The pandemic is over

Years ago my brother said the population is being immunized by infection.  Most infections with the current SARS-CoV-2 virus are asymptomatic.  The previous post https://luysii.wordpress.com/2024/05/02/useful-covid-information-which-should-have-been-published-2-years-ago/ contained the following information

Despite the fact that 1.3% of the Danish population  studied from 1/18 to 1/22 had a positive antibody test for SARS-CoV-2 (76,700 people) at most times, the maximum number of people hospitalized for COVID19 was 495, meaning that the infection was incredibly benign (495/76700 = .0065 an under 1% chance of hospitalization.

The statistic also means that most infections back then were asymptomatic.

Today’s Nature [ vol. 629 pp. 269 – 270 ’24 ] noted that zero of 35 volunteers could be infected with what was thought to be the current strain of the virus.  Why do such a thing, to find a virus which could be used to test vaccines against the pandemic virus.  This clearly implies that most of us are immune because of previous symptomatic or asymptomatic infection.

Also CNN reports that Astra Zenica is withdrawing its vaccine presumably because no one is using it and because other vaccines are available.

LondonCNN — 

AstraZeneca is withdrawing its highly successful coronavirus vaccine, citing the availability of a plethora of new shots that has led to a decline in demand.

The vaccine — called Vaxzevria and developed in partnership with the University of Oxford — has been one of the main Covid-19 vaccines worldwide, with more than 3 billion doses supplied since the first was administered in the United Kingdom on January 4, 2021.

Useful COVID information which should have been published 2 years ago

Useful information #1 Despite the fact that 1.3% of the Danish population  studied from 1/18 to 1/22 had a positive antibody test for SARS-CoV-2 (76,700 people) at most times, the maximum number of people hospitalized for COVID19 was 495, meaning that the infection was incredibly benign (495/76700 = .0065 an under 1% chance of hospitalization.

Useful information #2 If you were in a store using your credit card and someone else with a positive antibody test for the virus was in the same store also using a credit card and you both made the transactions within 5 minutes of each your risk of becoming positive increased by only 9% to 1.41%.

The data collection from the study ended 15 January 2022.   Yet the paper [ Proc. Natl. Acad. Sci. vol. 121 e2320194121 ’24 ] wasn’t submitted until 21 months later 23 October 2023, was accepted 24 March 2024 and finally published 27 April 2024, a lag of 27 months after data collection was complete.

This information would have been ever so much more useful if we had it January or February of 2022 instead of now.

Creepy information #1 The government had information on who had a positive antibody test and when, and was able to link this with the supposedly private credit card information about when and where they used the card.

Big brother is watching you, in this case something I approve of.

Cholesin

You wouldn’t think that there was anything more to be said about cholesterol metabolism after decades of work by med school classmate Mike Brown and a host other researchers.  But there is.

The body can synthesize cholesterol starting from scratch and Mike found out how this is inhibited when cholesterol levels get too high.  Here is a brief summary of how this happens from a recent paper [ Cell vol. 187 pp. 1685 – 1700 ’24 ]

“Cholesterol biosynthesis and uptake are tightly regu-lated through a negative feedback mechanism that senses the cellular cholesterol levels. When cells are deficient in cholesterol, SREBP2, along with its escort protein SREBP cleavage-acti- vating protein (SCAP), is transported in coat protein complex II (COPII) vesicles from the endoplasmic reticulum (ER) to the Golgi apparatus. In the Golgi, SREBP2 is sequentially cleaved by site-1 and site-2 proteases. The N-terminal domain of SREBP2, released by this cleavage, travels to the nucleus, where it func- tions as a transcription factor to enhance the expression of genes involved in cholesterol synthesis and uptake. Conversely, when cellular cholesterol levels rise, cholesterol molecules bind to SCAP, triggering its interaction with insulin-induced gene (INSIG). This interaction retains SREBP in the ER and prevents the subsequent activation of SREBP and the expression of genes involved in cholesterol metabolism”.

 

Well now you can see why this took decades to figure out.

