Studies of Caloric Restriction, Resveratrol and SIRT1 Gene

Regulation of Intermediary Metabolism and Mitochondrial

Biogenesis Demonstrate a Requisite ‘Metabotype’ Continuum
from Cellular Rejuvenation to Aging to Cancer, Independent
of Cancer Cell Growth Factors, Suppressors or
Immortalizers.
Gregory S. Bambeck, Ph.D.

Abstract
In the early to mid twentieth century Otto Warburg
hypothesized that cancer cells could be characterized by dramatically elevated
glycolysis and mitochondrial respiratory deficiency locked in a relationship he
called ‘aerobic glycolysis’. The respiratory deficiency claim was proven false
and cancer cell biochemistry shifted its studies toward oncogenes, cell growth
factors and their cascades, cancer cell growth suppressor systems, apoptosis,
telomeric immortalization, cell recognition and adhesion mechanisms etc. Any
global hypothesis devoted to mitochondrial inefficiency or dysfunction related
to the anabolic and catabolic control requirements to cancer cell function, such
as those proposed by G. Bambeck were refused funding or publication, and in
fact, were quietly chaperoned out of the halls of science. Now, some 30 and
some 50 years later, the pariahs may have been proven to be visionaries.
Glycolytic blocking and retrafficking agents, such as dichloroacetate etc. can
either kill or ‘renormalize cancer cells. Caloric restriction, the only known
mechanism for extending life well beyond its normal span, in everything from
roundworms to primates, is now known to rejuvenate aging cells and down
regulate cancer cell initiation and growth by turning on a suite of 745, or
more, genes that renormalize glycolysis and initiate phagocytosis of
inefficient mitochondria concomitant with biogenesis of efficient, new
mitochondria. Resveratrol turns on the exact same suite(s) of genes, and by
every measure, has the same cytological impact. The ‘metabotype’ continuum
from juvenile cell, to aging cell, to cancer cell, make even more sense in an
evolutionary context.

FORWARD

This is a general review article which is presented as a narrative. It is not

intended for peer review, in part, for reasons contained in the abstract and the
body of the text. That does not mean that this work is unimportant. In fact, it
might be very important, because it connects together lines of research that
converge on medical implications of great magnitude. It is written by a PhD
scientist with over thirty fruitful years of scientific research under his belt,
who hopes that he may have the time, later, to write a more formal article.
This information should be gotten out there informally and quickly, rather
than not to be gotten out at all.

There is no bibliography or references in this work, but at the end of this text,
a short list of search engine words and suggested readings are provided, so
that more than enough bibliography is provided to see ‘the big picture’
presented by the hypothesis contained, herein. This article is restricted to the
basic catabolic and anabolic changes in intermediary metabolism and how
they relate to juvenile, adult and cancer cells, because this area of cancer
research has been left fallow. Other very important areas of research, such as
the aforementioned cell growth factors etc., have tens of thousands of articles
that may be referred to.

This article is devoted to an area of research that has been relegated to a

backwater, in particular, in regards to cancer cell intermediary metabolics. If it
had not been for anti-aging and life extension research results, stumbling
through the metabolic ‘back door’, so to speak, some thirty years after a
modified Warburg hypothesis, coupled with some new cancer metabolism
blockers, the important connections could not have been made. Throughout
history, serendipities and/or convergences have come together to form holistic
emergent systems with more than just notable impact. Hopefully, this one of
those times.

Lastly, this article is written a la Scientific American in that it is, hopefully

designed to be understood by the educated lay person, while not short
changing the serious scientist, too much. Professional jargon will be held to a
minimum. It is my hope that a more in depth review won’t be necessary, if
enough interest ensues. It would certainly be far more gratifying if interested
parties would take up the discussion, or even initiate lines of investigation that
might knit the outlines of these semi-integrated patches into a more complete
fabric of either enhanced, or wholly new understanding. So, here we go, more
or less ‘off the cuff’.

Earlier Times

Approximately 96% of a cell’s organic biomass consists of carbohydrates,

fats, nucleotides and proteins. All four of these components can be either
burned as fuel to power the cell, or as construction materials to replace
defective parts in a quiescent cell or to build a new cell, as in the case of a
dividing cell. In a normal, healthy quiescent cell, there is a balance between
energy production and repair rates, so that the cell is said to be in homeostasis,
or balance. In a normal, healthy dividing cell, growth factors set up a cascade
of directives that order the cell to both increase its energy output and to import
raw materials and build those raw materials into the components of a new cell.
In short, nucleotides become RNA and DNA, amino acids become proteins,
sugars become complex carbohydrates and fatty acids become lipids in a
process called anabolism, while these same four starting materials can,
alternatively, be burned as fuel to create energy, in a process called
catabolism. The anabolic intermediate products can further assemble, by
anabolism, into molecular assemblies, such as ribosomes, enzyme complexes
etc., and, even further, into organelles such as mitochondria, lysosomes, a cell
nucleus etc. Eventually, a whole new cell is formed. Cancer cells do this to,
but as we shall see later, they don’t do it the same metabolic way as normally
dividing cells. Instead, they do it more like in an extended metabolic version
of aged cells.

