MCED Discovery


In some ways, the discovery of MCED reads like a movie script.  A cancer physician, laboring alone in his laboratory, has been peering though a microscope for hours.  Bleary-eyed, he squints at biopsy specimens obtained from cancer patients.  Each glass microscope slide he reviews displays the same cellular features he has encountered hundreds of thousands of times over a 35 year career as a medical oncologist and cancer researcher.  Suddenly, he notices something - a shape on the microscope slide catches his eye.  Is it an artifact of slide staining?  Perhaps debris of some sort?  He readjusts the focus to bring the shape into sharper relief.   He twirls the adjustment knob that controls the stage - the platform on which the glass microscope slide rests - in order to view a different field, a different area of the slide.  Another strange object looms into view, similar to the first.  And then another.

The objects Dr. Larry Weisenthal encountered that evening in his laboratory were endothelial cells.  More precisely, they were patterns of calcium staining associated with endothelial cells that happened to die in a certain way - a way that was unknown to science before Dr. Weisenthal discovered it.

Endothelial cells make-up the linings of our blood vessels - our arteries, veins, capillaries, and heart valves.  Dr. Weisenthal’s purpose in viewing those particular slides was to observe the effect of certain chemotherapy drugs upon the malignant cells of a relapsed cancer patient - a person battling to stay alive.  Dr. Weisenthal’s laboratory is largely focused on finding the most promising, personalized chemotherapy for each cancer patient (if you would like to learn more about that aspect of Dr. Weisenthal’s work, here is a link to a website that discusses it:  www.weisenthalcancer.com).  In order to understand how this relates to a discovery involving heart disease it is necessary to understand the nature of the biopsy specimen Dr. Weisenthal was studying. 

Tissue Microaggregates and Personalized Medicine

In the case of what is called a “solid” tumor type, such as breast cancer, lung cancer, colon cancer, or ovarian cancer, an actual tumor mass, containing living cancer cells, is surgically removed from the cancer patient.  The biopsy specimen is placed immediately into sterile medium - a liquid mixture of life-sustaining ingredients.  The specimen is then rushed to Dr. Weisenthal’s laboratory by overnight courier.  Upon arrival at the lab, a variety of techniques are applied to reduce the specimen to an almost pure concentration of living cancer cells.  Almost pure - that part is very important. 

The cancer cells are not completely isolated.  Instead, they are maintained as tiny, three-dimensional clusters, called microaggregates, in order to preserve important cell to cell signaling.  The microaggregates containing the still-living cancer cells are then distributed among the wells - shallow indentations about half the diameter of a pencil eraser - in a rectangular, polypropylene microtiter plate.

There are 96 individual wells on each plate.  A different chemotherapy drug, or combination of drugs is added to the microaggregates in each well.  

The plates, containing cells and drugs, are then placed in a temperature and CO2-controlled incubator for 96 hours.  At the end of this drug exposure period, the cells are deposited on glass microscope slides through a process called cytocentrifugation.  Biological stains are then applied.  These are special chemical dyes that color-stain certain cellular features and leave other features unstained. 

At this point Dr. Weisenthal examines the stained cells under a microscope in order to assess the effect of each chemotherapy drug upon each different group of cells.  Dr. Weisenthal’s mission, as stated perviously, is to learn which, if any, anti-cancer drugs were effective in killing each patient’s cancer cells and which drugs were not effective.  This enables the cancer patient to receive a chemotherapy treatment, personalized for him or her, which offers the best chance for remission or cure based upon its observed anti-cancer activity in the lab.

Now let’s return to the discovery.  As it turns out, in addition to cancer cells, Dr. Weisenthal’s microaggregates also contain endothelial cells.  (Dr. Weisenthal was later able to confirm the identity of these cells using special biochemical stains that react preferentially to this type of cell.)  Why did he notice these cells now when he had not noticed them before?  It was because of the extraordinarily vivid staining displayed by the endothelial cells on that specific microscope slide.  And why had these endothelial cells stained so vividly on this particular slide - and why had other researchers not discovered this particular staining pattern previously?  There are three reasons, really, and here’s where serendipity comes into the picture.

