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Out of Warburg Effect: an effective cancer treatment
targeting the tumor specific metabolism and
dysregulated pH
Laurent Schwartz1*, Thomas Seyfried2, Khalid O. Alfarouk3, 4, Jorgelindo Da Veiga
Moreira5, Stefano Fais6
1Assistance Publique des Hôpitaux de Paris France
2Biology Department, Boston College Chestnut Hill, MA, USA
3Al-Gahd International College for Applied Medical Sciences, Al-Madinah Al-
Munawarah, KSA
4Center for Evolution and Cancer, University of California San Francisco, USA
5 Laboratoire d’informatique Ecole Polytechnique Palaiseau France
6Department of Therapeutic Research and Medicine Evaluation, National Institute of
Health, Rome, Italy
* Corresponding Author:
Laurent Schwartz, MD
Assistance Publique des Hôpitaux de Paris
3 Avenue Victoria, 75004 Paris, France
Tel: +33681899030
E mail: dr.Laurentschwartz@gmail.com
1
Abstract
As stated by Otto Warburg nearly a century ago, cancer is a metabolic
disease, a fermentation caused by malfunctioning mitochondria, resulting in
increased anabolism and decreased catabolism. Treatment should, therefore,
aim at restoring the energy yield. To decrease anabolism, glucose uptake
should be reduced (ketogenic diet). To increase catabolism, the oxidative
phosphorylation should be restored. Treatment with a combination of α-lipoic
acid and hydroxycitrate has been shown to be effective in multiple animal
models. This treatment, in combination with conventional chemotherapy, has
yielded extremely encouraging results in glioblastoma, brain metastasis and
lung cancer. Randomized trials are necessary to confirm these preliminary
data. The major limitation is the fact that the combination of α-lipoic acid and
hydroxycitrate can only be effective if the mitochondria are still present and/or
functional. That may not be the case in the most aggressive tumors. The
increased intracellular alkalosis is a strong mitogenic signal, which bypasses
most inhibitory signals. Concomitant correction of this alkalosis may be a very
effective treatment in case of mitochondrial failure.
Keywords: Warburg’s effect, unification theory, ATP, metabolic treatment,
intracellular alkalosis, ketogenic diet.
2
Introduction
In the early 1920’s Otto Warburg demonstrated a unique feature of cancer
cells, namely an increased uptake of glucose and secretion of lactic acid by
cancer cells, even in the presence of oxygen (e.g. the aerobic glycolytic
phenotype) [1,2]. This aerobic fermentation is the signature of cancer [3].
Warburg also noticed a concomitant decreased number of mitochondria
(grana) [4]. In normal, differentiated cells, the yield of a molecule of glucose is
34 ATP. ATP is derived mostly from oxidative phosphorylation which takes
place in the mitochondria [5,6]. In the absence of mitochondria the energy
yield drops to two molecules of ATP per molecule of glucose [5,6]. As stated
by Warburg in the 1920’s, in cancer cells there is a decreased efficacy of the
mitochondria resulting in lesser yield. Despite increased glucose uptake, there
is a 50% drop in ATP level in human colon cancer cells compared to adjacent
benign cells [7]. This decrease in ATP is a consequence of impairment of the
oxidative phosphorylation [6–9].
To compensate for the decreased energy yield, the cell increases its glucose
uptake [7,10]. The decreased activity of the mitochondria has many
consequences, one of which is an increased secretion of lactic acid and
another one is the activation of the pentose phosphate pathway (PPP).
Another consequence is the activation of the glutaminolysis which is
necessary for nucleic acid synthesis [6–9].
The activation of the Pentose Phosphate Pathway results from an increase in
glucose uptake with a concomitant obstacle downstream of the pentose
phosphate shunt, most probably at the level of pyruvate dehydrogenase
and/or of pyruvate kinase [2,6,11]. The increased flux in the pentose
phosphate pathway results in:
•A shift toward anabolism due to increased synthesis of NADPH that
plays a crucial role in NDPH/NADP+ ratio that determines the redox
state of the cell via removal of reactive oxygen species (ROS) and so
prevents cellular death and controls cellular fate [7,11].
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•The shift toward the pentose pathway also results in the production of
ribose-5-phosphate, required for the synthesis of nucleic acids [5].
One other crucial consequences of the mitochondrial defect is intracellular
alkalosis [7]. Tumors show a 'reversed' pH gradient with a constitutively
increased intracellular pH that is higher than the extracellular pH. This
gradient enables cancer progression by promoting proliferation, the evasion of
apoptosis, metabolic adaptation, migration, and invasion [12–15].
There is evidence that an acidic extracellular pH promotes invasiveness and
metastatic behavior in several tumor models [14,16], proteolytic enzyme
activation and matrix destruction [17–19].
In normal cells, the intracellular pH (pHi) oscillates during the cell cycle
between 6.8 and 7.3 [7]. The oscillation of the pH during the cell cycle
matches the value of the decompaction of the histones, RNA polymerase
activation, DNA polymerase activation and DNA compaction before mitosis
[7,11].