Although, what has been presented so far, is a gross oversimplification,

suffice it to say, that by the middle of the twentieth century, a pretty
sophisticated outline of many hundreds of the molecular trafficking pathways
of the catabolic and anabolic interplay of organic molecular transitions, were
mapped out. Wall posters in classrooms demonstrated these processes in a
fashion analogous to watching automotive freeway traffic flow from an aerial
view over a large city. Many discoveries have been made by observing
disruptions in the flow, just as auto accidents cause clogs on off-ramps, on-
ramps and thru-ways on the freeway system.

Otto Warburg, in charge of a large research organization, and a highly

respected molecular researcher of his day, being the early to mid twentieth
century, thought he saw an intermediary metabolic perturbation both unique
and specific to all cancer cells. More precisely, he hypothesized that this
perturbation was critically confined to the catabolic, or fuel burning side of
metabolism, but also set in motion an anabolic shift as a consequence. Even
more specifically, he targeted the anomaly to the burning of a single fuel,
glucose, to the relative exclusion of other fuels.

Glucose is the primary, but, by no means, only fuel that is burned by cells of
the body, and it is burned in two complex systems, called glycolysis and the
Krebs cycle. Glycolysis obtains energy from glucose and other sugars by an
anaerobic (non-oxygen utilizing) mechanism to produce ATP, an energy
carrying molecule. The Krebs cycle, contained in an organelle called the
mitochondrion, burns the glycolytic end product, pyruvate, utilizing an
aerobic (oxygen consuming) mechanism to produce ATP. The burning with
oxygen process sequentially strips hydrogens from the glycolytic end product,
converting NAD to NADH2 and then couples the NAD hydrogens to oxygen
to form water, while releasing the sugar carbons to form carbon dioxide. The
formation of water is a stepwise process in which the energy of the hydrogens
and their electrons are rejoined in a stepwise process called electron transport,
while the energy of the process is captured by converting ADP to ATP, while
H and O form water, in a process called oxidative phosphorylation.
Anabolism is the opposite process in that NAD hydrogens and ATP energy
are utilized to build cell components. Thus the two swing molecules in the
process are ATP and NAD as they switch back and forth between their low
and high energy forms and their oxidation and reduction forms, respectively.

In a healthy homeostatic cell, many fuels are used, and about 5% of the ATP
is produced by glycolysis, while about 95% of the ATP is produced by the
mitochondria. Anabolism and catabolism are in a steady state balance with
mitochondria producing this ATP energy at about 99% efficiency. In a healthy
dividing cell, the entire energy production system is upregulated to make ATP
energy and NADH reducing power for the anabolic requirements to make a
new cell. Otto Warburg notice an uniqueness in the catabolism of cancer cells,
in that glycolysis was considerably elevated, that respiration was depressed
and that mitochondria in cancer cells appeared small, malformed or
disorganized. Furthermore, he proposed that, unlike fetal or other normally
dividing cells, the cancer cell was irreversibly ‘stuck’ in this metabolic
phenotype. Although the glycolytic part of his hypothesis was never refuted,
the respiratory defect notion was struck down in a furious debate in 1955-
1956. Mitochondrial respiratory deficiency, although found in many tumor
types, was not found in all tumor types, and was not considered required as a
fundamental requisite of the cancer cell condition.

As mentioned before, Warburg was a big name during his time. He made and
broke many scientific careers, and was known for having a bit of an irascible
nature. Well, the bigger they are, the harder they fall. Many of Warburg’s
detractors became journal editors, reviewers and laboratory directors. Woe be
it to anyone positing any form of mitochondrial defect/cancer hypothesis,
even twenty five years later.

After the Warburg hypothesis rejection, cancer research shifted away from
metabolic studies toward oncoviruses, oncogenes, cell growth factors and
their cascades, cell growth suppressor systems, apoptosis mechanisms,
telomeric immortalization, cell recognition and adhesion systems etc. These
studies have had a huge impact upon our understanding of the normal, to
cancer cell, transformation process. It has become obvious that evolution has
provided numerous impediments to lethal carcinogenesis in its attempts to
keep cell division under control. There is also no doubt that these new areas of
research would have opened up regardless of the outcome of the Warburg
hypothesis. However, it is also true that investigations of mitochondrial
interactions in the intermediary metabolic interplay in the cancer cell, would
not have all but dried up, as it most certainly did.

In 1975, some twenty years after the Warburg hoopla, a very politically naïve
graduate student, named G. Bambeck became fascinated with mitochondria, in
a Kent State University laboratory, that happened to have a mouse
lymphoblastic lymphoma model. He isolated mitochondria from many mouse
tissues and noted that the lymphoma mitochondria had uniquely low ADP:O
ratios. This means that these mitochondria were producing abnormally low
amounts of ATP energy per oxygen consumed. This could mean that either
ATP was being uncoupled from oxygen consumption via the respiratory
chain, that reduced NAD was either being decoupled from oxygen or being
exported from mitochondria in abnormally high rates, via some kind of
chemiosmotic shuttle, for anabolic purposes, or in some combination, thereof,
by some unknown mechanism(s).