Serendipity Knocks

First, let’s address the question of why other researchers had not made the discovery previously.

There is a term called “cell culturing” that refers to the process of either encouraging the growth of living cells (i.e. causing them to multiply) or else, as in Dr. Weisenthal’s case, not actually growing the cells but instead just keeping them alive in order to study their behavior in the laboratory.  Cell culturing always involves the use of a special medium - the liquid substance mentioned earlier containing ingredients that help to sustain living cells.   

It turns out that people who specifically study endothelial cells use a special type of culture medium that has been painstakingly designed, through much trial and error, to stop endothelial cells from dying while they are undergoing study in the laboratory.  As Dr. Weisenthal’s later experiments were to reveal, this endothelial cell culture medium is especially good at inhibiting MCED.  MCED is an abbreviation that stands for Massively Calcified Endosome Death.  It is the name Dr. Weisenthal assigned to the previously-unknown type of endothelial cell death that is the object of his discovery. 

Recall, however, that Dr. Weisenthal was not intending to culture endothelial cells.  As a matter of fact, he didn’t realize that endothelial cells were even present in his cancer cell microaggregates.  They just happened to be there - “hitch-hikers,” as he now calls them.  The culture medium used by Dr. Weisenthal was designed to sustain cancer cells and not endothelial cells.  Since there was nothing in Dr. Weisenthal’s culture medium to prevent the occurrence of MCED (which only affects endothelial cells and not cancer cells), many of the hitch-hiking endothelial cells on one particular slide - more about that in a moment - underwent MCED-mediated endothelial cell death. This, in turn, is what caused them to display the vivd staining pattern that caught Dr. Weisenthal’s eye. 

Two Types of Cell Death

We probably should say a word here about that vivid staining pattern and what caused it.  A defining feature of MCED, as opposed to to the previously known type of endothelial cell death, is the gradual accumulation of large amounts of calcium.  In non-MCED cell death, calcium levels spike and then dissipate rapidly in a matter of seconds or minutes.  In MCED mediated cell death, calcium levels rise steadily over several hours.  This meshes nicely with Dr. Weisenthal’s 96 hour drug-exposure method.  It is another serendipitous aspect of the MCED discovery.  It is precisely this large volume of accumulated calcium that stained so vividly on Dr. Weisenthal’s cancer slide. 

Why Didn’t You Think of This Sooner?

Now for the other question:  why was it that, in 35 years of studying cancer cell microaggregates, Dr. Weisenthal had not noticed the MCED staining pattern sooner?  The answer to that must be told in two parts - and here is where the serendipitous miracle truly occurs.

The first part of the answer may seem a bit dry.  It will seem less so, however, when combined with the second part.  It also figures largely in a cataclysm of events that occurred later which nearly sank the entire discovery.  But first things first. 

We mentioned earlier that endothelial cell culture medium contains, almost by accident, MCED inhibitors.  We say “by accident” because the developers of the medium did not know about the existence of MCED - all they knew is that their medium seemed to keep endothelial cells alive.  Also by accident, although Dr. Weisenthal did not know it at the time, some - but not all - batches of a specific ingredient Dr. Weisenthal had been using for many years in his lab’s home-brewed cell culture medium also contained these same anti-MCED factors.  Clearly, the manufacturer of the ingredient knew nothing about them either.  It was entirely luck of the draw, as later experiments were to confirm, that those particular cells upon which Dr. Weisenthal made his discovery had not been cultured in a medium that contained occult anti-MCED factors.

That was the technical part.  Now, here is where things become a little more dramatic.  In retrospect, Dr. Weisenthal now refers to the cancer specimen upon which the MCED discovery was made as “MCED Specimen Zero.”  It is the starting point for a series of follow-on discoveries involving other human tissue specimens. 