The intracellular pH of the cancer cells has been less studied. During the cell
cycle, it oscillates between 7.2 and 7.5. Intracellular alkalosis is probably a
consequence of the decreased oxidative phosphorylation resulting in
decreased secretion of carbon dioxide (CO2) and the CO2 reacts with water to
create carbonic acid. Cell transformation or enhanced cancer cell division and
resistance to chemotherapy are all associated with a more alkaline pHi [20–
23].
The very reason of the dysfunction of the mitochondria is still open for debate.
No tumor cell has yet been found with a normal content or composition of
cardiolipin, the signature lipid of the inner mitochondrial membrane. This lipid
regulates the efficiency of OxPhos [9,10].
The Warburg effect may be a direct consequence of the activation of
oncogenes [6]. Infection by an oncogenic virus or exposure to a carcinogen
inhibits the mitochondrial function and causes the Warburg’s effect [24–28].
As Warburg wrote in 1956 [4,29], “The chicken Rous sarcoma virus, which is
labeled today as a virus tumor, ferments glucose, and lives as a partial
anaerobe like all tumors.” Infection by an oncogenic virus or exposure to a
4
carcinogen inhibits the mitochondrial function and causes the Warburg’s effect
[24–28].
Alternatively, the Warburg effect results in oncogene activation and mutation.
Oncogene up-regulation is needed to drive fermentation after OxPhos
dysfunction. Mitochondria replacement will activate OxPhos and turn off
oncogenes [9,10].
This Warburg’s effect is responsible for the activation of the Pentose
phosphate pathway, glutaminolysis and subsequent anabolism [9] and all
thirty different oncogenes target and stimulate the anabolic pathways [9].
The introduction of normal mitochondria into cancer cells restores
mitochondrial function, inhibits cancer cell growth and reverses
chemoresistance [30,31]. Also the fusion of cancer cells with normal
mitochondria results in increased ATP synthesis, oxygen consumption and
respiratory chain activities together with marked decreases in cancer growth,
resistance to anti-cancer drugs, invasion, colony formation in soft agar, and «
in vivo » tumor growth in nude mice [31].
Reversing the Warburg effect:
Cytotoxic chemotherapy has had tremendous benefits for pediatric or
Hodgkin’s patients. However, for most solid tumors, while there is a sizable
response rate (complete and partial regression), there has been a limited gain
in survival.
The mechanism of action of cytotoxic chemotherapy is debated. Response to
cytotoxic drugs is assessed by PET scan. Effective treatment results in
decreased radiolabelled glucose. Most cytotoxic drugs target (indirectly) the
mitochondria and the Warburg effect [32–35]
As the Warburg aerobic glycolytic phenotype and its effects on metabolism
are key to cancer, the obvious question is whether drugs can be designed to
target it. To alleviate the Warburg effect, pyruvate should be converted into
Acetyl-CoA, which would decrease the bottleneck that results in the activation
of both the Pentose Phosphate Pathway and the glutaminolysis. The
mitochondrial yield should be increased to stimulate the synthesis of CO2 and
5
the increased secretion of CO2 would result in a decreased intracellular
alkalosis.
The combination of α-lipoïc acid and hydroxicitrate [36–39] has been reported
to slow cancer growth, in murine xenografts. This inhibition appears to be
independent of the primary tumor site and has been reproduced in different
laboratories [40,41].
The most likely mechanism of action for α-lipoic acid in its reduction of tumor
growth is the inhibition of pyruvate dehydrogenase kinase (the same target of
Dichloroacetic acid (DCA)). This enzyme inhibits the activity of pyruvate
dehydrogenase and is known to be up-regulated in cancer cells expressing
the Warburg aerobic glycolytic phenotype. Pyruvate dehydrogenase catalyses
the conversion of pyruvate to acetyl-CoA, the initial step of the final
conversion of glucose to carbon dioxide and water in the TCA cycle, with the
concomitant production of ATP. Therefore, it is reasonable to suggest that
blocking the activity of pyruvate dehydrogenase kinase will at least partially
restore the activity of pyruvate dehydrogenase, thereby increasing the flux of
pyruvate through the TCA cycle in the mitochondria, while simultaneously
reducing the production of lactic acid and most importantly decreasing the flux
in the pentose pathway shunt [9].
There are several reports of metabolic treatment utilizing a combination of α-
lipoic acid and hydroxycitrate together with conventional cancer therapy.
Starting in January 2013, metabolic treatment (α-lipoic acid/hydroxycitrate
with low doses of the chemotherapic plus Naltrexone) was offered to patients
sent home after the failure of conventional cytotoxic chemotherapy for
metastatic cancer (irrespective of the primary site) but with a Karnovsky
performance status above 70 (quantification cancer patients' general well-
being and activities of daily life) [42–46]. Of the first randomly selected eleven
patients, five were alive and reasonably well 30 months after the start of
treatment [43]. From our experience, the combination of α-lipoic
acid/hydroxycitrate with low dose Naltrexone was able to prevent tumor
recurrence in only two patients. The patients who survived for more than one
year were treated with a combination of standard care with metabolic
treatment.