Thus, he began a literature search and, among other things, he ran into the
Warburg debacle. But, being socially unsophisticated, he pressed on, and he
pressed on because he found something very interesting, so interesting, in fact,
that he virtually abandoned his wet lab (hands on) research for a mental form
of research. In mental research, one reviews the work of other researchers in
the hopes of ‘an unique synthesis of thought’, or, in other words an
overlooked ‘big picture’, that, in this case, might also be suitable for a PhD
dissertation.
First, G. Bambeck found that there were a lot of cancer cell mitochondria vs.
normal cell mitochondria papers out there, a number of them showing that
Warburg’s respiratory defect was not there, in more than just a few cases.
What he did find, in every cases where the data was taken, that net
mitochondrial ATP production on a per mitochondrion and/or, more
importantly, per cell basis, was significantly lowered compared to normal
dividing cells. He further noted that mitochondrial net ATP synthesis shortfall
could be due to low mitochondrial numbers per cell, inefficient electron
transport, increased NAD/NADH shuttle export of reducing power,
inefficient coupling of ATP formation to hydrogen chemiosmotic potential, a
shift in the ATP synthetase to ATPase equilibrium dynamics or some
fundamental metabolic Km shift in the chemical pathway linking glycolytic
end products to the Krebs cycle. Anything that would block the connection
between the NAD/NADH redox coupling to ATP formation, could tip the
balance of carbon flow to anabolism. Most importantly, these mitochondrial
shortfalls of ATP production could change the glycolytically produced to
mitochondrially produced ATP production ratio by over 1000%. He proposed
that this ratio differential forced fuel dependency upon glucose while
simultaneously forcing other metabolites, reducing power and ATP energy
flux toward anabolism. In the light of modern discoveries, to be discussed
later, one could say that such a system pre-adapts the cell, metabolically, for a
more uncontrolled growth format when not necessarily signaled by a growth
factor: a precancerous state or hyperplasic non-dividing growth condition, as
one might have it. If carcinogenesis is a multi-step process, why not have a
metabolic process, or a progression toward a metabolic state that predicates
the terminal process when suitable mutations and their associated stimuli
arrive?

G. Bambeck optimistically presented his conclusions to the scientific

community with the expected dismissive results. Even though his dissertation
was of a somewhat heretical nature, he received his PhD because the logic
was essentially sound. But with nowhere to publish and nowhere to work on
his findings, he evaporated from obscurity to nothingness in the annals of
cancer research. Instead, he plied his trade in medical diagnostics and research
tool technologies. The tools of 1975-1980 were not available to address a
rationale for the collapse of mitochondrial efficiency in cells. For one, there
seemed to be too many ways for it to happen. Gene switches and gene
switching systems were just in their early days of initial elucidation. There
were simply too many unknowns and alternatives. G. Bambeck took a stab at
the problem, and notioned that reactive oxygen species (ROS), called free
radicals at the time, might be peppering the mitochondrial and nuclear
genome, causing a sequential randomizing of the mitochondrion and
manifesting itself as a progressive decline in fuel burning efficiency.
Paraphrasing, he said that, ‘by whatever means, the per cell mitochondrial
ATP production deficiency is there’. We now know this to be true, and not
just in cancer cells, but in aging cells as well. Its most severe manifestation
appears to occur in the cancer cell. The anaerobic to aerobic ratio of ATP
production appears to increasingly exacerbate as cells age and become
transformed. Cell heat production and adaptation to hypoxic conditions are
hallmarks of glycolytically adapted cells with poorly coupled ATP formation,
as is witnessed by the limited success with hyperthermic, hypobaric and lactic
acid export blocking cancer therapies. G. Bambeck hypothesized that more
specific blocking agents to glycolytic and mitochondrial systems, might have
greater efficacy, either for instituting differential kill or cancer cell
renormalization. The now known facts that glycolytic blocking agents can kill
or that fetal pyruvate kinase blocking agents can renormalize cancer cells to a
non-anabolic and non growing state, and that conditions and agents that create
efficient aerobic catabolism via mitochondrial biogenesis, reduce cancer
incidence and increase cell rejuvenation, show a more mechanistic support for
the concept. However, much of these data come from research areas originally
perceived as only tangentially related, or not at all related, to cancer.

Toward Present Times

From the 1980’s to the present, there has been a lot of work with ROS and
their mutagenic and aging effects on cells. Basically, ROS are highly reactive
oxygen species containing an unshared electron which allows the ROS to
react with just about any available organic molecule, including DNA, RNA,
proteins etc. Naturally, ROS are mutagenic and, therefore, carcinogenic, akin
to ionizing radiation, causing the highly specific and organized genome and its
protein products to become more randomized, or nonsensical. Because they
are immersed in an oxygen atmosphere, either directly or by blood delivery,
body cells produce ROS and utilize free radical scavenger molecules to mop
up these nemeses. Anti-oxidants, such as vitamin C, vitamin E, bioflavinoids
etc. are such free radical scavengers, and experiments with elevated doses of
anti-oxidants and their effects upon cancer induction and aging rates are rife.
In general, the results are positive, helping organisms to more closely achieve
their well nourished natural life expectancy potential, but not beyond that
potential.

The mitochondrion produces more ROS than any other part of the cell because
it is the seat of oxidative fuel burning, where the electron cascade of the
respiratory chain is used couple the energy of the Krebs cycle intermediate
hydrogen electrons to ATP formation, then, ultimately to oxygen, to yield
water. To protect itself from its own internally produced ROS, the
mitochondrion utilizes endemic anti-oxidants and a special enzyme called
SOD to mop up ROS.