However a recently discovered protein cholesin cuts off cholesterol synthesis when you eat and absorb cholesterol, which is much more proactive as it doesn’t wait for cholesterol levels to increase.   Cholesin is secreted into the blood by the gut when cholesterol is absorbed (secretion into the blood is what makes it a hormone).   Human cholesin contains 195 amino acids and works its magic by binding to a G Protein Coupled Receptor (GPCR) called GPCR146 which shuts off signaling by protein kinase A (PKA). This prevents  SREPB2 from turn on cholesterol synthesis (primarily in the liver).

So obviously GPCR146 and cholesin do a biochemical dance together.  Amazingly, dance is more than a metaphor, and the two proteins are coded (and entwined) on opposite strands of the same genetic locus of chromosome #7 with the code for GPCR146 on one strand inside the code for cholesin on the other.

I find this both bizarre and fantastic.  The discoveries of molecular biology never cease to amaze (me at least, and you too if your molecular biological soul isn’t completely dead).

The viruses in our brains

PNMA2 (ParaNeoplastic antigen MA2) is a protein initially found as the target of the immune response (autoantibodies) producing a nasty dementing neurologic disease (Paraneoplastic encephalitis).  The PNMA2 protein is exclusively expressed in neurons which implies that neurons are using it for something.   This is teleological thinking, usually looked down on, but always needed in molecular biology and cellular physiology.

What PNMA2 does is amazing.  It forms icosahedral viral capsids which are released from cells (in culture) as nonEnveloped capsids.  It isn’t clear if this normally happens in our  brains.    Probably it doesn’t, and when the capsid somehow gets out of the producing cell or neuron immunological hell breaks loose and autoimmune encephalitis is the result.

PNMA2 is derived from one of the long terminal repeat retrotransposons (LTR retrotransposons), viral remnants that make up 8% of the human genome (https://en.wikipedia.org/wiki/LTR_retrotransposon). This explains why it makes particles that look like viruses.  Such particles can contain RNA, so big pharma is interested in them as a way of delivering mRNA drugs.

Totally off topic but yesterday I read a paper about E. Coli DNA gyrase, an amazing enzyme which untangles DNA ( Science vol. 384 pp. 227 – 232 ’24 ).

Here is what it does.   If you’ve got some venetian blinds in your home twist it 20 or so times (keeping the ends fixed, and you have the DNA double helix, with two strands winding around each other.  Now to read or copy a single strand, you must grab both strands where you want this to happen  and pull them apart keeping the ends of the venetian blind fixed.  This immediately increases the coiling elsewhere. Since there are only 10 nucleotides/turn of the double helix, copying a gene for a 100 amino acid protein means you are removing 33 twists from the separated strands (and producing new ones elsewhere).   The cords of the venetian blind quickly become a tangled mess when this happens.  This is where DNA gyrase comes in.  It cuts both strands of the DNA double helix, holding on to the cut ends, and slides an intact double helix of the twisted DNA through the cut.   Sounds fantastic doesn’t it?  Hard to see how evolution could come up with something like this but it did.

The paper contains the following passage toward the end

“A second model based on a sign-inversion reaction wassuggested to describe introduction of (–)SC by this enzyme (28). This model proposed that the enzyme binds to a positive crossover followedby a DNA strand passage through a DNA double-strand break that results in a sign inversion.”

(28) is 28. P. O. Brown, N. R. Cozzarelli,Science206, 1081–1083 (1979).

The paper is 45 years old and has now been shown to be correct.  N. R.  Cozzarelli is my late good friend and Princeton classmate Nick, and it is very nice to see him honored here.

A few words about Nick.  Although Princeton was full of rich kids, they still had the brains to take in someone like Nick whose father was an immigrant shoemaker in Jersey City.  Nick worked his way through Princeton waiting on tables in commons (where all Freshmen ate).  I can still see the time that some rich preppie jerk gave him a hard time about the service.

Nick got his PhD at Harvard and later became a professor at Berkeley where he did his great work.  Nick later edited the Proceedings of the National Academy of Sciences (USA) for 10 years before his very untimely death over 20 years ago from Burkitt’s lymphoma.  R. I. P. Nick.

