related to
intracellular hydrogen peroxide generation
only be obtained by
intravenous administration of AA
Preferentially kills neoplastic cells
Is virtually non-toxic at any dosage
Does not suppress the immune system, unlike most chemotherapy
agents
Increases animal and human resistance to infectious agents by
enhancing lymphocyte blastogenesis, enhancing cellular immunity,
strengthening the extracellular matrix, and enhancing bactericidal
activity of neutrophils and modulation of complement protein
Strengthens the structural integrity of the extracellular matrix
which is responsible for stromal resistance to malignant invasiveness
1969, researchers at the NCI reported AA was highly toxic to Ehrlich
ascites cells in vitro
In 1977, Bram et al reported preferential AA
toxicity for several malignant melanoma cell lines, including four
human-derived lines
Noto et al reported that AA plus vitamin K3
had growth inhibiting action against three human tumor cell lines at
non-toxic levels
Metabolites of AA have also shown antitumor activity in
vitro
The AA begins to reduce cell proliferation in the tumor cell
line at the lowest concentration, 1.76 mg/dl, and is completely cytotoxic to
the cells at 7.04 mg/dl
the normal cells grew at an enhanced rate at the low dosages (1.76
and 3.52 mg/dl)
preferential toxicity of AA for tumor
cells. >95% toxicity to human endometrial adenocarcinoma and pancreatic
tumor cells (ATCC AN3-CA and MIA PaCa-2) occurred at 20 and 30 mg/dl,
respectively.
No toxicity or inhibition was demonstrated in the normal,
human skin fibroblasts (ATCC CCD 25SK) even at the highest concentration of
50 mg/dl.
the use of very high-dose intravenous AA for the treatment of
cancer was proposed as early as 1971
Cameron and Pauling have published
extensive suggestive evidence for prolonged life in terminal cancer patients
orally supplemented (with and without initial intravenous AA therapy) with
10 g/day of AA
AA, plasma levels during infusion were not monitored,
the long-term, oral dosage used in those experiments (10 g/day),
while substantial and capable of producing immunostimulatory and
extracellular matrix modulation effects, was not high enough to achieve
plasma concentrations that are generally cytotoxic to tumor cells in
culture
This low cytotoxic level of AA is exceedingly rare
5 — 40 mg/dl of AA is required in vitro to kill 100% of tumor
cells within 3 days. The 100% kill levels of 30 mg/dl for the endometrial
carcinoma cells and 40 mg/dl for the pancreatic carcinoma cells in Figure 2
are typical
normal range (95% range) of 0.39-1.13 mg/dl
1 h after beginning his first 8-h infusion of 115 g AA (Merit
Pharmaceuticals, Los Angeles, CA), the plasma AA was 3.7 mg/dl and at 5 h
was 19 mg/dl. During his fourth 8-h infusion, 8 days later, the 1 h plasma
level was 158 mg/dl and 5 h was 185 mg/dl
plasma levels of over 100 mg/dl have been maintained in 3
patients for more than 5 h using continuous intravenous infusion
In rare instances of patients with widely disseminated
and rapidly proliferating tumors, intravenous AA administration (10 — 45
g/day) precipitated widespread tumor hemorrhage and necrosis, resulting in
death
Although the outcomes were disastrous in these cases, they are
similar to the description of tumor-necrosis-factor-induced hemorrhage and
necrosis in mice (52) and seem to demonstrate the ability of AA to kill
tumor cells in vivo.
toxic effects of AA on one normal cell line were observed at 58.36 mg/dl and
the lack of side effects in patients maintaining >100 mg/dl plasma levels
Although it is very rare, tumor necrosis, hemorrhage, and subsequent
death should be the highest priority concern for the safety of intravenous
AA for cancer patients.
Klenner,
who reported no ill effects of dosages as high as 150 g intravenously over a
24-h period
Cathcart (55) who
describes no ill effects with doses of up to 200 g/d in patients with
various pathological conditions
following circumstances: renal
insufficiency, chronic hemodialysis patients, unusual forms of iron
overload, and oxalate stone formers
Screening for red cell glucose-6-phosphate
dehydrogenase deficiency, which can give rise to hemolysis of red blood
cells under oxidative stress (57), should also be performed
any cancer therapy should be started at a low dosage to ensure
that tumor hemorrhage does not occur.
patient is orally supplementing between infusions
a scorbutic rebound effect can be
avoided with oral supplementation. Because of the possibility of a rebound
effect, measurement of plasma levels during the periods between infusions
should be performed to ensure that no such effect takes place
Every effort
should be made to monitor plasma AA levels when a patient discontinues
intravenous AA therapy.
