Strap on your hardhat, we are going into the factory

• Once on the big warehouse floor, the protein synthesis is ready to happen. It all starts at the Ribosome, the protein making machine for the cell (of which there are millions inside a cell). These little machines are made of two subunits, small and large, and staple two types of RNA together. The first is the mRNA blueprint information and the other is transfer RNA (tRNA) which has the amino acid raw material.
  • Reading 3 codons at a time, the ribosome reads the mRNA and fits the corresponding tRNA together. It then goes clunk and spits out the old tRNA to get a new one. The result is a chain of amino acids strung together like popcorn strings at Christmas. With this simple set up of connecting the blueprint and raw material we can create small integral-for-life proteins like Thyroid Releasing Hormone (TRH) which is 234 amino acids long or Titin, the largest protein in the human body, at about 35,000 amino acids to help keep our muscles springy.

• Sticking raw materials together isn’t enough to make a product, some assembly is required. In this case we can define four levels of protein structure:
  • Primary Protein Structure refers to the physical sequence of the amino acids. This specific sequence is what is going to lend stability and flexibility to the later levels. Mutations at this level can be catastrophic or completely harmless.
  • Secondary Protein Structure is how the amino acid sequence folds in on itself. Like a piece of string you let go of, it will land and loop and knot around itself. Another way to look at this is if the primary structure is letters, the secondary is words. How you order the letters is what makes words which when done correctly make coherent words.
  • Tertiary Protein Structure is now a completed protein with a three dimensional shape. At this point the words have come together to make a sentence that has proper grammar and sense. If the protein is meant to be a monomer, it would be completely functional at this stage and can be whisked off to where it belongs. Else, it goes to the next stage.
  • Quaternary Protein Structure is our final structure and most complex. Here many monomer proteins are stuck together to make a larger, more complex 3D shape. We have taken our sentences to make paragraphs or blog posts. This is also what constitutes many of the cellular components that are necessary for complex multicellular life, like Hemoglobin which is made of 4 globin proteins or Insulin which is made up of two chains glued together at both ends.

• This whole folding business is all well and good but we need workers in the factory that actually facilitate the creation of more complex proteins. In the cell this is accomplished by Chaperone Proteins which help proteins fold in the correct orientation to ensure function. Without them the lanky amino acid sequence could fold incorrectly due to changes in the cellular environment, especially heat and pH. This is best accomplished as the protein comes out of the ribosome so that there is little time between creation and folding.
  • For the most part the process handles itself. By a clever bit of evolutionary magic both the chaperone and baby protein prefer interaction rather than dissociation. Because of this, the conformation of binding, folding, and then releasing the protein is a favored process rather than something requiring enormous amounts of energy. In the drawing above you can see how the chaperone captures the unfolded protein, gives it a big ol’ hug, and lets it go in the correct conformation.
  • Sometimes however the protein escapes chaperone involvement or there is a decrease in chaperone synthesis. The result is a collection of unfolded proteins collecting together in an Aggregate. It’s this issue that is the basis of our post today.

“Lots of Love and Lots of Patience”

Life before the diagnosis was just a normal life, doing groceries, reading a lot, making the house in order, and sending birthday cards to people. Playing golf with friends and the occasional dinner, vacations etc. She deteriorated week by week. It all started with dizziness, then she fell, received a walker, and then eventually a wheelchair. She became more anxious and would see things in the mirror that wouldn’t immediately recognize (she’d see herself but it wouldn’t register which would startle her.) The stairlift helped with getting around the house. Eventually after six months we decided that hospice care was the best place to take care of her.

She’d forget things. First the simple things like making sandwiches, coffee, house cleaning. Then she became less mobile and I would help her to the bathroom. Eventually she’d not understand left from right and you’d have to be patient and steer her in the right direction. A simple bathroom trip could take 30 minutes and lunch would take 45. All the while trying to keep her focused on the tasks. We had a whole system: take the doorknob with your hand, move in the direction while pointing, walk behind her while half carrying her under the armpits in the direction she’d have to go.