Each mitochondrion has several copies of its own DNA, and when ordered to
do so, mitochondria can multiply, similar to cells. But unlike cells,
mitochondria can do something amazing. Via a mostly unresolved process,
mitochondrial biogenesis (to be distinguished from simple division), results in
new efficient mitochondria arising from old inefficient mitochondria. Cell
nuclear genomes pass on the mistakes that their DNA repair systems fail to
detect or faultily repair, as do mitochondria, when simply dividing, but not
when undergoing biogenesis. Perhaps it is because a single cell has only one
nuclear genome (or at most, two half genomes), but as many as over a
thousand mitochondrial genomes that might be coupled to some selection
process, maybe based upon some kind of selection for fidelity of consensus
sequence. It seems obvious that there must be some kind selective renewal
system, because mitochondria pass from generation to generation via
oogenesis, on average, without a single point mutation. Without such a
process, it would not take many generations for eggs to contain a gibberish of
mitochondrial sequences, rendering the following of the matrilineal line to a
virtual impossibility. The fact that we can follow the matrilineal line through
thousands of generations, supports this notion. I envision something like a
polytene chromosomal sequence comparator mechanism of some kind, or
perhaps, a highly protected and sequestered mitochondrial ‘mother genome’
somewhere in the cell.

Evidence for real biogenesis mounts. In adult body cells, mitochondria

progressively degenerate into inefficient couplers of hydrogen, electrons and
oxygen to ATP and water production. And not just one thing, but a host of
things go wrong with mitochondria as adult cells age, with one of the hottest
causal suspects being ROS mediated mutagenesis. Concomitant with reduced
mitochondrial efficiency over time, is an increase in glycolytic ATP
production. Thus, as cells age, the anaerobic to aerobic ATP production ratio
rises, and begins to appear more like the pattern seen in cancer cells. Since
most adult cells are terminally differentiated, and can no longer divide, such a
ratio shift does not pose a cancer risk, as would adult stem cells, which still
have functional telomeres, and could possibly become telomerase
immortalized. Telomere shortening to terminal differentiation is a well
established mechanism, putatively, for avoiding cancerous progression in
adult cells.

However, non-terminally differentiated adult cells and adult stem cells can
divide, and they generally divide to replace dead or missing adult tissue, under
the directive of growth factors. Under such conditions, tissues are replaced but
they are not rejuvenated. Instead of young dividing new cells replacing old
dead cells, old dividing cells are replacing them, in part because mitochondrial
biogenesis is not occurring. Cells are being replaced, but the tissue is not
being rejuvenated. It appears that as we age, the metabolic phenotype
(metabotype) of the aging cell operates more and more like a cancer cell
metabotype. Somehow, it seems that that there is no great evolutionary
pressure to put additional roadblocks to such a metabotype progression,
especially with the, aforementioned, long list of non-metabolic quality
assurance mechanisms already in the toolkit. In fact, such a progressive
metabolic transition probably assists the evolutionary process. From an
evolutionary perspective, it is preferred at a certain point to dispose of the old
bodies, which represent yesterday’s genetic experiment, with the next
generational gene mix experiment. After all, it is the genome that has a shot at
perpetuity, and not the vehicles it employs to get there. In summary, it appears
that adult tissues are aged metabotypes of juvenile tissues and adult stem cells
are aged metabotypes of fetal stem cells, and that their metabotypes progress
toward the cancer cell metabotype, irrespective of other components of the
cancer cell transformational process. There appears to be a blended metabolic
continuity between these basic temporal cell types, and recent science on
cancer and aging are yielding insights that show metabolic crossover
applications between these once disparate fields of research. It appears that we
are beginning to achieve a degree of control over both aging and cancer, at
least from a metabotype perspective.

To a rather remarkable degree, the aging cell and cancer cell metabotype have
recently been reverted to normal with dramatic increases in life extension and
incidence reduction in cancer, from groups of apparently unrelated
experiments that only share their fortunate results by viewing them from a
shared metabolic perspective. By life extension, it is meant to mean ‘age
beyond its normal well nourished maximum’ This not to be confused with
achieving the natural maximum by delaying premature death, but instead, by
going beyond its natural healthy maximum by a considerable extent. Also, it is
far too early to talk of cancer ‘cure’, but the early data point to a significant
cancer renormalization or kill. But first, there is a need for a bit of a preamble.

Until recently, the only technique known to cause genuine life extension is to
initiate a condition known as caloric restriction. Caloric restriction and its life
extension affects hails back seven decades, but its many manifold mechanisms
of action are just now coming to light, due to new technologies grown out of
the genomics revolution. Studies had shown that caloric restriction
rejuvenated cells, by virtually every measure, and in virtually every tissue and
organ of the body, from neuroregeneration, to delayed and reversed muscular
wasting, visual impairment, skin wrinkling etc. These rejuvenation-like
phenomena occurred in organisms, as lowly as yeast, upward through the
multicellular organism evolutionary chain ranging from roundworms to mice.
Just this past year, a 25 year caloric restriction study on rhesus monkeys
extended these findings to the primate order, of which humans are a member.
Preliminary results on humans are yielding parallel results. By hindsight, such
findings make evolutionary sense, because feast and famine cycles go back to
the dawn of time. There is a distinct survival advantage in being able to sweat
out the lean times until nutrient availability of fat times allows energy
availability to support cell division in single cell organisms or procreation in
multicellular organisms. Hibernation or estivation may work for periodic
circumstances such as winter snow or summer drought, as it does for northern
bears or desert toads, but variable and unforeseen conditions have led to a
much more ancient and flexible metabolic solution, as observed by caloric
restriction. It seems apparent that untold millions of generations have been
honed by caloric restriction to utilize it as a generic life extender as a means of
avoiding extinction.

