**Modulation of One-Carbon Metabolism by B Vitamins: Implications for Transformation and Progression of Prostate Cancer**

Glenn Tisman *Cancer Research Building, Whittier, CA USA* 

#### **1. Introduction**

190 Prostate Cancer – From Bench to Bedside

Zapata, P.D., Ropero, R.M., Valencia, A.M., Buscail, L., López, J.I., Martín-Orozco, R.M.,

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> Extensive laboratory, epidemiological and clinical investigations suggest that prostate cancer might be affected by enhanced folate or B12 ingestion and or other perturbations of one-carbon (CH3—) metabolism. Over the last decade, largely due to government mandated dietary fortification with folic acid (FA), our clinic patients experienced a 4-6-fold increase in the median level of serum folate (5 ng/ml 24 ng/ml). The National Health and Nutrition Examination Surveys (NHANES) confirm similar elevated levels Figure 1 (Dietrich et al, 2005; McDowell et al, 2008; Yang et al, 2010).

Fig. 1. Median serum/plasma blood folate levels NHANES and GT data.

Modulation of One-Carbon Metabolism by B Vitamins:

**neoplasia** 

Kim, 2008; Smith et al, 2008).

Implications for Transformation and Progression of Prostate Cancer 193

The idea that B12 could accelerate chronic myelogenous leukemia in a patient with pernicious anemia and B12 deficiency was first demonstrated by Jose Corcino in 1971 while working in Dr. Victor Herbert's lab in the Bronx, NY (Corcino et al, 1971). In 1994, Dr. Ralph Green confirmed that observation in other patients with pernicious anemia (Green, 1994). Finally, in 2009, Tisman first demonstrated that B12 administration accelerated the growth of the epithelial prostate tumor in a patient with pernicious anemia and untreated prostate

cancer, while at the same time correcting his anemia (Tisman et al, 2009).

cancer to higher risk with decreased survival (D'Amico et al, 2005) Figure 2.

**3. Contemporary observations spur interest in folate, B12 and prostate** 

In 1998 the US government mandated that the US diet be fortified with FA in an attempt to prevent birth defects such as spina bifida and anencephaly. Subsequent to governmentmandated fortification of US, Canadian and Chilean diets with FA, numerous reports appeared documenting a higher incidence of certain cancers (colon, rectum, breast, prostate), reviewed by Young-In Kim and others (Hirsch et al, 2009; Kim, 2007; Kim, 2007;

The newest data relate elevated serum and prostate tissue folate to increased Gleason's grade and proliferation of prostate tumors compared to normal donor prostate tissue (Tomaszewski et al, 2011). FA supplementation was associated with a 2.6 fold increase in incidence (Figueiredo et al, 2009) and stage (Lawson et al, 2007) of prostate tumors. Collin noted serum folate-related increase in PSA velocity (Collin et al, 2010) enough to advance low-risk prostate

Fig. 2. Higher serum/plasma folate, associated with PSA velocity > 2.0 may increase mortality.

Exposure to varying concentrations of FA and other B vitamins may transform benign cells to malignant and/or accelerate tumor growth. This review explores clinical observations and biochemistry of B vitamin mediated one-carbon metabolism with general emphasis on neoplasia and particular focus on prostate cancer.

## **2. Folic acid found to enhance tumor growth: Historical note**

Sidney Farber, a pediatric pathologist practicing in Boston's Children Hospital in the mid 1940s, is generally given credit for establishing that FA stimulates human leukemia cell proliferation; however, a careful review of the literature does not confirm that assessment. In fact, Sidney Farber's clinical research group's seminal paper did not include treatment with FA (Farber et al, 1947). This manuscript, published in 1947 in the popular journal Science, reported on Farber's experience with the di- and triglutamate derivatives of oxidized folic acid (diopterin and teropterin, both supplied by Lederle Laboratories of Pearl River, NY under the direction of Yellapragada Subba Row, PhD). The fact is that Farber's landmark paper, routinely referenced in the literature, never stated FA could accelerate leukemia or any other malignancy.

Dr. Richard Lewisohn, famous for his excellence in surgery and his 1917 research on citrated blood as the preferred anticoagulant for transfusion was semiretired in the early 1940's and working in a trivial basement laboratory at Mount Sinai Hospital in NYC. At that time, he was given two folate compounds that were isolated at the Lederle Laboratories: liver L. casei factor (folic acid) and fermentation L. casei factor. The mislabeling of these compounds was the source of confusion for his initial paper mistakenly stating that folic acid was an inhibitor of spontaneous mouse mammary cancer (Leuchtenberger et al, 1945). The compound he thought was FA was in fact pteroyltriglutamate or teropterin.