• Let’s say you travel to a far off land and stumble across some locals who are exceptionally kind and you are sharing dinner with them. As you are plied with drinks you can’t pronounce, the food starts rolling out of the kitchen—plates piled high with flat pocketed breads, tangy and spicy sauces, and mountains of grilled vegetables you think you recognize from old episodes of Anthony Bourdain. Stealing the show however is the great pot of meat stew that is placed in front of you. The scent of warm spices touch your nose and you can’t help yourself from indulging in the meaty heaven (vegetarians and vegans play along for now). Your guide returns from booking the hotel and asks you how you like the meal. You nod your approval and ask what the meat is. He says sheep brain. You chew slower.
  • Dramatization aside, the first disease we will discuss is Creutzfeldt-Jakob disease (krutz-feld yay-kob) which is a fatal neurodegenerative disorder. CJD is part of a larger group of diseases called Spongiform Encephalopathies which get their name from the sponge-like texture of the brain due to the causative agent: Prions. Unlike other transmissible diseases caused by living things like bacteria or fungi, Prions are not alive (the same with viruses). Prions are just misfolded proteins that cause other nearby proteins to misfold. In this case the Prion misfolded protein enters the body and is taken up into a cell where it causes other proteins to misfold. This has two effects: the first is the creation of exponentially more Prions as each bad protein misfolds more and more native proteins. The second is prevention of digestion from lysosomes and proteasomes to degrade the misfolded proteins. Eventually the cell dies and bursts, releasing all those proteins to other cells.

The dementia would get worse each week and she developed tremors that looked like Parkinson’s. After 6 months, moments like: “beautiful weather outside right? Yes mom it is.” “Beautiful weather outside right? Yes mom it is.” “Beautiful weather outside right? Yes mom it is.” “Beautiful weather outside right? Oh I think I asked that before. That’s okay mom, yes, the weather is very nice.” After a few months she didn’t want or couldn’t hug me anymore; she didn’t really want to be touched. This disease is one of the most destructive things—it leaves the person without dignity. Something is lost every single week and you mourn while this disease is eating away the person you once knew. This was during the lockdowns so I had to isolate myself from my friends and family but I went to her so much. I asked her everything I ever wanted to know and I'm glad I did. The more the disease progressed, the less communication you’d have. And it got so hard…

• Creutzfeldt-Jakob Disease is caused by the transmission of infectious proteins (the prion) due to ingestion of contaminated products or by having contaminated body parts transplanted OR, in Ruth’s case, her doctor’s think that an immune reaction led to an initial misfolding that spiraled into spontaneous CJD. In some instances, human corneal and skin grafts have resulted in CJD but nowadays we screen for these complications way before transplant. More common is ingesting animals infected with prions like in the case of Bovine Spongiform Encephalopathy or more commonly known as Mad Cow Disease. Mad Cow is an incurable and fatal neurodegeneration where the cow slowly loses muscle control, loses weight rapidly from lack of eating, and then eventual delirium. It generally takes a few weeks to a few months for Mad Cow to be evident.
  • It’s thought that cattle contract Mad Cow from being fed meat-and-bone meal that contains the meat of other cows that spontaneously developed the disease or from another animal species that had prions. Prion testing in the meat industry is mandatory but the tests can be… wonky. The EU mandates that all cattle be tested at 30 months but most are slaughtered at 2-2.5 years old. Japan tests all cattle at time of slaughter but prion tests are high for false negatives. The only sure-fire way of knowing if a cow is infected is to perform a necropsy on the brain and search for the characteristic sponge texture. Before I scare you off from eating meat ever again, know that the rate of Mad Cow to human transmission is 1 case per 1 million people (which is still kind of high).