In just the last couple years, the genetic mechanisms behind the caloric
restriction phenomenon have become much more elucidated. First, caloric
restriction turns on a gene called SIRT1 that activates a suite of, at least, 745
genes that is normally turned on in juveniles, but is turned off in adults. At the
very least, SIRT1 is turning on a hugely complex gene system in adult cells
that is normally only operational in juveniles, tempting the notion of
‘rejuvenation’, especially since life extension is the result of their activation. It
appears obvious that this must be an ancient system for it to be composed of
such a high number of orchestrated components, and in such a pan specific
manifestation, with a particular bent toward extending organism life length by
mimicking, or attempting to mimic the juvenile period and imposing it on
adult cells. The impact seems to be most pronounced on metabolic efficiency.
For one thing, the entire metabotype of the aging cell shifts over to a juvenile
metabotype when SIRT1 turns on. Surprisingly, homeostatic non-dividing
adult cells undergo mitochondrial biogenesis, in which, old inefficient
mitochondria become replaced by new efficient mitochondria. This is not just
inefficient mitochondria dividing to produce more inefficient mitochondria, as
is the case in aging adult replicating cells and cancer cells, but an actual
biogenesis of young behaving mitochondria yielding cells of a juvenile
metabotype. This probably, helps account for the reduction in insulin
resistance, by dramatically reducing the cell’s glucose dependency, and for a
host of rejuvenating effects caused by rebalancing catabolic energy
production, and, probably ROS reduction, or ROS scavenging. There is
support for the notion that mitochondrial biogenesis alone can have very
substantial impact upon cell rejuvenation, as direct elicitors of mitochondrial
biogenesis (PGC1 alpha) in mice, reduces muscular wasting and visual
impairment. We can expect to see an explosion of research in this area, now
that mitochondria are known to be so vital to cellular health, far more so, than
was thought, in the traditional sense.

However, the amount of caloric restriction needed to institute these

alterations, is draconian; tantamount to humans trying to live for 110 years in
hell-like semi-starvation vs. living in relative comfort for only 80 years. But,
stay tuned. There are indications that caloric restriction gene systems might
be, in good part, chemically inducible.

There is a new kid on the block, called resveratrol. Over the last decade, or so,
resveratrol has become hailed as an anti-aging super anti-oxidant. Originally
touted as a component of the lycopene/tomato, omega 3/olive oil and
resveratrol/ red wine Mediteranian diet, resveratrol has emerged as a heavy
weight contender in its own right, and for some good reason. Shotgun gene
affinity experiments, which measure up and down regulation of over 20,000
genes at a time by measuring mRNA gene transcript annealing to DNA
sequence leaders, demonstrate that more than` 745 genes are regulated by
resveratrol in the same fashion as done by caloric restriction, in mouse tissues.
In fact, the number and direction of regulation shows a remarkable 99.7%
concordance with caloric restriction, the remaining 0.3%, which somehow
bypasses SIRT1. Similar results were obtained with other tissues. The effect
even occurs in fat mice, even though the jury is still out on its impact on
longevity in these animals. Importantly, resveratrol initiates mitochondrial
biogenesis and a laundry list of cytological and tissue regenerative affects
similar, if not identical to caloric restriction. The probability of such a high
gene activation concordance is so minute as to border on the incalculable. The
fact that it does so via some kind of bypassing of the putative cascade
initiation control genes SIRT1 and Insulin-like Growth Factor (IGF), is
initiating some rather heated discussion between pharmaceutical companies
with vested interests on both sides of the resveratrol gene control issue, and
whether it is a true caloric restriction mimetic. Based upon the impacts seen
from mitochondrial biogenesis, I would guess that a very sizeable percentage
of the genes induced by SIRT1 or resveratrol, are devoted to mitochondrial
biogenesis and its consequences on aging and cancer cell metabotype.

There can no longer be a doubt that many, if not most cancer cells have some
entrenched metabotype that is fundamentally more fixated than in normal
cells. Briefly put, in-vitro studies of glucose mimicking glycolytic blocking
agents such as 2-deoxy glucose and 5-thio D glucose, can kill up to 99.999%
of cancer cells in a few hours while leaving normal dividing cells alive, with
the cancer cells being radiation sensitive and the normal cells being radiation
insensitive. These agents slam glycolytic ATP production to a halt leaving
only pyruvate starved mitochondrial ATP production to remain. Being
glycolysis end product dependant, much more so than most normal cells,
cancer cells die, while normal cells can utilize, or switch off to alternate fuels,
more readily. Unfortunately, these results don’t translate to in-vivo use
because the therapeutic dose is to close to the contraindicating dose, probably
because they are glucose analogues. Brain cells are highly dependant upon
glucose, for instance.
 In cancer cells, glycolysis often produces many times more pyruvate than the
inefficient mitochondria can assimilate, so the excess pyruvate is converted to
lactate for cell export to the liver, where it is converted to glucose for re-
export to the tumor, in a closed loop system called the cori cycle. Other cells,
such as hypoxic cells and low oxidative fast twitch muscle cells, can
reversibly utilize the cori cycle, while cancer cells are much more entrenched
in the cori loop. Hyperacidification by lactic acid export blockers has
instituted differential cancer cell kill, in many cases, but usually fails to finally
eradicate tumors that adapt to hypoxic conditions. Also, it is necessary, as
with glucose feedstock blocking, to thread a narrow path between efficacy and
contraindication.