Pancakes?

MCED Specimen Zero happened to come from a neuroendocrine tumor.  The specific tumor type isn’t as important as the fact that these cancer cells, when tested in the laboratory, were tenaciously resistant to injury by any of the chemotherapy drugs tested against them.  And so, as Dr. Weisenthal labored his way through the glass microscope slides - more than 50 of them for this one cancer patient - he found that slide after slide was crowded with big, bright, fully-intact cancer cells.  It was like performing a close-up inspection of a Monet painting in which all of the lily pads were painted bright pink.  Actually, Dr. Weisenthal uses a different metaphor.  He describes the unaffected cancer cells as big pink pancakes.  And, because these cells had all been exposed to standard chemotherapy drugs - designed to kill cancer cells and not endothelial cells - even if MCED had occurred on any of the slides, it was rare, largely obscured by the abundance of intact cancer cells, and easily mistaken for debris. 

One Shot in a Million

But the last slide was different.  The cells on that slide had been exposed to a drug called bevacizumab.  Bevacizumab’s main purpose is not to kill cancer cells directly.  Bevacizumab works by depleting a substance called vascular endothelial growth factor, commonly abbreviated as VEGF.  Without VEGF, endothelial cells die.  Why should an anti-cancer drug target endothelial cells?  We will explain that just a little farther along.  Meanwhile, our story has finally reached its climax - the precise moment of discovery.     

When Dr. Weisenthal viewed this slide he found it crowded with big, pink tumor cells, as before - but he also found, scattered among the tumor cells, conspicuous purple splotches.  Here is how Dr. Weisenthal describes it, using his “pancake” metaphor:

“So, basically, I looked at close to 50 cytospins in a row and they were all monotonously the same - filled with nice clean, pink pancakes. The very last slides were the bevacizumab slides. They looked different.  Instead of being filled with plain pink pancakes, they were filled with pink blueberry pancakes.”

Opportunity Meets the Prepared Mind

At first, Dr. Weisenthal was puzzled as to the meaning of this unusual staining pattern.  He circled back and reviewed hundreds of archival slides among the thousands in his extensive library.  These had been created by Dr. Weisenthal in the past while performing chemotherapy drug testing for hundreds of cancer patients.  Many of these patients had died long before.  And yet their preserved cells remain as a resource to guide progress on behalf of persons still living or not yet born. 

Dr. Weisenthal then set about conducting hundreds of individual experiments, very often using his own blood - quarts of it, it seemed.  The experiments involved different staining methods, different cell culture techniques, different types of chemotherapy drugs, sera from different human donors, and exposure of the cells to other substances which he felt might have some effect upon them.  He scoured the medical literature, looking for clues that might help to corroborate or refute theories that were beginning to crystalize in his mind. 

Finally, as the picture became increasingly clear, Dr. Weisenthal recognized that he had discovered a brand new mechanism of endothelial cell death - one that had never before been described in the medical literature.  He named the new mechanism MCED - Massively Calcified Endosomal Death.  And so now he had a new mechanism and also a new name for it.  But what did it all mean?

MCED in Cancer Treatment

As a cancer physician and researcher, it first occurred to Dr. Weisenthal that he had just invented an exquisite test to guide the use of a new class of anti-cancer agents called angiogenesis inhibitors.  Bevacizumab, discussed above as the drug that produced the “blueberry pancake” slide, is one of these drugs.  Anti-angiogenic drugs work by interfering with the growth of capillaries which deliver oxygen and nutrients to a tumor mass.  They also serve to remove waste products.  By destroying the capillaries, tumor cells are starved of life-giving blood and they die. 

The problem with these promising new drugs is that they do not work for a large percentage of patients.  And when they do work, they may work well for a while and then stop working altogether.  Also, anti-angiogenesis drugs are enormously expensive and they often produce dangerous side-effects.  And so a way was needed to determine exactly which patients would benefit from anti-angiogenesis drugs and which patients would not.  Dr. Weisenthal’s new test afforded an ideal solution.  At least, so it seemed.