6
In the update of a subsequent study, patients with multiple brain metastasis
(n=4) or glioblastoma (n=6) were treated with a combination of conventional
and metabolic treatments (α-lipoic acid/hydroxycitrate) as well as ketogenic
diet. Five out of six patients with glioblastoma were alive and stable after two
years, while two of the four patients with multiple brain metastases are alive
and well three years later [46]. These glioblastoma patients had concomitant
radiation therapy and chemotherapy (temozolamide).
In another study, four lung patients (primary adenocarcinoma) were treated
with a combination of the small molecule EGFR inhibitor, Gefitinib, and
metabolic treatment (α-lipoic acid/hydroxycitrate). They all responded to
treatment and are stable one year after the start of therapy. These very
encouraging results need to be confirmed by further randomized clinical trials.
Correcting the intracellular alkalosis :
Correcting the intracellular pH may be an alternative or an adjunct to a
metabolic treatment as there is extensive literature that many effective cancer
treatments decrease the intracellular pH (pHi) [2,23,47]: literature on
increased survival support for the combined use of antacids (which prevent
proton extrusion from the tumour cells) with standard chemotherapy [15,48–
51]. Hyperthermia [50] decreases the intracellular pH.
There are two possibilities to decrease the intracellular pH (pHi). The first is a
calorie restricted ketogenic diet [8], which will reduce the availability of
glucose, the principal metabolite for glycolysis and the PPP. This diet will
result in increased levels of acidic ketone bodies that cannot be metabolized
by the cancer cells (while normal cells are able to do so) and this probably
results in a decrease of the intracellular pH. Similarly, it has been proposed
that an acid diet in general might lead to a supply of lactate (exogenous
lactate) that might lead to regression of the tumour growth [51].
The rapid turnover of aberrantly dividing cancer cells within the tumor mass,
sugar fermentation and increased ATP hydrolysis, all lead to the production
and release of large amounts of protons into the extracellular compartment
[1,52]. The H+ accumulation, in turn, leads to the progressive setting of a
highly hostile microenvironment, mostly characterized by low pH. The acidic
7
microenvironment produces a sort of “selective pressure,” that, in fact, favors
the cells that are best adapted to survive in these hostile conditions. The
hostility of the microenvironment is increased by hypoxia and low nutrient
supply, making it virtually impossible for “normal” or more differentiated cells
to survive in these unsuitable conditions. To thrive in such an unfavorable
microenvironment, tumor cells must develop systems to actively extrude
excess protons [48,52–55]. These mechanisms include the V-ATPase, the
Na+/H+ exchanger (NHE), monocarboxylate transporters (MCTs) and carbonic
anhydrase 9 [52]. Of course, we must consider these proton exchangers as
key “survival options” for cancer cells, and, therefore, depriving cancer cells of
their function should inevitably lead to a rapid cell death due to internal
acidification, as it has been shown for different cancer cell types in pre-clinical
settings [56,57]. Unfortunately, it is very hard to convince clinical oncologists
to use this approach in a first line treatment of cancer patients. However, it
was possible to increase the proof of principle that at least proton pump
inhibitors may be included in new anti-cancer protocols together with the
standard treatments. Clinical evidence has been provided in human tumor
patients with either osteosarcoma [58], breast cancer [59] or GI cancers [60].
Further evidence has been accumulated in clinical trials have been performed
in domestic animals with spontaneous tumors of different histotypes, with very
encouraging results [61,62]. Altogether, these pre-clinical and clinical results
suggest that a way to counteract intracellular alkalinity might be to inhibit the
above mechanisms designed to avoid intracellular acidification.
Conclusion
Today, cancer is thought to be a set of very complex diseases with thousands
of different mutations. That apparent complexity has led to personalized
medicine. However, modern biology has confirmed the universality of the
Warburg aerobic glycolytic phenotype. Furthermore, the fact that the
combination of α-lipoïc acid and hydroxycitrate slows down cancer growth in
every tumor model studied to date suggests that at least some targets are the
same in a large spectrum of tumors.
8
The Warburg effect can be caused by two intertwined phenomena. The first is
a metabolic bottleneck, which can be corrected. The second is the
destruction/disappearance of the mitochondria.
Cytotoxic drugs injure or even destroy the mitochondria [32–35]. At the time of
failure of chemotherapy, there is a sharply increased glucose uptake such as
seen on PET scan [63]. Resistance to chemotherapy has been correlated with
mitochondrial functionality (perhaps oxidative phosphorylation) [64] and
alkaline pHi [1,23]. The oxidative phosphorylation is further reduced; the pH is
more alkaline, and the PPP is activated. Cancer grows relentlessly, and
further chemotherapy is usually ineffective [9].
Further studies should assess the role of low dose chemotherapy together
with a combination of metabolic treatment and treatments to lower the
intracellular pH.
Conflict of Interest statement:
The authors declare that they have no competing interests.
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