What if our most common assumption about Alzheimer’s disease is wrong?

Although the “Abeta protein aggregates cause Alzheimer’s disease” has had quite a run, it is not our most common assumption about Alzheimer’s disease.

 

Any guesses?

The assumption is hidden in the deep in the semantics of Alzheimer’s disease.  By simply naming it we are tacitly assuming that Alzheimer’s disease is just one thing.   The history of medicine is the history of splitting diagnostic categories with the passage of time due the accumulation of  causal knowledge.

Infections were characterized by the type of fever they produced before Pasteur.  Breast cancer was characterized by pathology before it was molecularly split depending which hormone receptors were present.  No one would dream of treating it all the same way today.

Yet here we have massive clinical trials of single therapies in Alzheimer’s disease because we don’t know any better.

Because people vary, in all clinical trials the responses to a given drug vary patient to patient. It is worth studying those responding best and those responding worst to a therapy in terms of the data taken on entry (MMSE, age, sex, education, pre-existing disease, smoking drinking, etc. etc.).  Such analysis might tell us something about the underlying causes in addition to predicting who will and who won’t respond in the future.

In particular, Cassava Sciences’ recent release of two years of open label Simufilam administration should be studied this way.  The second year is problematic as some dropped out, some continued to receive the Simufilam, some did not, but all patients still in the study at one year had been on the drug for a year, and their clinical data (ADAS-Cog etc. etc.) is in Cassava’s possession.

How did the group responding best differ from those responding the least.  Lindsay is too busy dealing with the slings and arrows of outrageous fortune (courtesy of Science, the Wall Street Journal etc. etc.) to climb an academic totem pole to write it up.

I am particularly interested in the 5/50 patients Lindsay reported 8/21 who likely showed a 50% improvement at 9 months. As a clinical neurologist with decades of experience with demented patients, I never saw this degree of improvement at 9 months.  How did the 5 do at one year and, if continuing on Simufilam, how did they do at two years?   What was different about the 5 as a group vs. the 45 that didn’t do as well?

To have a look at the actual data Lindsay presented back then follow this link — https://luysii.wordpress.com/2021/08/25/cassava-sciences-9-month-data-is-probably-better-than-they-realize/

Why me, O Lord, Why me?

It was very hard for my multiple sclerosis (MS) patients to understand why they were singled out for MS, given the publicity given to theories of viral causation, popular at least since I started getting seriously interested in neurology as a 3rd year medical student in 1964.  Herpes simplex (fever blisters) was a popular culprit, but we all know lots of people who’ve had them without coming down with MS.

The best explanation I could give them was of my med school classmate Marty, a Jewish kid from Pittsburgh.  Graduating in 1966 at the height of American involvement in Vietnam, all my classmates entered the service within a few years.  Marty was sent to Vietnam  as a GMO (General Medical Officer).  He was quickly sent back stateside as he developed a severe anemia.  Why?

Well malaria was endemic in Vietnam, and anyone going over received an antiMalarial as prophylaxis.  The malarial parasite does its damage by infecting red blood cells.  The antiMalarial drug he received inhibited a red cell enzyme Glucose 6 Phosphate Dehydrogenase (G6PD), essentially starving the parasites.  Marty had a partial deficiency of this enzyme.  Such deficiencies are relatively common in areas endemic for Malaria, as it is protective, just as the sickle cell trait is protective against Malaria in Africa.  A variety of other mutations in different red cell proteins arising in endemic areas are also protective (example Thalassemia in Greece, etc. etc.)

So if Marty had never been sent to Vietnam he would never have become anemic.  I’d tell my patients that they had some biochemical difference (totally unknown back then) that made them susceptible to complications of infection with a common organism.  Not very satisfying, but it was the best I could do.

In the case of another virus Epstein Barr Virus (EBV) which causes infectious mononucleosis, this explanation (50+ years later) turned out to be exactly correct.    Not only that it shows the extreme subtlety of what ‘causation’ in medicine actually means.

Not only must the unlucky people getting MS after EBV infection be different biochemically, they must be infected with a particular variant of EBV (not all EBV is the same, just as not all people or SARS-CoV-2 are the same).