Later, Hutchings and Stokstad of Lederle informed Lewisohn's group that the correct tentative designation of liver L. casei factor as used in Lewisohn's first report was actually fermentation L. casei factor (the triglutamate of pteroylglutamic acid or teropterin (Angier et al, 1946).

In Lewisohn's subsequent paper (Lewisohn et al, 1946) he confirmed that the mislabeled folic acid (liver L. casei factor) was actually fermentation L. casei factor or teropterin. Clarifying this confusing issue, the initial study revealed that teropterin injection inhibited spontaneous mouse mammary cancer while folic acid (liver L. casei factor), as confirmed in subsequent experiments, stimulated mouse mammary primary tumor growth and its pulmonary metastases while shortening overall survival (Lewisohn et al, 1946). To our knowledge, this is the first literature-documented study revealing FA stimulation of cancer cells (mouse mammary cancer) and their metastasis.

Next, in 1948, the hematologist Robert Heinle (Heinle & Welch, 1948) and later in 1950 the laboratory researcher Howard Skipper (Skipper et al, 1950) were first to document in the clinic and laboratory respectively that FA stimulated chronic myelogenous leukemia in man and acute leukemia in the rat.

The first mention that Farber was aware of what he termed an "acceleration phenomenon" appeared in his work published in 1948 in the NEJM (Farber et al, 1948). In that paper, he referred back to the Science 1947 manuscript stating that the reported children with leukemia displayed acceleration of the leukemic process within the marrow in response to diopterin or teropterin therapy. However, that finding had never been mentioned in the 1947 landmark Science report.

Exposure to varying concentrations of FA and other B vitamins may transform benign cells to malignant and/or accelerate tumor growth. This review explores clinical observations and biochemistry of B vitamin mediated one-carbon metabolism with general emphasis on

Sidney Farber, a pediatric pathologist practicing in Boston's Children Hospital in the mid 1940s, is generally given credit for establishing that FA stimulates human leukemia cell proliferation; however, a careful review of the literature does not confirm that assessment. In fact, Sidney Farber's clinical research group's seminal paper did not include treatment with FA (Farber et al, 1947). This manuscript, published in 1947 in the popular journal Science, reported on Farber's experience with the di- and triglutamate derivatives of oxidized folic acid (diopterin and teropterin, both supplied by Lederle Laboratories of Pearl River, NY under the direction of Yellapragada Subba Row, PhD). The fact is that Farber's landmark paper, routinely referenced in the literature, never stated FA could accelerate

Dr. Richard Lewisohn, famous for his excellence in surgery and his 1917 research on citrated blood as the preferred anticoagulant for transfusion was semiretired in the early 1940's and working in a trivial basement laboratory at Mount Sinai Hospital in NYC. At that time, he was given two folate compounds that were isolated at the Lederle Laboratories: liver L. casei factor (folic acid) and fermentation L. casei factor. The mislabeling of these compounds was the source of confusion for his initial paper mistakenly stating that folic acid was an inhibitor of spontaneous mouse mammary cancer (Leuchtenberger et al, 1945). The

Later, Hutchings and Stokstad of Lederle informed Lewisohn's group that the correct tentative designation of liver L. casei factor as used in Lewisohn's first report was actually fermentation L. casei factor (the triglutamate of pteroylglutamic acid or teropterin (Angier

In Lewisohn's subsequent paper (Lewisohn et al, 1946) he confirmed that the mislabeled folic acid (liver L. casei factor) was actually fermentation L. casei factor or teropterin. Clarifying this confusing issue, the initial study revealed that teropterin injection inhibited spontaneous mouse mammary cancer while folic acid (liver L. casei factor), as confirmed in subsequent experiments, stimulated mouse mammary primary tumor growth and its pulmonary metastases while shortening overall survival (Lewisohn et al, 1946). To our knowledge, this is the first literature-documented study revealing FA stimulation of cancer

Next, in 1948, the hematologist Robert Heinle (Heinle & Welch, 1948) and later in 1950 the laboratory researcher Howard Skipper (Skipper et al, 1950) were first to document in the clinic and laboratory respectively that FA stimulated chronic myelogenous leukemia in man

The first mention that Farber was aware of what he termed an "acceleration phenomenon" appeared in his work published in 1948 in the NEJM (Farber et al, 1948). In that paper, he referred back to the Science 1947 manuscript stating that the reported children with leukemia displayed acceleration of the leukemic process within the marrow in response to diopterin or teropterin therapy. However, that finding had never been mentioned in the

compound he thought was FA was in fact pteroyltriglutamate or teropterin.

cells (mouse mammary cancer) and their metastasis.

and acute leukemia in the rat.