Prepared Mind Meets the Real World

No matter how good a new medical test appears - any new test - it must be validated. This means correlating what is observed in the test with what actually occurs in the patient. For example, Dr. Weisenthal’s test for other standard and targeted anti-cancer drugs was proven highly-accurate in more than 100 clinical studies, including a large, prospective randomized trial.  In every study, results yielded by Dr. Weisenthal’s test correlated closely with patient response and/or with patient survival.   

Dr. Weisenthal approached the company that produces the most widely-used of the new anti-angiogenesis drugs to see if it would be interested in participating in a correlation study of his new anti-angiogenesis test.  Worldwide sales of their drug are estimated at over $6 billion.  However, problems were arising for the company.  Insurance companies were refusing to pay for use of the drug in many cases and the FDA had withdrawn its approval in breast cancer because not enough breast cancer patients were being helped by it. 

Dr. Weisenthal assumed the company would be thrilled to learn that a test existed that potentially could identify patients most likely to benefit from its blockbuster drug.  He was wrong.  The company was not interested in participating in even a small pilot study.  Dr. Weisenthal offered to donate his time, his labor, and his expertise to the project entirely for free and to pay for all costs of testing out of his own pocket.  The company stated, in effect, that it had no interest.  No explanation was provided and so it is hard to guess what the company’s thinking might have been in the matter.  A cynic might assume that the company would prefer that all cancer patients receive their expensive drug in order to identify the few who will benefit from it.

Another Critical Discovery is Made

Throughout all of this, Dr. Weisenthal’s research moved forward.  Further exhaustive experimentation and much innovative deduction led Dr. Weisenthal to the conclusion that specific factors - substances that could be detected in the blood - are associated with the occurrence of MCED.  Importantly, he also discovered that different human sera - essentially blood from different individuals - have differing abilities either to promote or inhibit MCED.  But still, what does this have to do with heart disease?

MCED in Heart Disease

Despite much progress made in the prevention and treatment of heart disease, it remains the leading cause of death worldwide.  Moreover, many mysteries remain unsolved.  Why, for example, do some persons with high serum lipid profiles - high amounts of cholesterol and other fatty acids in their blood - have lovely, unobstructed arteries while other persons, whose serum profiles are deemed favorable, suffer arterial blockage, heart attack, and stroke?  High cholesterol is often associated with heart disease but, inconveniently, not always.  Why is that?  Clearly, cholesterol seems to be a factor but, just as clearly, it is not the factor.

It is also not clearly understood by scientists exactly what causes vessels to become inflamed.  Inflammation, by the way, is the new way of referring to the narrowing of an artery.  It is now fairly universally understood that fat does not literally become deposited on the inner walls of blood vessels like sludge in an old sewer pipe.  Instead, the wall of the vessel itself undergoes an apparent thickening as part of an inflammatory process.  This causes the lumen, the channel in the middle of the vessel through which blood flows, to become progressively narrower until, one day, a turbulent fluttering of blood against the narrowed, uneven surface of the vessel wall serves as a signal to summon platelets.  This results in formation of a blood clot.  It is the same highly-useful mechanism that stops a minor cut from bleeding endlessly.  However, in this case, it delivers a death-blow by creating the final blockage that triggers a heart attack or stroke, depending upon where the clot ultimately lodges. 

But what causes this inflammation to occur?  A few theories have been proposed and these have achieved varying degrees of acceptance within the scientific community.  But all of the theories fall short in accounting for apparent contradictions which are known to exist in the genesis and behavior of atherosclerosis.  All of the theories fall short, that is, except one.

MCED appears to explain everything.  In fact, having formulated his MCED hypothesis, Dr. Weisenthal then undertook a series of experiments ruthlessly designed to disprove it.  Far from refuting the hypothesis, each experiment revealed yet another insight that seemed to support it.  The most recent of these insights - enhancements to the original hypothesis which more fully elucidate the MCED mechanism -  only recently emerged. 

