Vitamin D and Breast Cancer in Obesity: Therapeutic Trials

Institution: Stanford University
Investigator(s): David  Feldman , M.D. -
Award Cycle: 2011 (Cycle 17) Grant #: 17OB-0071 Award: $1,166,615
Award Type: Translational Research Award
Research Priorities
Innovative Treatments>Gene therapy and other treatments: new frontiers



Initial Award Abstract (2011)

For women, being overweight or obese (we use the term obese for the combined group) is a well-known risk factor for developing breast cancer (BCa) and for having a worse prognosis. The BCa-associated hazards are: (1) inability to respond to insulin normally (insulin resistance) causing elevated levels of insulin and growth factors, (2) increased estrogen formation by the breast fat, and (3) influx of inflammatory cells into the breast that further stimulate estrogen formation and growth. Obese women are frequently vitamin D deficient. Vitamin D exerts many anti-cancer actions to reduce cancer growth but is especially useful to treat BCa because it suppresses the obesity-induced hazards.

First, we will study the effect of vitamin D in mouse models of human breast cancer. In previous work we have shown that these tumors respond to estrogens and that vitamin D suppresses this activity. For the present studies we will use an innovative mouse model of estrogen receptor positive (ER+) BCa induced by stem cells injected into the mouse mammary fat pads. The cells will contain a gene allowing quantitative, continuous assessment of tumor growth. We will monitor tumor growth and possible metastases using state of the art bioluminescent imaging. The study will compare normal and obese mice generated by a dietary-induced obesity regimen and assess the chemo-preventive activity of vitamin D to delay or prevent the development or progression of BCa.

In parallel work we will validate the beneficial actions of vitamin D by assaying biomarkers in women with newly diagnosed BCa. We will assess the womenís blood and the profile of activated genes in BCa tissue specimens. For these studies, we will recruit 60 BCa volunteers, half of whom will be obese, 40 will receive high dose (10,000 IU) vitamin D and 20 will receive a minimal dose (400 IU) as a control group. The vitamin D is given orally in the interval between the diagnostic biopsy and the surgery several weeks later. We use advanced molecular methods to study blood biomarkers and tissue genes that predict disease prognosis. We will compare the ability of vitamin D to improve the hazards of insulin resistance, inflammation and increased estrogen formation and to change the biomarkers and genes from a pattern that predicts poor prognosis to one that indicates an improved outcome.

Taken together, our findings will indicate to physicians and the BCa community the importance of avoiding vitamin D deficiency and the need to screen for and treat deficiency when it is found. We expect that the benefit of high dose vitamin D will be more dramatic in obese women but that there will be a benefit to all subjects including African Americans who are especially at increased risk for vitamin D deficiency and BCa. We envision that vitamin D will be routinely added to other therapies to enhance their activity by inhibiting estrogen formation in the breast and protecting bones from osteoporosis. These findings will provide the basis for a large clinical trial that will show compelling evidence for the value of avoiding vitamin D deficiency as well as the benefit of vitamin D therapy in women with BCa. The Breast Cancer Connections (BCC), advocacy group in Palo Alto, will advise us and assist in recruitment, development of educational materials; especially in the outreach efforts to underserved and minority women.




Progress Report 1 (2012)

This CBCRP grant has two Aims. Aim I proposes to study the benefit of dietary vitamin D (VitD) compared to calcitriol in mice assessing the anticancer effects of VitD in lean compared to obese mice. Aim II is a randomized clinical trial (RCT) in women, both lean and obese, with newly diagnosed BCa. They will be treated during the neo-adjuvant period with daily VitD at low (400 IU) or high (10,000 IU) doses in the interval from biopsy to surgery. The women will be stratified by BMI to evaluate the effects of obesity. We will assess biomarkers in the womenís blood as well as changes in gene expression profiles in the BCa tissue comparing their biopsy to their surgical specimens. We hope to demonstrate that dietary VitD supplementation is beneficial in both lean and obese women as reflected by changes in blood and tissue biomarkers that would predict a better outcome.

Our findings in Aim I showed that we could induce substantial weight gain (~ doubling) in C57BL6 mice by feeding them a high-fat (HF) diet. In our experiments, obesity accelerated the appearance and growth of MMTV-Wnt-1 tumors established in the mammary fat pads of HF diet fed mice. Control experimental groups included mice on a calorie-restricted (CR) diet, which provided fewer fat-derived calories. The effects of dietary VitD supplementation and calcitriol administration were tested in mice fed the HF diet as well as the CR diet. Weight gain in mice on the obese mice was associated with an acceleration of tumor appearance and continued tumor growth. Dietary VitD and calcitriol delayed the appearance of tumors in both lean mice (on CR diet) and obese mice (on HF diet). Furthermore, not just the first appearance but the continued rate of tumor growth was also slowed by both VitD and calcitriol therapy, resulting in decreases in tumor size at the end of the experiments. Serum and tumor tissue were analyzed. DIO was associated with significant elevations in serum leptin levels as expected in obese mice. However, there was also increased leptin expression locally in the tumors and mammary fat. Treatment with calcitriol and VitD inhibited local leptin expression in the tumors and mammary fat and also resulted in decreased circulating serum leptin levels. DIO did not cause significant changes in tumor or mammary fat adiponectin expression or serum levels. Calcitriol and VitD treatments did favor a modest trend toward increasing adiponectin expression. The measurement of estrogens and inflammatory biomarkers in the sera obtained from these animals is currently under way as are the effects of VitD deficiency.