Bypassing cell glucose importation and lactate export systems may

efficaciously demonstrate an alternative outcome. A recent strategy, applied
across a broad spectrum of tumors in mice, utilizes dichloroacetate (DCA) to
block a fetal puruvate kinase (FPK) enzyme in these tumors, thus
renormalizing metabolic flow away from anabolism, and stopping cell growth.
Apparently the switchout from adult PK to FPK is very common in such
mouse tumors. Unfortunately, the results are so promising, and
dichloroacetate, being a common unpatentable reagent, have led to a growing
illegal market for dichloroacetate, among desperate cancer victims.

Caloric restriction and antioxidants both have impact on cancer incidence and
severity. Carcinogenesis is both delayed, and once initiated, growth rates are
slowed. It would be very interesting to see the results of resveratrol and
dichloroacetate together. One could postulate a synergistic effect, in which
dichloroacetate renormalizes glycolytic flow, and resveratrol reinforces the
effect via anti-oxidation and/or mitochondrial biogenesis. Then again, things
may not be so simple. The FPK system might not work the same way in
humans as it does in mice, because people using DCA are reporting huge cell
kill rates in their tumors and the associated dead cell load problems, indicating
that DCA is acting more as a blocking agent than a pathway switching agent.
If true, it could all be for the better, as kill is far more preferred than
renormalization, as it allows the therapy to be stopped, at some point, instead
of having to remain interminable. Besides, long term use of DCA is
hepatotoxic.

The DCA effect has resurrected the name of Otto Warburg, and rightly so. For
the first time in 50 years, metabolism is again, moving into the forefront of
cancer research. Please note that the metabotype referred to herein is only one
of a number of required steps in cellular transformation to a full blown
metastatic cancer. However the metabotype now assumes a required status,
uniquely different from other dividing cells, such as fetal cells and adult stem
cells. Cancer, it seems, is not just related to aging, but might be a mutated
stepwise progression form of it. These integrated set of results give rise to an
interesting evolutionary perspective.

A Short Evolutionary Perspective

Now that we have whole genomes, from bacteria to human beings sequenced,
many obvious first questions arise. One of the most tempting is: How did
something as complex as a mammal, with well over 200 different cell types,
operating in a highly integrated symphony, evolve to have a few times more
genes than a eukaryotic single cell life form, such as yeast. What makes this
even more puzzling, is that our 25,000, or so, genes seem mostly to have
arisen from a couple of duplicating of, perhaps a 5,000-7,000, or fewer, gene
containing archaic genome. One notion is that duplicating whole genomes
creates a regulatory opportunity for multicellularity, as it allows for
differential expression of each genome in each of two cells that fail to separate
after division, allowing each cell to perform unique tasks that enhance the
survival of each, together. Over time, this leads to multicellularity, and simple
repeating body plans, such as segmented worms, developing segments into
complex body parts and plans, such as insect mandibulata or mammalian inner
ear etc. Over time, gene remnants fail to transcribe, sister chromatids cross
over millions of times, viral transposons shuffle DNA around, evolutionary
pressures select for gene gains, losses and mutations, gene regulatory systems
advance, more and more cell types and there regulatory systems evolve, and
create an ever diversifying array of body plans etc, until, at present times, the
archaic genome, although readily statistically apparent, is barely still visible in
the mish-mash. This is not quite like doing more with less, as it is like doing a
helluva lot more with more of the same. Not to take anything away from
Darwin, but nature does seem to ‘learn’ from, or pseudo-repeat, her successes.
The genome duplication demonstrates how a gigantic array of regulatory
elements in multicellular organisms can take an existing array of genes to new
heights of diverse expression and function without actually having to create
real ‘new’ genes.

The new field of epigenetics is a case in point, because it pulls an apparent

Lamarckian ‘acquired characteristics’, end around, the more temporally linear
notion of Darwinian survival selected fitness traits resulting from serially
generational genomic mutagenesis. Briefly, the epigenome responds to
environmental conditions, such as famine in the sex cells of the parents, by
selectively methylating genes to up and down regulate them so that these
regulatory systems are passed on to the offspring, purportedly to pre-adapt the
offspring to the existing environmental conditions experienced by the parent.
The number of possible variations of epigenetics dwarfs the size of the
genome itself. This area of research is in its infancy, but one of its discoveries
is serendipitous with this narrative. One epigenomics experiment shows that
feast and famine leave a differential impact on sons and daughters that
significantly affects their life expectancy. More so, the epigenetic signature
passes through several generations before it fades out. From a Darwinian
perspective, the only way to have the appearance of a Lamarckian outcome, is
for pre-adaptation to have arisen from numerous, widely spaced, earlier or
many, highly selective, recent exposures to a phenomenon. The more gene
systems affected in a more orchestrated fashion, the more ancient and more
protracted the system must be. Open ended circular arguments apply to life,
because life has ‘been there’ before, especially in the case of famine, and
because, successful gene systems replicate and fan out over time. In regards to
the subject focus of this document, there appears to be a much more ancient
and a much larger and much more controlled famine gene manipulation
system than epigenetics at work.
I broached these topics to put forth the notion that ancient systems, such as
intermediary metabolism, mitochondrial function and anciently occurring
phenomena, such as feast and famine, grew along with the growth of the
metabolic pathways themselves. Whole suites of genes are involved with their
control, but over time the system evolves to arrive at a relatively few fixed
sets of ‘tried and true’ tactics that have become, more or less, successful fixed
response systems. Other, more flexible life components, such as length of time
as a juvenile, or length of life as an adult, may be operating in an intermediate
time spectrum, as they are responding to phenomena much longer than
epigenetics, but much shorter than raw metabotype, although all three systems
outlined, here, from a feast and famine standpoint, can leave their mark upon
metabotype expression.