Before moving on, it might be useful to recap where things stood just before the new insights came to light.    

MCED Discoveries, 2006 - 2013

Without becoming overly-technical, here is a summary of Dr. Weisenthal’s conclusions as of late 2013.  If you wish to read more detail, kindly consult the “Publications” page of this website.  Links to Dr. Weisenthal’s MCED journal articles, abstracts, and poster presentations are provided.

  1. MCED occurs only in endothelial cells and not in normal or cancer cells.

  2. MCED can be induced by certain substances which are known to be lethal to endothelial cells.

  3. Among the MCED-inducing substances are anti-angiogenic drugs and oxidized lipids.

  4. Other chemicals, including standard chemotherapy drugs, induce only non-MCED-mediated cell death.

  5. MCED can be distinguished from the non-specific cell death.

  6. MCED is an energy-dependent process that can be inhibited by non-specific cytotoxic agents and by sub-optimal culture media.

  7. MCED involves a specific, multi-step pathway.

  8. Immune cells appear to interact with MCED cells, suggesting that the latter present an antigenic focus.

  9. A ligand, such as an oxidized lipid, probably interacts with a receptor and initiates an MCED cascade. This would provide a new and highly specific target for prevention and/or treatment of cardiovascular disease.

  10. The most important cause of susceptibility to atherosclerotic coronary vascular disease probably is the presence or absence of circulating MCED factors and not cholesterol or other serum lipid levels.

  11. It is probable that some circulating serum factors inhibit MCED while other factors promote it.

  12. Most likely, MCED factors will be detectable and measurable in blood and other sera.

  13. The prevalence of MCED promoting and inhibiting factors varies among different individuals.

New Insights - from MCAD to MCED

The insight that endothelial cells die through the mechanism of MCED did not spring fully-formed from Dr. Weisenthal’s intellect and imagination.  For several years, Dr. Weisenthal discussed, published, and otherwise referred to his findings using the acronym MCAD, which stands for Massive Calcium Accumulation Endothelial Cell Death.  The name derived from the calcium “lakes,” that form around endothelial cells undergoing this newly-discovered form of cell death.  However, while MCAD was and is a defining feature of the inflammatory mechanism there is more to the story.   

Following nearly eight years of painstaking research, marked both by encouraging successes and disappointing set-backs, Dr. Weisenthal was able to drill down further into the step-by-step process by which an endothelial cell actually dies.  In doing so, he answered additional questions about what specifically causes vessels to become inflamed and obstructed.

MCED Explained

Based on Dr. Weisenthal’s most recent research, here is his assessment of what probably happens: 

Endothelial cells need substances called pro-angiogenic factors, in order to stay alive.  Pro-angiogenic factors work by contributing something of value to a broader pro-angiogenic pathway.  In the world of cell biology, the word “pathway” is used to indicate a multi-step biochemical process.  If something happens to interfere with one or more pro-angiogenic factors, the entire pro-angiogenic pathway can be disrupted.  When this occurs, a different pathway - one that leads to endothelial cell death by the mechanism of MCED - can be set into motion.

No matter how MCED begins, the pathway always leads to the same destination - the formation of endosomes

An endosome is a tiny transport vesicle - think of it as a self-contained shipping department that springs spontaneously into existence, sorts-out various substances, and then either transports them into the interior of the cell or else ushers them unceremoniously back out onto the street, depending upon whether or not a specific substance is deemed useful to the cell. 

When an endothelial cell is dying by the mechanism of MCED, the endosomes become highly calcified and are expelled from the cell.  An endosome that is expelled from a cell changes its name and is thereafter called an exosome.  As seen through a microscope, calcified exosomes cluster around dying endothelial cells like bunches of grapes.         