That’s the view from 30,000 feet.  You can stop here but the full explanation is unsparingly technical.  It is to be found in Cell vol. 186 pp. 5675 – 5676, 5708 – 5718 ’23 ]

Here goes.

Long term control of Epstein Barr Virus is mediated by cytotoxic T lymphocytes which recognize parts of EBV proteins.  One such protein is EBNA1, and antibodies to amino acids #386 – #405 of EBNA1 cross react with amino acids #370 – #389 of a human protein called GlialCAM (for Glial Cell Adhesion Molecule) which is important in maintaining the myelin sheath around axons in the brain (MS is basically a destructive immune attack on myelin).

Such an antibody is called autoreactive, in the sense that it is reacting to a normal human protein.  Cells producing autoreactive antibodies (autoreactive cells) are normally eliminated by cytotoxic natural killer cells.  In the case of EBV specific T cells they are eliminated by natural killer cells expressing proteins NKG2C and NKG2D.  They target the autoreactive GlialCAM specific autoreactive B cells.  Some people have deletion of the gene coding for NKG2C rendering them more susceptible to MS after EBV infection.

But wait, there’s more. There are many EBV variants and some of them upregulate another human protein HLA-E by containing another protein (LMP1) which stabilizes HLA-E.  HLA-E blunts the natural killer cell attack on autoreactive GlialCAM cells.

So it’s a delicate dance of unfortunate events ‘causing’ MS.  A mild genetic defect in the human, and a genetic variant in the virus, both of which must occur for causation.

Who knew that medical causation could be so subtle.

At last a Science article about Alzheimer Rx that doesn’t trash Cassava Sciences

An article “Immunotherapies for Alzheimer’s Disease”  (Science vol. 382 pp. 1242 – 1244 ’23) avoids hype about these therapies and doesn’t trash Cassava Sciences or Simufilam (by ignoring it).   They note that “aducanumab, lecanemab, and donanemab are far from curative, but they all slow cognitive decline by ~25 to 30% over 18 months.   Continuing to avoid hype they say “reduction of brain Aβ in early symptomatic AD has a positive, but modest, clinical effect.”

They don’t ignore side effects. “ARIA-E (edema in the brain parenchyma or sulcal effusion—i.e., extravasated fluid in the leptomeninges along the sulcal spaces) and ARIA-H (hemosiderin deposits resulting from red blood cell breakdown products—i.e., microhemorrhages) (9). Although the incidence of ARIAs is increased by antibody treatment and is observed in roughly one-third of treated participants, in most individuals, it is asymptomatic.

This is similar to the rationale put out for the ‘asymptomatic hemorrhages’ seen after intravenous tissue plasminogen activator (TPA) for stroke.

Ask yourself, would you want any of them (ARIA-E, ARIA-H, asymptomatic hemorrhage) ?  I wouldn’t and the long term effects of such things are unknown.

They also deal with the cost of immunotherapy. “In the US, aducanumab, lecanemab, and donanemab each have an annual cost of $26,500, but associated imaging and monitoring means that costs for a year of treatment may exceed $75,000. ”

All in all, a very fair minded article.

But what really caught my eye was the following statement. “In cancer, many drugs are approved with modest effects on overall survival, but over time, data emerge showing exceptional responses in select individuals who are cured or are in very-long-term remission.”

This is exactly what I saw in my examination of Cassava’s data on the first 50 patients to complete 9 months of treatment on Simufilam.  Basically 5/50 had a greater than 50% improvement in their ADAS-Cog11.   This is why I’m so excited about the drug.  No antimicrobial cures all infections (because they are different).  Similarly, there is no compelling evidence that all Alzheimer’s disease is the same.

Now I am a retired clinical neurologist after 33 years of training and practice.  You never see results like this in Alzheimers disease.  I likely saw 1 demented patient a week during this time.  Although the 9 month results were not double blinded and open label, long clinical experience tells me that these results are spectacular and unprecedented (even if they only occur in 10% of patients getting Simufilam).