My point is this: even though there are hundreds of cell growth factors, the
unique mixes, of which, tell each cell type when and how fast to grow and
divide, there seems to be a veritable paucity of system(s) setting mitochondrial
biogenesis in motion, with its apparent requisite reestablishment of the 5/95
glycolytic/Krebs ATP production ratios, and the resultant catabolic/anabolic
rate shift. This rejuvenating biogenic system appears to paint with a broad
brush, saying essentially ‘on’ in juvenile cells, ‘off’ in adult cells, and when in
the ‘off’ position, mitochondria slowly age (as in humans), or rapidly age (as
in mice), toward the inefficient cancer-like metabotype. This becomes a
precancerous, or hyperplasic condition awaiting growth signal and signal
related mutations, to transform it into the cancerous state. Amazingly, the
juvenile system can be turned back on in adult cells, but only when the
organism is in dire straights of a type recurrent from the dawn of metabolic
evolution. Interestingly, most mammals fail to ovulate if they are not carrying
enough fat supply, so caloric restriction carries extended life in exchange for
the loss procreative capacity, thus allowing species replacement numbers to be
deferred to a more nutrient plush future. Conversely, when feast times return,
fat storage ensues, high ATP production efficiency is no longer a life
sustaining requirement, and after a few procreative cycles, the now shorter
lived adults, are disposed of. It is rather interesting that nature has also found a
mechanism for failing mitochondria to act in a metabolic default toward cell
growth, the caveat of it, being linked to life saving energy supply availability,
on the one hand, and the potential to assist in the death dealing carcinogenic
process, on the other.

There does not seem to be any real overt need to modify this ancient system,
save to change the length of the juvenile and adult stages to suit the
environmental conditions demanding such. These life stage length differences
between mice, dogs, horses, humans etc. are independent of metabolic rate,
body size, the brain to body ratio, just as is the time from birth to the elevated
cancer risk incidence curve. Instead, they seem more tied to the mitochondrial
decay rate as per life expectancy. It is as if the vast array of environmental
constraints set the necessary life length of the organism and that cellular
metabolism doesn’t have a very difficult time complying by setting juvenile
and adult stage length as needed. Of mice and men; old at age three or old at
age 100; several thousand percent difference in life expectancy and 99%
identical metabolic pathways. This gives argument to the notion of not what,
or how you do something, but when you do it. It probably works the same in
mice as it does in men, with the switches being turned off and on at different
times and at different strengths. The central metabolic system, and its core
regulatory elements haven’t changed much since the most advanced animal
was a sponge.

In retrospect, glycolysis and aerobic respiration are systems entrenched in

times so ancient, that it was before earth even had an oxygen atmosphere.
When early oxygen producing photosynthesis first evolved in the ocean,
anaerobic glycolytic bacteria, in one or more branches of life, began
oxidatively extracting energy from glycolytic end products by removing
hydrogen and adding oxygen in a stepwise process. Eventually this oxidative
process evolved to extract the maximum obtainable energy by extending the
oxidative chain all the way to the natural carbon dioxide and water, as waste
product, endpoint. A symbiosis was formed when a nucleus containing
archaeobacterial species housed an aerobic bacterial life form within itself,
probably providing predatory protection in exchange for abundant energy.
This is a relationship similar to that which exists today when coral polyps
house algae within themselves. Before the first sponge-like simple
multicellular life forms evolved, a nearly billion year relationship between the
host and its symbiont had been maturing. Eventually, this relationship became
so inextricable that, by the time that the first eukaryotic fungal/animal
precursor cell arrived, the aerobic symbiont had been mostly reduced to the
status of slave to its host, giving up most of its genome in exchange for
providing critical energy producing advantages to its host (except for the
notable, and rather disturbing distinction of retaining a host cell killing
command center, called apoptosis). We call this relic, the mitochondrion.
Today, the animal cell mitochondrion contains only the genes for its
respiratory chain and some of its protein synthesis and replication machinery,
cell suicide mechanism, included. All other ancient pre-mitochondrial
functions are now sequestered in the DNA of the cell nucleus, and their gene
products are imported into the mitochondrion from the cytoplasm. This is true
in even though most primitive animal life forms, and in even earlier life
forms, such as single celled yeast. Thus, aerobic and anaerobic systems were
intricately enmeshed, in terms of each others operation and control
mechanisms for a period of time longer than the existence of the Kingdom
Animalia. It is no small wonder that we see the birthings of a SIRT1 control
system in organisms that predate the animal kingdom.