1947 landmark Science report.

neoplasia and particular focus on prostate cancer.

leukemia or any other malignancy.

et al, 1946).

**2. Folic acid found to enhance tumor growth: Historical note** 

The idea that B12 could accelerate chronic myelogenous leukemia in a patient with pernicious anemia and B12 deficiency was first demonstrated by Jose Corcino in 1971 while working in Dr. Victor Herbert's lab in the Bronx, NY (Corcino et al, 1971). In 1994, Dr. Ralph Green confirmed that observation in other patients with pernicious anemia (Green, 1994). Finally, in 2009, Tisman first demonstrated that B12 administration accelerated the growth of the epithelial prostate tumor in a patient with pernicious anemia and untreated prostate cancer, while at the same time correcting his anemia (Tisman et al, 2009).

## **3. Contemporary observations spur interest in folate, B12 and prostate neoplasia**

In 1998 the US government mandated that the US diet be fortified with FA in an attempt to prevent birth defects such as spina bifida and anencephaly. Subsequent to governmentmandated fortification of US, Canadian and Chilean diets with FA, numerous reports appeared documenting a higher incidence of certain cancers (colon, rectum, breast, prostate), reviewed by Young-In Kim and others (Hirsch et al, 2009; Kim, 2007; Kim, 2007; Kim, 2008; Smith et al, 2008).

The newest data relate elevated serum and prostate tissue folate to increased Gleason's grade and proliferation of prostate tumors compared to normal donor prostate tissue (Tomaszewski et al, 2011). FA supplementation was associated with a 2.6 fold increase in incidence (Figueiredo et al, 2009) and stage (Lawson et al, 2007) of prostate tumors. Collin noted serum folate-related increase in PSA velocity (Collin et al, 2010) enough to advance low-risk prostate cancer to higher risk with decreased survival (D'Amico et al, 2005) Figure 2.

Fig. 2. Higher serum/plasma folate, associated with PSA velocity > 2.0 may increase mortality.

Modulation of One-Carbon Metabolism by B Vitamins:

normal of serum prostate specific antigen Figure 4 (in press JMCR).

Fig. 4. Patient 2, PSA & withdrawal of B12 and folates

**vitamin nutrition** 

**4. Mechanisms of modulation of one-carbon metabolism associated with B** 

generating methionine, later to be metabolized to S-adenosylmethionine (SAM).

Figures 5 and 6 help visualization of the nomenclature and structure of folate vitamers of the "one-carbon" pool. A normal diet supplies methyl groups through methionine and choline, however physiological needs exceed dietary intake. Man makes up the difference by *de novo* synthesis of methyl groups (CH3—). As illustrated in Figure 7, both dietary methionine and choline supply CH3— groups. Note the re-methylation of homocysteine

**3.2 Patient 2** 

Implications for Transformation and Progression of Prostate Cancer 195

A 71 year-old man was diagnosed in 1997 with Stage T1c prostate cancer, Gleason's score = 3+4 = 7. Primary therapy included intermittent androgen deprivation to resistance. While receiving docetaxel chemotherapy for 18 weeks with a continually increasing PSA, withdrawal of ingestion of 10 daily doses of a supplement composed of (500 mcg of vitamin B12 as cyanocobalamin, and 400 mcg each of folic acid as pteroylglutamic acid and 400 mcg of L-5-methyltetrahydrofolate = 800 mcg of mixed folates) was associated with a return to

Others related high B12 levels to prostate cancer (Hultdin et al, 2005; Johansson et al, 2009; Weinstein et al, 2006; Vlajinac et al, 1997). We reported direct stimulation of prostate cancer by administration of B12 to a B12 deficient patient (Tisman et al, 2009) and by a supplement containing a combination of B12 and mixed folates Figures 3 and 4 (Tisman & Garcia, 2011). Patients with prostate cancer frequently ingest a variety of B vitamin-containing supplements (Velicer & Ulrich, 2008; Bailey et al, 2010) including FA and B12. We confirmed this almost universal finding in our clinic. Many are oblivious that their supplements contain larger than needed doses of vitamins. Others take comfort in supplement ingestion immediately after a cancer diagnosis while some use them in an attempt at prophylaxis (Holmes et al, 2010). Holmes' group noted folic acid supplement use before a colorectal cancer diagnosis was 35.4%. This statistic increased to 55.1% after receiving a diagnosis. We start our review by briefly presenting two of our patients with prostate cancer whose clinical course was adversely impacted by the administration of B12 and mixed folates. This is to be followed by a rather in depth review of B vitamin metabolism as relates to
