**11. What to do next?**

considered inconsequential for long time. This erroneous view, has been recently reviewed, as data have emerged, demonstrating that leukemic cells are sensitive to the oxygen tension, and hypoxia influences leukemic cell proliferation, differentiation, and resistance to chemo-

Migliavacca and Coll. have recently demonstrated oncogenic function of HIF-1α, in the M5 Fab subtype of AML [99]. In particular, the authors have demonstrated that in M5 AML, HIF-1α mediates the ability of leukemic cells to migrate and invade extramedullary sites. The same group has demonstrated that PML-RARα and other fusion proteins involved in the pathogenesis of acute promyelocytic leukemia (APL) behave as transcriptional coactivators of HIFs, and both HIFs and PML-RARα enhances the progression of APL, by promoting cell

Further investigations [102] have demonstrated that HIF-1α plays critical and pleiotropic

Globally, elevated levels of HIF-1α have been reported in AML [103–106], APL [100], acute lymphoblastic leukemia (ALL) [107], and chronic myelogenous leukemia (CML) [108, 109]. Furthermore, HIF-1α overexpression conditions disease severity and outcome in both AML

Overall, the available data show that hypoxia and HIF-mediated signaling play a crucial role in leukemia, and targeting HIF with specific drugs or natural inhibitors, such as Vitamin C,

Decreased TET expression and loss of 5hmC have been observed in a wide variety of solid tumors, as well as in many hematological malignancies, including acute myeloid leukemias,

Recent experimental studies suggest that pharmacological dose of Vitamin C may represent a potentially important strategy in leukemia therapy through a stimulatory effect on TET2 activation and restoration in leukemic cells. Vitamin C is a co-factor of TET2 enzyme and is capable of interacting with the catalytic domain of TET2, enhancing the enzymatic oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) [115]. This epigenetic modulation elicited by Vitamin C is able to improve the generation of pluripotent stem cells [116]

Two recent studies explored the possible epigenetic effects of Vitamin C on leukemia models, mediated by activation and restoration of TET2 activity in leukemic cells. In the first one, authors used a murine model of IDH1-dependent acute myeloid leukemia [118], and 2-phosphate l-ascorbic acid (Asc 2-P). Asc 2-P, unlike native Vitamin C, remains oxidatively stable under standard cell culture conditions [119], and possesses the same modulatory effects of Vitamin C, but, unlike Vitamin C, it does not induce cytotoxic effects of through stimulation of

migration, homing to bone marrow, and bone marrow neo-angiogenesis [100, 101].

roles in the pathogenesis of chronic lymphocytic leukemia (CLL).

represents a potentially useful approach to its treatment [113].

myelodysplastic syndromes, and clonal hematopoiesis [114].

**10. Vitamin C as a powerful modulator of TET2 activity**

and to induce a blastocyst-like state in mouse embryonic stem cells [117].

and myelodysplastic syndrome (MDS) [110–112].

therapy [98].

166 Myeloid Leukemia

The anticancer properties of Vitamin C are known, since at least six decades, even though its use in clinical practice has only recently re-emerged, after the demonstration that in relatively high concentrations, it can selectively kill a number of different human tumor cells, both in vitro and in vivo.

The proof of the anticancer efficacy of Vitamin C in high doses, administered by mouth, has been reported four decades ago, by Linus Pauling [54–57], and further confirmed, very recently, by experimental in vitro and in vivo data [30–32, 66, 69].

Vitamin C is a natural compound, and it is an antioxidant and a life-saving nutrient with multiple beneficial effects on the human body. Man, some primates, and a few other mammals do no longer produce it. Beyond being a natural and essential nutrient, Vitamin C shows, in vitro, an outstanding efficacy in killing a number of different cancer cells, with an efficiency that no other anticancer drug, presently available on the market, has ever shown.

Vitamin C is extremely selective since it kills only cancer cells, by sparing, at the same time, all the other cells of the organism. As a consequence, it is very well tolerated, and devoid of any significant side effects. In fact, the only (relative) contraindication to its use, is the lack of the enzyme glucose-6-phosphate-dehydrogenase (G6PDH), a rare genetic condition also known as "favism." More importantly, within an expensive and often artificially inflated market, such as that of the anticancer drugs [122, 123], Vitamin C, with its low cost, represents an outstanding opportunity for both the patients and the healthcare system.