These eight distinct cancer types included: DCIS, breast (ER(+) and ER(-)), ovarian, prostate, lung, and pancreatic carcinomas, as well as melanoma and glioblastoma. Doxycycline was also effective in halting the propagation of primary cultures of CSCs from breast cancer patients, with advanced metastatic disease (isolated from ascites fluid and/or pleural effusions)
Doxycycline behaves as a strong radio-sensitizer, successfully overcoming radio-resistance in breast CSCs
cancer cells can indeed escape the effects of Doxycycline, by reverting to a purely glycolytic phenotype. Fortunately, the metabolic inflexibility conferred by this escape mechanism allows Doxycycline-resistant (DoxyR) CSCs to be more effectively targeted with many other metabolic inhibitors, including Vitamin C, which functionally blocks aerobic glycolysis
Vitamin C inhibits GAPDH (a glycolytic enzyme) and depletes the cellular pool of glutathione, resulting in high ROS production and oxidative stress
DoxyR CSCs are between 4- to 10-fold more susceptible to the effects of Vitamin C
Doxycycline and Vitamin C may represent a new synthetic lethal drug combination for eradicating CSCs, by ultimately targeting both mitochondrial and glycolytic metabolism
inhibiting their propagation in the range of 100 to 250 µM
metabolic flexibility in cancer cells allows them to escape therapeutic eradication, leading to chemo- and radio-resistance
used doxycycline to pharmacologically induce metabolic inflexibility in CSCs, by chronically inhibiting mitochondrial biogenesis
This treatment resulted in a purely glycolytic population of surviving cancer cells
DoxyR cells are mainly glycolytic
MCF7 cells survive and develop Doxycycline-resistance, by adopting a purely glycolytic phenotype
Cancer stem cells (CSCs) are thought to be the “root cause” of tumor recurrence, distant metastasis and therapy-resistance
the conserved evolutionary similarities between aerobic bacteria and mitochondria, certain classes of antibiotics inhibit mitochondrial protein translation, as an off-target side-effect
Vitamin C was more potent than 2-DG; it inhibited DoxyR CSC propagation by > 90% at 250 µM and 100% at 500 µM
IC-50
DoxyR CSCs are between 4- to 10-fold more sensitive to Vitamin C than control MCF7 CSCs
Berberine, which is a naturally occurring antibiotic that also behaves as an OXPHOS inhibitor
treatment with Berberine effectively inhibited the propagation of the DoxyR CSCs by > 50% at 1 µM and > 80% at 10 µM.
Doxycycline, a clinically approved antibiotic, induces metabolic stress in cancer cells. This allows the remaining cancer cells to be synchronized towards a purely glycolytic phenotype, driving a form of metabolic inflexibility
Doxycycline-driven aerobic glycolysis
new synthetic lethal strategy for eradicating CSCs, by employing i) Doxycycline (to target mitochondria) and ii) Vitamin C (to target glycolysis)
Doxycycline inhibits mitochondrial biogenesis and OXPHOS,
hibits glycolytic metabolism by targeting and inhibiting the enzyme GAPDH
CSCs act as the main promoter of tumor recurrence and patient relapse
a metabolic shift from oxidative to glycolytic metabolism represents an escape mechanism for breast cancer cells chronically-treated with a mitochondrial stressor like Doxycycline, as mitochondrial dys-function leads to a stronger dependence on glucose
Vitamin C has been demonstrated to selectively kill cancer cells in vitro and to inhibit tumor growth in experimental mouse models
many of these actions have been attributed to the ability of Vitamin C to act as a glycolysis inhibitor, by targeting GAPDH and depleting the NAD pool
here we show that DoxyR CSCs are more vulnerable to the inhibitory effects of Vitamin C, at 4- to 10-fold lower concentrations, between 100 to 250 μM
concurrent use of Vitamin C, with standard chemotherapy, reduces tumor recurrence and patient mortality
after oral administration, Vitamin C plasma levels reach concentrations of ~70-220 μM
intravenous administration results in 30- to 70- fold higher plasma concentrations of Vitamin C
pro-oxidant activity results from Vitamin C’s action on metal ions, which generates free radicals and hydrogen peroxide, and is associated with cell toxicity
it has been shown that high-dose Vitamin C is more cytotoxic to cancer cells than to normal cells
This selectivity appears to be due to the higher catalase content observed in normal cells (~10-100 fold greater), as compared to tumor cells. Hence, Vitamin C may be regarded as a safe agent that selectively targets cancer cells
the concurrent use of Doxycycline and Vitamin C, in the context of this infectious disease, appeared to be highly synergistic in patients
Goc et al., 2016, showed that Doxycycline is synergistic in vitro with certain phytochemicals and micronutrients, including Vitamin C, in the in vitro killing of the vegetative spirochete form of Borrelia spp., the causative agent underlying Lyme disease
Doxycycline, an FDA-approved antibiotic, behaves as an inhibitor of mitochondrial protein translation
CSCs successfully escape from the anti-mitochondrial effects of Doxycycline, by assuming a purely glycolytic phenotype. Therefore, DoxyR CSCs are then more susceptible to other metabolic perturbations, because of their metabolic inflexibility