She didn’t have any “physical” pain but the brain told her there was pain when she got touched. She received an IV morphine box to help manage it. CJD patients do not like to be touched, it overloads their system so the morphine worked pretty well. When she was at home she fell and we tried to get her admitted to the hospital since we could not take care of her at home but we were sent home. The next day daily home care came twice a day to help clean but when she fell for a second time we knew we had to move her bedroom to the living room with 24/7 care. Suddenly she got mean to the 24/7 staff which is not her nature. I suspected she might have a bladder infection which turned out to be true and the cause of her new delirium. She didn’t get antibiotics in the hospital but at this point we were moving her to hospice and she received them. She got “better” with the medication and her sweet self again. In hospice she got Haloperidol (Haldol, a neuroleptic with multiple uses) but she got way too drowsy so we told her to stop giving it. We could see the next day that she was happy the stuff was stopped—she could communicate (a few words or eye movements) and looked happier.

• In the image below you can see how the brain’s anatomy changes due to the disease. The first change is in the number of wrinkles in the brain which increase the total amount of surface area and thus more neuronal density. As CJD progresses, the wrinkles in the brain start to decrease which explains the confusion and decrease in brain function a person experiences. You'll also notice in the middle of the brains the hollow spaces called ventricles. Normally these tight fluid spaces carry cerebrospinal fluid that helps nutrients flow to neurons and toxins flow out of the brain. As the neural matter decreases, the ventricle space becomes bigger which again contributes to a smaller surface area.

• Again to reiterate, Prion diseases are all part of the Spongiform Encephalopathy family but where they attack is what dictates the specific disease we call them. Cattle are not the only animals that can spontaneously develop prion diseases or transmit them to each other. Chronic Wasting Disease also known as Zombie Deer Disease is a rare genetic disorder that naturally occurs in Ungulates (deer, moose, elk, etc). It is characterized by weakness and repetitive walking patterns despite the presence of predators. Luckily it can’t be spread to humans but it can be spread to other deer.

• The other not so common but definitely higher way of contracting CJD is cannibalism. Among the Fore people of Papua New Guinea is a disease called Kuru which is a form of Spongiform Encephalopathy. While CJD is a spontaneous prion disease, Kuru is characterized by getting prions due to cannibalism rather than bad luck. Kuru is characterized by shaking (of which it gets its name) and for having victims laughing hysterically and going quiet just as suddenly. It is believed that Kuru spread among the Fore tribe due to ritualistic funerary cannibalism in which recently dead family members would be cooked and eaten. When the brain and eyes are subjected to high temperatures, prion formation is rampant and concentrates which would infect tribal members.
  • When humans are discovered to have a genetic disorder it is referred to as Fatal Familial Insomnia. The disease has 4 stages—it starts with insomnia that becomes worse and worse and eventually sleep deprivation leads to hallucinations and panic attacks lasting months. At about 8 months the person is completely unable to sleep and needs anesthesia in order to attain unconsciousness. Three months later the onset of dementia wipes the brain of its memories until the person forgets how to talk and eventually forgets how to breathe; death occurs after six months of dementia.
    • The average age of onset is about 50 years of age but there was a case of a 12 year old boy contracting the disease and dying a year later. Since it is a genetic disease, the hunt for the causative gene was performed and discovered in the early 2000s. On chromosome 20 they found that two amino acid changes (Methionine to Asparagine in position 129 and Aspartic Acid to Asparagine in position 178) cause the fatal disease.

Some final thoughts from Jane and Nicolette

You can read up all you want about this disease but each and every case is different and you cannot prepare for it. What you need to do is watch closely…observe. Try to guess what the needs are because you can never be prepared for what comes next. Read up on all insurances, social workers, churches, and anyone that can help with care taking or even just have a cup of tea with the sick person. Stay close with your family doctor. Make a dossier with all the phone numbers you can find and the information of the person it's about. Document what happens and follow up the next day when you order things because stuff can and will go wrong. Call the cjdfoundation.org although Reddit is actually the only place I found some useful information. Try and find a professional somewhere who has dealt with this disease before that you can call but this is difficult to find. Ask the ones caring for them if they have experience with the disease. If not, take the time to explain it. Either way, always give them this literature from the CJD Foundation. Don't forget to take time for yourself even though it seems impossible.
If you know someone who comes down with CJD, don’t say “is there anything I can do?” People will just say no thank you. Be more concrete—focus on their needs. When you go to the supermarket, offer to pick something up. Come over and help with cleaning or an errand. Someone with CJD is often unable to do long visits, so keeping it short like 15 minutes is all that’s needed.
Spend every second with your loved ones, especially at this point you can still get answers to your questions.You can never know what happens next, so just observe and be patient and loving. Lots of love. Lots and lots of patience, you will need it. Take breaks. All I keep thinking now is “FUCK my mom was brave” when she was going through this whole thing.

A Mind is a Wonderful Thing to Taste

• Without repeating the same story over and over again, as far as we can tell, if an animal or human contracts a prion disease they will die; the only question is how long it will take. So the big question becomes why are prions so fatal? Above you see two Pontiac G6s. My question for you is which one is broken? Just by looking at the structure we can’t tell and if enough poorly functioning cars are sold, eventually you get people crashing and hopefully a recall. In a sense, we can only prevent a problem ahead of time if the processes that identify problems work properly before the car hits asphalt. It's no different with prions.

• Here you can see the structure of the protein normally responsible for misfolding into a prion. Fittingly the gene that encodes it is called Prion Protein (PRNP) gene and it encodes for the production of the major Prion Protein (PrP). The Prion Protein is produced all throughout the nervous system which is why it accumulates in the brain and attracts other non-nervous system prions to itself (like a batman signal). The Prion Protein can exist in many different isoforms or structural forms. The normal Prion Protein is denoted PrP(c) where C stands for normal Cellular. In the picture of PrP(c) you can see some twisty alpha-helix sections that are created by five octapeptide sequences (total of 40 amino acids). This specific sequence allows for the generation of a copper binding domain that is extremely pH dependent.
  • When the folding of PrP messes up it turns into a disease-causing mis-isoform. Now, proteins fuck—I mean mess up—folding all the time and that’s where those degradation organelles come in to destroy the bad protein. This would be the safety checks testing the car before it gets sold to the customer. There is one bad isoform that is resistant to the degradation: PrP(Sc) where Sc stands for Scrapie (the prion disease that affects Sheep). PrP(Sc) has the same sequence (primary structure) but its secondary and tertiary structure is muddled, turning out nice alpha helices into a hybrid helix-flat sheet. This mixed character resists the Proteasomes that break it down and slowly it starts to propagate misfolding in other proteins.
  • It is thought that PrP plays a major role in the synapse ability of neurons. PrP(c) is present in the entire neuron cell signaling compartments and its ability to hold onto copper allows for neurons to correctly communicate with each other. In this sense, the PrP(c) acts as a homeostasis protein, or in other words, it promotes returning cells from extremes to normal functioning. Loss of PrP(c) functionality causes changes in neuronal talking and neurotransmission which ultimately disrupts the brain's ability to coordinate between structures, make new connections, and recall memories.
• Spongiform Encephalopathies, and especially CJD, are scary and devastating diseases and I think I should lay down the etiology, or cause, of prion diseases to at least offer some solace. Of CJD, you have a 1% chance of acquiring the prion disease by ingesting infected beef or while getting surgery. About 10 to 15% of CJD is familial and is due to various gene mutations that affect the Prion Protein gene. About 85% is sporadic, meaning we don’t have a clear cause, like in Ruth’s case (again, her doctor’s think it was due to an immune response). Okay that’s not really comforting is it? Well try to hang on to the idea that 1 in 1 million people will get CJD annually… hey at least you’re more likely to find a four leaf clover (1:10,000 clovers) or getting food poisoning (1:6). :D