Here is a link to a post analyzing these results in far greater detail https://luysii.wordpress.com/2021/08/25/cassava-sciences-9-month-data-is-probably-better-than-they-realize/

Why studying the cell is like the blind men and the elephant

I’ve been reading about stress granules for over 20 years.  My notes on them contain over 60,000 characters and have my summaries of the information in over 50 papers.  They go by a lot of names — processing body, P body etc.  They are an example of phase separation in the cell, similar to other better known players such as the nucleolus.  They are formed by the cellular response to a variety of stresses — starvation, lack of oxygen, reactive oxygen species, changes in cellular pH, problems with mRNA translation into protein, etc. etc.

One protein called G3BP1 is constantly found in them, but like a lot of phase separated bodies their composition isn’t fixed and they contain lots of different proteins and RNAs which come and go from the body.  Heraclitus would have loved the stress granule, you never step into (study) the same stress granule twice. They may be a new form of matter — https://luysii.wordpress.com/2015/12/06/a-new-form-of-matter/.  They are fascinating to the chemist as their composition (stoichiometry) isn’t constant — https://luysii.wordpress.com/2022/07/20/bye-bye-stoichiometry-2/  and chemists, particularly physical chemists have spent a lot of time studying them.

Most of their components (mostly proteins and RNAs) are characterized by having multiple areas which can bind to other areas.  Protein sequences able to bind to RNA are common in them, as are areas of proteins which never settle down to a single structure, and areas of proteins with a very simple amino acid composition (low complexity domains).  So phase separated bodies are a fascinating field of study for cell biologists, molecular biologists, protein chemists, physical chemists, physicists and (not so much) organic chemists, all busily studying away about their chemical properties.

Into this mix comes a completely different way of looking at stress granules, e.g. as molecular plugs for holes arising in the cellular membranes found in lysosomes and endosomes.  This is an entirely new (and very important) function for stress granules, which hadn’t even been considered (until now).  Here, their physicality rather than their chemical nature is what matters, allowing another set of blind men to study the stress granule elephant in a completely different way.  For details please see Nature vol. 923 pp. 919 – 920, 1062 – 1069 ’23.  Given the subject matter, I find it fascinating that one Alex S. Holehouse is of the authors of 919 – 920

What you can measure isn’t always what’s important

Back in the bad old days some residents would make sure that dying patients were in electrolyte (sodium, potassium, chloride CO2) balance, even though they were irrelevant to why the patient was dying (cancer, stroke, cardiac failure, etc. etc.).  Severe electrolyte imbalance can kill.  It was basically CYA.  While ‘lytes could easily be measured they were not what was important.

Similarly, the economy is great — inflation is no longer so bad and is decreasing, unemployment is low and the gross national product is increasing.  Tell that (Bidenomics) to the 60%+ of Americans living paycheck to paycheck according to three different polls conducted this year.  Inflation rate, unemployment rate, GNP are just irrelevant numbers to them.  They see diminished purchasing power every time they buy groceries (not included in inflation), buy gas, or try to eat out.

Similarly the controversy over protein electrophoretic patterns of Simufilam and whether they have been fudged is irrelevant to the far more important question of whether it helps people with Alzheimer’s disease think.  Here the important number is their scores on cognitive tests and their functioning on the activities of daily living.  I think it does.  For an elaboration please see — https://luysii.wordpress.com/2021/08/25/cassava-sciences-9-month-data-is-probably-better-than-they-realize/

Phase separation strikes again

Phase separated droplets of protein, RNA and God knows what else have gotten everyone’s attention.  For background see after the ***.

Neurologists know that phase separated TDP43 droplets are involved in a variety of neurologic diseases.  For background see after the &&&

Now it’s everyone’s concern because a recent paper [ PNAS vol. 120 e230355120 ’23 ] shows that phase separation is involved in the construction of the pandemic virus SARS-CoV-2.

Cellular Nucleic Acid Binding Protein (CNBP) is part of the interferon generated response to RNA virus infections of our cells.  In response to infection CNBP is phosphorylated and translocates from the cytoplasm to the nucleus where it turns on the interferon beta genes.