The presence of highly-calcified exosomes and endosomes provokes a response in which the usual cast of inflammatory immune cells - monocytes, lymphocytes, neutrophils, etc. -  attack what they perceive to be unwanted debris.  In the course of the clean-up effort a zone of chronic inflammation arises throughout a field of dead and dying endothelial cells.

A Deadly Cycle

Despite the body’s best efforts not all of the calcified endosome and exosome debris can be cleared away.  And so the body does what it always does with an irritation it can’t get rid of - it tries to wall it off.  It is roughly the same process that occurs when a splinter stays in the body too long or when a cyst is formed around an infection.

In the case of vascular MCED, the body walls-off the calcified debris and chronic inflammation by laying-down a covering of smooth muscle along the interior of the blood vessel.  New endothelial cells are then recruited.  These arrange themselves over the smooth muscle layer.  (You may recall when we stated earlier that blood vessels are always lined with endothelial cells.) 

Unfortunately, the new endothelial cells are now vulnerable to injury by exactly the same factors that killed the previous layer of endothelial cells.  And so the whole process repeats itself.  As each endothelial layer dies, becomes inflamed, and is covered-over by a new layer, the the interior of the vessel becomes narrower and narrower.  This progressively restricts the flow of blood through the vessel until a blood clot is formed and stroke or heart attack results.

Dr. Weisenthal’s newest findings are important because, as more of the MCED cell-death pathway is mapped, more opportunities arise along its entire length to intervene with drugs that disrupt the pathway, block MCED, and halt or prevent vascular inflammation.  

Bumps in the Road

The MCED discovery process hasn’t always gone smoothly.  There have been set-backs along the way, some of them gut-wrenching.  At one point, Dr. Weisenthal lost nearly a year when MCED simply stopped working in his experiments for no apparent reason.  Dr. Weisenthal finally discovered that the problem was caused by variable batches of a certain cell culture medium ingredient - one of many ingredients he uses in his medium.  Diabolically, the ingredient was supposed to be identically-formulated from batch to batch and it was all purchased from the same supplier.  With additional staff to conduct experiments and with money to test multiple batches of medium simultaneously he could have resolved the mystery in a week. 

The Road Ahead - From Discovery to Cure

The work on MCED continues.  Much has been accomplished and much remains to be accomplished.  But the paradigm-changing aspect - the discovery of MCED - has now been made.  Identifying and isolating one or more MCED factors should be reasonably straightforward.  But it requires funds and it requires bio-pharmaceutical collaborators.  The purpose of this site, therefore, is to raise awareness - to let people know that this crucial work is being carried-out by a lone scientist who is both under-funded and under-staffed. 

The history of medicine is conspicuously littered with scores of instances in which critically-important discoveries were ignored by a medical establishment, apparently caught-up in the inertia of existing dogma, that sometimes seems more comfortable with “process” than with true discovery.  And in some cases discovery can be downright inconvenient, especially when entire academic careers and billions of industry dollars are fully-invested in older schools of thought.  Fortunately, truth, like cream, always rises to the top.  Invariably, after years or decades of benign neglect, the establishment catches-up and “rediscovers” what was discovered previously.  Typically, when this happens the original discoverer is acknowledged.  The discovery may even be named in his or her honor.  All too often this honor is accorded posthumously.

One way or the other, MCED will be recognized, along with its originator.  How many years or decades will pass until that happens?  And in that time, how many lives will be diminished by heart disease and stroke?  How many lives will be lost to it?


       

 

New Discoveries in Cancer and Heart Disease

“Our own discovery of MCED was entirely serendipitous.”






















































“As the picture became increasingly clear, Dr. Weisenthal recognized that he had discovered a brand new mechanism of endothelial cell death.”
















































“Having formulated his MCED hypothesis, Dr. Weisenthal then undertook a series of experiments ruthlessly designed to disprove it.”





















































“MCED appears to explain everything.”












































“The purpose of this site is to let people know that this important work is being carried-out by a lone scientist who is under-funded and under-resourced.” 





















“Truth, like cream, rises to the top.”