Alzheimers is similar but follows a different etiology

• Alzheimer’s incidence (rate of acquisition) and prevalence (number of individuals) increases with age and follows a pretty consistent pattern. From ages 65 to 74 years old, the rate is 400 per 100,000, the next 10 years of age is 3200 per 100,000 and finally ages 85 and older have a 7600 in 100,000 rate of acquiring Alzheimer’s. Early onset Alzheimer’s (<65yo) accounts for 10% of all AD cases. It affects women more than men at a rate of 1.5:1.
  • We divide Alheimer’s symptoms into two groups: cognitive and noncognitive. The cognitive symptoms are the ones we expect—short term memory impairment that progresses into long term memory impairment although procedural memory is usually intact until the end (procedural memory is things we know implicitly like walking, riding a bike, playing an instrument, etc.). One of the first signs is language impairment that first starts as naming impairments (like forgetting a noun) then impaired comprehension and finally a decrease in fluency. Often Alzheimer’s patients will lose their sense of time or where they are. This is a common reason why Alzheimer’s patients get lost from their nursing homes; they leave the building and try to get home because they forget they are in a new home.
    • Noncognitively, Alzheimer’s patients can develop other symptoms that characterize what it is like to work with these patients. Behavioral changes such as apathy and irritability are common as the person becomes frustrated with failing to remember things. Often depression can develop especially as the person feels socially isolated even though their loved ones visit often. Eventually as the brain forgets more and more the person can lose the ability to hold their urine and may hallucinate. Generally those with mild to moderate Alzheimer’s are able to maintain a social facade and can preserve skills like bathing and dressing.

• With such an insidious disease, what causes the symptoms we see? Macroscopically we see changes in the structure of the brain. Above you can see a normal, healthy brain (A is a side view while B is a crosssection) and C and D is an Alzheimer’s patient. You can see in Alzheimer’s that the brain tissue starts to Atrophy or lose neuronal volume. Damage to the hippocampus (D) are the earliest changes we can see in the brain since the hippocampus is responsible for forming and retaining episodic and semantic memories. Likewise we see a decrease in surface area of the sulci (sul-ki; the valleys) and the cortex which means there is less surface area i.e. decreased neurons. We also see the development of ventricles that are normally absent. These ventricles are hollow spaces inside the brain that normally carry cerebrospinal fluid and nutrients but in Alzheimer’s they represent an internal decrease in neuronal density.
  • Microscopically we see four things almost simultaneously. First a decrease in cholinergic neurons inside the nucleus basalis of Meynert. One older hypothesis of Alzheimer’s is that a decrease in the neurotransmission of acetylcholine is what causes Alzheimer’s progression. The more modern theories of Alzheimer’s are called the Tau hypothesis and the beta-Amyloid hypothesis. The Tau hypothesis states that there are intracellular protein tangles of the Tau Protein that we think is responsible for causing progressive damage in neurons. These tangles are made up of insoluble proteins that become wrapped around inner-cell structures.

• The beta-Amyloid theory (which I personally subscribe to) states that a protein called the Amyloid Precursor Protein (APP) is responsible for the production of Senile Plaques that clump together causing neurotoxic damage. Normally APP is cleaved in two parts: first alpha-Secretase cleaves off the alpha portion of APP (the tip and tail) and then gamma-Secretase takes off the back end leaving a soluble protein to be degraded. In Alzheimer’s it is thought that a different enzyme cleaves only the tail off of the Amyloid protein leaving the beta-Amyloid insoluble protein that starts to clump together. These clumps hide in the synapses of the neuron and block transmission which leads to damage.

• The treatment of Alzheimer's is still early in development but there are some good treatments that are seeing good(ish) results. The first treatment is to replete the acetylcholine that is decreased due to neuronal loss. The first drugs used were the Acetylcholinesterase Inhibitors which are drugs that stopped the degradation of acetylcholine. The first drug we will look at is Galantamine (Razadyne) which hit the market in the 1950s in the Soviet Union. Galantamine was successfully extracted commercially from the Common Snowdrop (Galanthus nivalis) and Red Spider Lily (Lycoris radiata) in 1956 by Bulgarian chemist Dimitar Paskov. The drug was extremely hard to extract (only about 0.1% of Galantamine could actually be isolated) making large scale cultivation necessary. It was marketed as Nivalin in 1959 but became a lost medication due to the instability of the Soviet Union during this period.