SARS-CoV-2 evades detection by our RNA sensing pathways (a story in itself), so CNBP is retained in the cytoplasm.

CNBP has another trick up its sleeve, binding to the 3′ and 5′ long terminal repeats (LTRs) of the viral genome, competing with the viral nucleocapsid protein.  This prevents the two from liquid liquid phase separation (LLPS — another name for macromolecular phase separation) which is critical for viral replication.   Did you know this? I didn’t.  The paper gives 4 references in journals I don’t read all appearing in the last few years.

Cells and animals lacking CNBP have higher SARS-CoV-2 viral loads after infection.

****

Bye bye stoichiometry

I’m republishing this old post from 2018, to refresh my memory (and yours) about liquid liquid phase separation before writing a new post on one of the most interesting papers I’ve read in recent years.  The field has exploded since this was written.

Until recently, developments in physics basically followed earlier work by mathematicians Think relativity following Riemannian geometry by 40 years.  However in the past few decades, physicists have developed mathematical concepts before the mathematicians — think mirror symmetry which came out of string theory — https://en.wikipedia.org/wiki/Mirror_symmetry_(string_theory). You may skip the following paragraph, but here is what it meant to mathematics — from a description of a 400+ page book by Amherst College’s own David A. Cox

Mirror symmetry began when theoretical physicists made some astonishing predictions about rational curves on quintic hypersurfaces in four-dimensional projective space. Understanding the mathematics behind these predictions has been a substantial challenge. This book is the first completely comprehensive monograph on mirror symmetry, covering the original observations by the physicists through the most recent progress made to date. Subjects discussed include toric varieties, Hodge theory, Kahler geometry, moduli of stable maps, Calabi-Yau manifolds, quantum cohomology, Gromov-Witten invariants, and the mirror theorem. This title features: numerous examples worked out in detail; an appendix on mathematical physics; an exposition of the algebraic theory of Gromov-Witten invariants and quantum cohomology; and, a proof of the mirror theorem for the quintic threefold.

Similarly, advances in cellular biology have come from chemistry.  Think DNA and protein structure, enzyme analysis.  However, cell biology is now beginning to return the favor and instruct chemistry by giving it new objects to study. Think phase transitions in the cell, liquid liquid phase separation, liquid droplets, and many other names (the field is in flux) as chemists begin to explore them.  Unlike most chemical objects, they are big, or they wouldn’t have been visible microscopically, so they contain many, many more molecules than chemists are used to dealing with.

These objects do not have any sort of definite stiochiometry and are made of RNA and the proteins which bind them (and sometimes DNA).  They go by any number of names (processing bodies, stress granules, nuclear speckles, Cajal bodies, Promyelocytic leukemia bodies, germline P granules.  Recent work has shown that DNA may be compacted similarly using the linker histone [ PNAS vol.  115 pp.11964 – 11969 ’18 ]

The objects are defined essentially by looking at them.  By golly they look like liquid drops, and they fuse and separate just like drops of water.  Once this is done they are analyzed chemically to see what’s in them.  I don’t think theory can predict them now, and they were never predicted a priori as far as I know.

No chemist in their right mind would have made them to study.  For one thing they contain tens to hundreds of different molecules.  Imagine trying to get a grant to see what would happen if you threw that many different RNAs and proteins together in varying concentrations.  Physicists have worked for years on phase transitions (but usually with a single molecule — think water).  So have chemists — think crystallization.

Proteins move in and out of these bodies in seconds.  Proteins found in them do have low complexity of amino acids (mostly made of only a few of the 20), and unlike enzymes, their sequences are intrinsically disordered, so forget the key and lock and induced fit concepts for enzymes.

Are they a new form of matter?  Is there any limit to how big they can be?  Are the pathologic precipitates of neurologic disease (neurofibrillary tangles, senile plaques, Lewy bodies) similar.  There certainly are plenty of distinct proteins in the senile plaque, but they don’t look like liquid droplets.

It’s a fascinating field to study.  Although made of organic molecules, there seems to be little for the organic chemist to say, since the interactions aren’t covalent.  Time for physical chemists and polymer chemists to step up to the plate.