It seems incredible how both deeply entrenched and unchanged the caloric
restriction response system has remained over time. We find it throughout the
entire animal kingdom, from worms, through insecta, up the vertebrate tree,
from ancient to modern life forms. The system is also remarkable in its
versatility, and in its short term, intermediate term and its long term
applications to organismic, species and Kingdom survival. When we look at
mammals, in our attempts to utilize models more akin to our human selves,
we see the sweeping SIRT1 system applies to every tissue and organ tested.
Although the SIRT1 cascade ‘rejuvenation’ is nowhere near to being confused
with the word ‘immortalization’, it is an apt nomer, nonetheless. Among other
things, the system institutes production, via mitochondrial biogenesis, of
efficient catabolism, which yields survival advantages during nutrient energy
shortages, and yields energy efficient rapid growth during the growth factor
directed juvenile period of rapid body expansion. It reverts adult cells to a
juvenile state, inasmuch as it can within mutagenic constraints, causing a true
30% to 80% increase in life span, depending upon which model is used.

After puberty, under ad libitum conditions, it becomes pretty clear that the
SIRT1 system is, not only, not needed, but is not utilized, probably because it
becomes disadvantageous in the statistical evolutionary survival sense. In the
adult phase it, appears that cell senescence is an obligatory rule that can only
be violated if survival of the next generation is at risk, due to the risk to the
survival of the present generation, via starvation. Thus it does not matter if,
under feast conditions, with procreation accomplished, that cancer becomes a
natural outcome of declining metabolic efficiency, combined with the other
required changes in cell growth control systems. In the adult system,
inefficient catabolism diverts anabolic processes toward fat storage and
pushes cells to become more dependant upon the cori cycle as they become
more fuel dependant upon glucose by diverting excess pyruvate to lactic acid
for export to the liver for gluconeogenesis. Such cells are metabotype ‘pre-
adapted’ for cancer, as well as for organism aging or death, due to disrepair,
because it really doesn’t matter how the organism is disposed of, as long as it
is disposed of, usually by predation or disease, of course, due to its aged and
weakened condition, Only under nutrient fuel paucity conditions does this
natural decay process seem to be interrupted. The fact that SIRT gene
knockout mice age rapidly and die very early, only underscores these
conclusions.

SIRT1 up regulation has just the opposite effect. Mitochondrial biogenesis

occurs, the metabotype reverts to a juvenile metabotype , adult life span
lengthens, cancer incidence declines, regain of tissue function, such as that of
muscle, liver, neurological etc. takes place and total organismal energy output
efficiency and general vitality ensues. The fact that resveratrol mimics this
general condition, is a most fortunate discovery. The fact that the
pharmaceutical giants are at war over resveratrol and its caloric restriction
mimetic capacities, shows just how far human folly can escalate when so
many billions of dollars are at stake, especially if the leading candidate drug is
unpatentable. Although the affinity chip data indicate that resveratrol bypasses
SIRT one and IGF, the fact that it activates 99+% of its downstream cascade,
in at least three tissue types, speaks volumes. The physiological and
cytological studies, which support the rejuvenative effects, arise from too
many sources for it to be a collusion, accident or folly.

Regardless, continued research on actual SIRT1 activators, resveratrol, its

analogues or SIRT1 downstream cascade activators, these recent
breakthroughs should spur a huge amount of basic research in both aging and
cancer, and should, especially, reinvigorate a long overdue reinvestigation of
the therapeutic efficacy of cancer cell intermediary metabolic interventions.
This is particularly true, in light of the preliminary results from the
employment of DCA, and the pivotal position between glycolysis and the
mitochondrion occupied by pyruvic acid.

At the present time, the list of anti-aging supplements that need to be taken to
ward off the plethora of cell systems in decline in aging organisms, is
mammoth, to put it lightly. Many of these supplements (but, by no means all,
such as, vitamins, anti-oxidants, phytonutrients etc.) are provided abundantly
by juvenile cells, but are produced in progressively lessening quantities by
aging organs, tissues and cells. However, by turning on a SIRT1 rejuvenation
gene cascade, it appears that life extension occurs in the absence of such
supplementation. This fact alone has incredible implications, because it proves
that the vast majority of youthful cell vigor information, remains intact, and is
recoverable with the right inducement(s). The implications are gargantuan,
because it declares with a delightful certainty, that normal aging is not, so
much so, a process of irreversible decay, as it is a form of highly controlled
preplanned obsolescence. The investigation of modified supplementation, in
tandem with rejuvenation gene cascade initiation and mitochondrial
biogenesis control systems, could offer a fertile field for the cultivation of a
broad area of applications. It is becoming progressively more clear that
glycolytic and mitochondrial catabolic control systems, in and of themselves,
have deep impacts upon cellular function, outside the governance of cell
growth control signaling pathways. The notion that such a system has, at its
roots, control over such sweeping concepts, as aging, biogenesis, rejuvenation
and cancer, is almost stupefying. After all, these systems, and their core
control elements, predate multicellularity and the need for differential cell
type control.

Afterword

I know that this a brief sketch, but I do believe that the fundamental elements
and their chief interactions have been presented. I know that I cast a very
broad net in an age of narrow specialists, so I beg the reader to leave the
details for future investigation, while relegating present thoughts to seeing
‘the big picture’ in the context of a renaissance mind.

Acknowledgement

I thank Michael Wolfson for his stimulating conversations on life extension,

which spurred my investigations on this topic and formed the relationship of
ideas with my previous work. Without his unique perspective and irrepressible
humor, I doubt that I would have undertaken this task.

Gregory S. Bambeck Ph.D.

Kent, Ohio, USA 44240

e. mail: gregorybambeck@yahoo.com
February 21, 2010