In Aim II, we are conducting a clinical trial in women with newly diagnosed BCa treated with low dose or high dose VitD. We will examine the issue of obesity and BCa, since about half of our subjects have elevated BMI. At this time 21 women newly diagnosed BCa have been enrolled with 17 evaluable. This slow rate of recruitment remains a problem. The reasons for this problem and our proposed remedies are fully discussed in the section on Human Subjects. Most of the subjects have ER-positive BCa and one-half of them are overweight or obese. As recommended by our statistician, blood tests and tissue arrays will be done in a batched manner after accumulating a sufficient number of samples. Our plans include gene expression arrays and measurement of serum metabolic and inflammatory biomarkers to determine the mechanisms by which VitD and calcitriol benefit both lean and especially obese mice and women with BCa.




Progress Report 3 (2014)

Aim I proposes to study the benefit of dietary vitamin D (VitD) compared to calcitriol (Cal) in ovariectomized mice assessing the inhibitory effects of VitD on breast cancer (BCa) in lean compared to obese mice. Our findings showed that we could induce substantial weight gain (~ doubling) in C57BL6 mice by feeding them a high-fat (HF) diet compared to mice fed a calorie-restricted (CR) diet. Obesity accelerated the appearance and growth of MMTV-Wnt1 BCa established in the mammary fat pads of HF diet fed mice. Dietary VitD and Cal delayed the appearance of BCa in both lean mice and obese mice as well as inhibited the rate of tumor growth resulting in decreased tumor sizes at the end of the experiments. Obesity was associated with significant elevations in serum leptin and insulin levels indicating the development of insulin resistance. In mice treated with VitD or Cal, serum leptin and insulin were reduced, suggesting improvements in insulin sensitivity. Expression of aromatase, estrogen receptor (ER), COX-2 and leptin locally in the tumors and mammary fat were also elevated in the obese mice. The treatments decreased the expression of these genes in the tumors and mammary fat demonstrating their suppressive effects on estrogen synthesis and signaling and inflammation.

Aim II is a randomized clinical trial (RCT) in women with newly diagnosed BCa. The women are treated during the neo-adjuvant period with daily VitD at low (400 IU/placebo) or high (10,000 IU) doses in the interval between biopsy and surgery. The women will be stratified by BMI to evaluate the effects of obesity. We will assess changes in serum biomarkers as well as gene expression profiles in the BCa tissue, comparing the biopsies to the surgical specimens. We hope to demonstrate that dietary VitD is beneficial in both lean and obese women as reflected by changes in blood and tissue biomarkers that would predict a better disease outcome. At this time 35 women have been enrolled with 30 evaluable. Most have ER-positive BCa and ~one-half of them are overweight or obese. As recommended by our statistician, blood tests and tissue arrays have been batched and are now being analyzed. Our plans include gene expression arrays and measurement of serum metabolic and inflammatory biomarkers to determine the mechanisms by which VitD may benefit both lean and especially obese women with BCa. Partial and preliminary analyses measuring the association of important serum biomarkers with serum concentration of 25-hydroxyvitamin D (25D) reveal the following: (1) BMI is negatively associated with 25D levels; (2) HOMA-IR, a measure of insulin resistance, is strongly reduced by Vit D; (3) the inflammatory cytokine IL-1a is negatively associated with increasing 25D while IL-6 and IL-8 show similar trends; (4) VEGF, a marker of angiogenesis shows a negative association with increasing 25D. Gene arrays have been done in only 2 subjects so far. The analysis indicates that 116 genes show a >8-fold change in expression when pre- and post-intervention samples are compared with approximately equal numbers of genes being stimulated as suppressed. These preliminary data demonstrate that dietary VitD treatment of BCa patients for only a short interval of time can modify the expression of many genes and biomarkers some of which are critical to BCa proliferation, inflammation, insulin resistance and angiogenesis.

The requested No Cost Time Extension will allow more time to complete the mouse studies and also to increase recruitment for the human trial and complete the analysis of the serum and gene expression biomarkers. Preliminary data seem to demonstrate a beneficial effect of VitD leading to gene expression and serum biomarker profiles that would predict an improved outcome in human BCa.



Equivalent anticancer activities of dietary vitamin D and calcitriol in an animal model of breast cancer: Importance of mammary CYP27B1 for treatment and prevention.
Periodical:Journal of Steroid Biochemistry and Molecular Biology
Index Medicus: J Steroid Biochem Mol Biol
Authors: Krishnan AV, Swami S, Feldman D.
Yr: 2013 Vol: 136 Nbr: Abs: Pg:289-95

Equivalent anti-cancer activities of dietary vitamin D and calcitriol in an animal model of breast cancer: Importance of mammary CYP27B1 for treatment and prevention.
Periodical:Journal of Steroid Biochemistry and Molecular Biology
Index Medicus: J Steroid Biochem Mol Biol
Authors: A. Krishnan, S Swami and D. Feldman
Yr: 2013 Vol: 136 Nbr: Abs: Pg:289-95

The role of vitamin D in reducing cancer risk and progression.
Periodical:Nature Reviews. Cancer
Index Medicus: Nat Rev Cancer
Authors: Feldman, D., Krishnan AV, Swami S, Giovannucci E, Feldman BJ
Yr: 2014 Vol: 14 Nbr: 5 Abs: Pg:342-57