• In 1985 another acetylcholinesterase hit the market: Rivastigmine (Exelon) that proved effective in increasing brain levels of acetylcholine. Originally formulated orally, a transdermal patch reduced side effects significantly and is almost exclusively used nowadays. Due to efficacy in Alzheimer’s it caused researchers to rediscover Galantamine and reintroduce it in 1992. It was Jannsen that discovered a more marketable synthetic process of making Galantamine that avoided using the daffodil flowers. Eventually another drug Donepezil (Aricept) hit the market in 1996 as another transdermal patch that treated Alzheimer’s. Today Donepezil is the 120th most common medication with over 5 million annual prescriptions. Donepezil is being investigated in ADHD as well to reduce the symptoms of that disease (although the data is middling).
• The ability to block acetylcholine destruction allows for the brain to have more neurotransmitters than it would otherwise have access to. Although these drugs don’t preserve the number of cholinergic neurons they do attempt to preserve the amount of acetylcholine that they would be producing. A second thought is that another receptor found on cholinergic neurons, NMDA, was allowing too much Calcium into the neuron. Calcium coordinates the release of neurotransmitters but it's known that too much Calcium use inside the neurons can be neurotoxic. The use of NMDA Receptor Antagonists like Memantine (Namenda) or Riluzole are paired with acetylcholinesterase inhibitors to boost the effects of either.
• If you weren’t aware, you are living in the next age of pharmacy in which previously incurable diseases are slowly finding treatments or cures. The magic here are monoclonal antibodies or mabs which have been in use for over a decade (despite the news claiming COVID birthed them). mabs are a form of synthetic antibody that we can inject into the body to seek and destroy specific structures, molecules, or cells (like cancer). Because of that mabs have incredibly specificity and low cross reactivity and new novel mabs have very few side effects. If you’re interested in a deep dive of these life saving meds, check out my post here!

• One of the big silent breakthroughs of Alzheimer’s disease is the development of a ‘vaccine’ against the beta-amyloid plaques that form. The vaccine Adalimumab (Aduhelm) is a humanized antibody that targets these aggregate plaques and alerts the immune system to DANGER DANGER! WEE WOO WEE WOO!!! causing the immune system to slurp up the plaques. Adalimumab is usually well tolerated with dizziness and nausea being common side effects but it does have a common side effect of edema (swelling) in the brain which can lead to massive headaches. Don’t forget too that the amyloid protein is part of the lining of blood vessels. Normal amyloid proteins can be flagged as bad and removed by the drug which leads to microhemorrhage.
  • Aduhelm entered phase I testing in 2015 as the first new drug to be indicated for Alzheimer’s since 2003. Passing human safety phase I testing, it went through phase II and phase III testing where it failed to meet its primary endpoint of slowing or halting Alzheimer’s progression. At the end of 2019 Biogen, the company developing Aduhelm, announced it would be restarting FDA approval after analyzing a larger dataset that showed evidence of slowing Alzheimer’s progression. In the EMERGE trial, the drug slowed disease progression 23% more than placebo but the ENGAGE trial failed to show similar results—only a 2% decline in disease progression and it wasn’t significant. In any regard, Aduhelm was approved by the FDA in June 2021 with as a first-of-its-kind treatment of Alzheimer’s. Aduhelm went through the accelerated approval process, which isn’t unusua for these experimental drugs, and Biogen is required to perform tight post-marketing information of side effects and benefits. 3 advisors to the FDA did resign due to the decision to approve Aduhelm despite a lack of clear efficacy.
• Now it’s important to keep things in context for Aduhelm and why drugs like it get approved. We covered a similar topic when we talked about the Orphan Drug Act but let me set the scene: you're 56 with two children graduated from college and another in their third year of college. Your wife and you both work 9-5 jobs to afford your middle class lifestyle. You spent the last 20 years working at the same company to get to the position you achieved and then your memory starts to fail. It’s a few words at first but then it’s longer moments of confusion. You’ve been diagnosed with early onset Alzheimer’s disease.
  • It may seem dramatic but the average lifespan after diagnosis is four to eight years. What would you and your family try to extend your life or possibly cure you? Would you try an experimental drug, one that could lead to brain swelling and microhemorrhage? What is the price for your memory? Luckily you and I can play devil advocate and debate the pros and cons, but for the person who’s memory is fading or their families, this is a real question.