the Journal of King Saud University – Science.

I funded the simulation myself, about $10,000, in a discipline I had no training in and had to learn as I went. It was the hardest research I have undertaken, done for no reason beyond public benefit in the middle of a crisis. Peer review then took over a year, which blunted whatever timeliness the findings might have had — the pandemic did not wait for the reviewers.

It got through. In a year when almost everyone was looking at the same face of the same protein, we looked at the other side of it and found something worth reporting. A molecule that docks well is a lead and not a cure, and I would not claim otherwise. But the target was novel, and it came out of this log.