Unfortunately, in spite of all the above characteristics, Vitamin C has never been easily or favorably accepted as an anticancer drug, by the western Medicine. This also explains why, although the data on its anticancer efficacy are outstanding and straightforward, many scientists still prefer to consider "controversial" the role of Vitamin C in the treatment of cancer.

which decline with differentiation [138]. Importantly, human HSCs and multipotent progeni-

High Doses of Vitamin C and Leukemia: In Vitro Update http://dx.doi.org/10.5772/intechopen.71484 169

Using "GULO" mice (deficient in Vitamin C because of the lack of gulonolactone oxidase, the last enzyme in the synthesis of Vitamin C starting from glucose), Agathocleous and colleagues have shown that Vitamin C deficiency induces an increased number of HSCs. A FLT3 internal tandem duplication (ITD) mutation, found in approximately a quarter of patients with de novo AML, imparts a particularly poor prognosis. Using "GULO" mice (deficient in Vitamin C because of the lack of gulonolactone oxidase), Agathocleous and colleagues have shown that Vitamin C deficiency induces an increased number of HSCs. Therefore Vitamin C deficiency, and TET2 mutations, are likely to cooperates with FLT3-ITD to induce leukemia development in

Given the above evidence, it will be worth mentioning, once more, that the biochemist Irwin Stone, in his book "The healing Factor: Vitamin C against disease," published in 1972 (45 years ago!), had already warned the scientific community on the role of Vitamin C as a main factor in the prevention and treatment of leukemia. In his words, "*In a leukemic, the biochemical stresses of the disease process has reduced the body stores of ascorbic acid to very low levels … Any ascorbic acid circulating in the blood has been scavenged and locked in the excessive numbers of white blood cells contained in the blood. The plasmas level of ascorbic acid is usually zero or close thereto. A zero level in the blood plasma means that the tissues of the body are not being supplied with this most important metabolite. The ascorbic acid contained in the leukocytes are unavailable for the tissues. The tissues are in a condition of biochemical scurvy and this explains why these depleted tissues are so susceptible to the characteristic hemorrhaging of leukemia and the infections that kill so many of the leukaemics. A leukemic is not only suffering from leukemia but also from a bad case of biochemical scurvy. To correct this condition, ascorbic acid has to be administered in sufficiently large doses not only to saturate the excess of white blood cells but to provide adequate spill over into the blood plasma and tissues so that the seriously ill leukemic will be given a fighting chance to combat the disease. This may require the administration of ascorbic acid at the rate of 25 or more grams per day, as noted in the* 

The rationale behind the use of high doses of Vitamin C in the treatment of acute leukemia is

**b.** While it is currently supposed to kill cancer cells by inducing pro-oxidant damage, Vitamin C is also very effective as an antioxidant by inhibiting the hypoxia inducible factor (HIF);

**c.** The mechanistic explanation of the pathogenesis of myeloid leukemia, includes the possibility that a Vitamin deficiency may induce the neoplastic transformation of myeloid precursors, through an upregulation of the HIF, and the consequent cascade of HIF-de-

**a.** Leukemic patients, almost invariably show a severe deficiency of this nutrient;

tor cells (MPPs), such as murine HSCs, display high Vitamin C levels.

*following case of leukemia treated with megascorbic levels of ascorbic acid.*" [3].

murine models of FLT3-ITD-driven leukemia. [138].

**13. Concluding remarks**

pendent cancer genes;

strong and very well grounded. In summary:

As we have seen, the idea that the oral administration of Vitamin C, in high doses, is not effective against cancer is a conceptual artefact, originating from questionable interpretations of pharmacokinetics data, after oral and/or intravenous administration. On the other hand, the idea that Vitamin C, administered in high doses by intravenous infusion, behaves as a pro-drug of H<sup>2</sup> O2 beyond being experimentally questionable, has not led to clinically significant results or outcomes [124–128]. More importantly, encouraging results have emerged from unbiased interpretation of the available data [129]. In particular, as it has been shown up to 110 g/m<sup>2</sup> / day are very well tolerated by the majority of patients, and even in the absence of any significant clinical remission, intravenous Vitamin C is almost invariably associated with a clear-cut improvement in patient's quality of life.

As a result, History repeat itself! … and just as Vitamin C was dismissed as ineffective, against cancer, more than 30 years ago, on the ground of questionable clinical trials [130, 131], nowadays, it runs again the risk of being definitively discarded, in spite of the large amount of scientific evidence, demonstrating its extraordinary efficacy in fighting cancer!

It is clear that much remains to be understood about the cytotoxic effects of Vitamin C against cancer, and much more can (and must!) be done, to both improve the intravenous therapy and further investigate the oral administration route of the high doses of the nutrient.

Improving the intravenous treatment can (and should!) be achieved, by considering:

