Genetic, Dietary and Environmental Influences on Vitamin D Metabolism
Condition: Vitamin D Deficiency
MeSH terms: Vitamin D Deficiency
Study type: Interventional

Summary: 
      Determine differences in serum vitamin D metabolism by genetic ancestry.
    

Detailed description: 
      Vitamin D metabolites are well-recognized to stop cancer cell growth in culture. However, a
      clear definition of a sufficient level of serum vitamin D (currently measured as 25(OH)D) for
      disease prevention has been hampered by inconsistent results from both observational studies
      and randomized clinical trials. Observational cancer studies report both increased and
      reduced risks of cancer in subjects with higher serum levels of 25(OH)D, while large
      randomized trials report no significant benefit of vitamin D supplementation for breast
      cancer (incidence), colon cancer (incidence and mortality), lung cancer (mortality), or
      benign proliferative breast disease.

      The 25(OH)D metabolite of vitamin D (comprised of 25(OH)D3 and 25(OH)D2), is the principal
      hydroxylated metabolite in serum and is considered a reasonable functional biomarker of
      vitamin D status. This is because, although subject to seasonal variation, multiple
      measurements in the same individual are relatively consistent over time. The most
      biologically active vitamin D metabolite is 1,25(OH)2D3, and may therefore be the most
      relevant to long-term health outcomes. Nonetheless, 1,25(OH)2D3 is not frequently used as a
      biomarker in epidemiologic studies as it displays diurnal variation and has a short in vivo
      half life.

      Notably, in vivo treatment with 1,25(OH)2D3 is associated with increased metabolic clearance
      of 25(OH)D3, decreased serum 25(OH)D3 levels, and a higher 25(OH)D3 to 24,25(OH)2D3
      conversion rate in humans and animals. This suggests that use of 25(OH)D as single biomarker
      of vitamin D status does not fully capture the entire picture. Paradoxically, one individual
      may be classified with 'low' 25(OH)D as a consequence of low dietary intake and little sun
      exposure while yet another individual classified with 'low' 25(OH)D may actually have a
      higher concentration of the 1,25(OH)2D3 metabolite-possibly due to genetic differences in
      enzymes that metabolize vitamin D (CYP2R1, CYP27B1, and CYP24A1).

      Genome-wide and candidate gene association studies of serum 25(OH)D levels have suggested a
      role of single nucleotide polymorphisms (SNPs) near the vitamin D binding protein (GC) and
      CYP2R1. However, the investigators and others have shown that serum 25(OH)D levels are also
      associated with genetic ancestry. Because existing studies do not adjust for genetic ancestry
      and the function of these SNPs have yet to be established, it is not known whether these SNPs
      actually play a role or whether these may be spurious associations due to ancestral
      background (i.e., population stratification), correlations with truly causal SNPs, or to
      random chance. In addition, because of the number of SNPs that must be distinguished,
      genome-wide studies do not capture regions with high homology or sequence repeats, such as
      the promoter region of CYP24A1 which has a higher guanine-cytosine content. Improved
      classification of 'low' 25(OH)D levels within the context of a particular genetic background
      through identification of "rapid" versus "slow" vitamin D metabolizers will likely have
      important implications for cancer risk.

      It is well-recognized that individuals with African Ancestry have substantially lower
      (~2-fold) serum 25(OH)D levels compared with other racial/ethnic groups. These differences
      have been attributed primarily to skin pigmentation. However, the relation between serum
      25(OH)D levels and health outcomes is complex and involves a number of variables including
      diet, sun exposure and hormone status. For example, despite lower average dietary intake of
      both calcium and vitamin D, and lower serum 25(OH)D levels, African Americans have higher
      bone mineral density and a 3-fold lower risk of hip fracture relative to European Americans.
      There are some lines of evidence which suggest that African Americans may have comparatively
      higher circulating levels of 1,25(OH)2D3, although most studies are small and do not account
      for age and/or diurnal variation, and this may explain why other studies report no
      difference.

      Among Europeans alone, a moderately high heritability of serum 25(OH)D levels (~50 to 70% for
      25(OH)D) has been observed, as well as associations between 25(OH)D and several genetic
      polymorphisms in vitamin D metabolizing enzymes, although the functional consequences of
      these polymorphisms are not known. Small and large clinical trials among healthy individuals
      have also noted substantial inter-individual differences(as much as 5-fold) in the increases
      of serum 25(OH)D levels among participants given the same oral dose of vitamin D, the same
      minimal erythema dose (MED) of UV-B light, or the same dose of UV light. The importance of
      understanding inter-individual differences in vitamin D synthesis, metabolism and effect have
      been underscored by results from high dose randomized trials reporting that those in the
      vitamin D supplementation groups experienced a 31% higher bone fracture incidence, and
      significantly lower prostate cancer survival. There is a clear need to understand individual
      differences in the metabolism of vitamin D before future high dose trials proceed.

      Previous Data: Studies by the investigators' lab and others have identified significant
      racial/ethnic differences in the frequency of variants in genes responsible for vitamin D
      metabolism (i.e., CYP2R1, CYP27B1 and CYP24A1). In collaboration with the Pike Laboratory
      (University of Wisconsin), the investigators' laboratory has identified several novel genetic
      variants in recently identified regulatory regions of the gene which encodes for the major
      vitamin D catabolic enzyme (CYP24A1).

      Study Rationale:The role/function of these polymorphisms has not been tested in either
      epidemiologic association studies or in cell-based assays. It will be important to complement
      genetic association studies with multiple types of molecular assays that can tell a
      consistent story about the likely role of a particular genetic variant.

      The investigators hypothesize that there is a significant influence of genetic variants on
      serum vitamin D metabolism that is independent of genetic ancestry, skin melanin content,
      sunlight exposure and dietary/supplement intake.

      Study Objectives:A small randomized study in healthy subjects will investigate temporal
      changes in the balance of metabolites (25(OH)D3, 25(OH)D2, 24,25(OH)2D3 and 1,25(OH)2D3)
      following a two-month course of vitamin D3 (800 IU/d).

      This study overcomes limitations of existing genetic association studies that may report
      spurious results due to lack of control for genetic ancestry, skin reflectance, collection of
      blood specimens during summer months, or the use of assays that do not distinguish 25(OH)D2
      and 25(OH)D3. The advantages of this study are: 1) a matched design that incorporates
      temporal assessment of vitamin D metabolism, 2) genetic Ancestry Informative Markers (AIMs),
      3) melanin skin index, 4) a sensitive and reliable assay for four serum vitamin D
      metabolites, and 5) functional molecular assays related to specific genetic polymorphisms.

      The investigators' research addresses one of the major research gaps in the understanding of
      the health-related benefits of vitamin D identified by a recent Institute of Medicine Review.
      While vitamin D metabolites have significant anti-cancer properties, a better understanding
      of the genetic influences upon vitamin D metabolism is needed in order to improve the
      identification of cancer risks associated with vitamin D.

      Study Design: This study employs two designs-first a cross-sectional study of participants is
      selected.

      Among the cross-sectional study participants, a subset is randomly selected and matched based
      on genetic ancestry, and randomized to either the intervention (800 IU/day of vitamin d) or
      placebo group.

      Statistical Plan:In the first portion of this study, an estimated 400 participants will be
      screened to determine eligibility. Eligibility screening will be completed online through
      RedCap.

      In the second portion of this study, participants randomly chosen from the original 400 for
      the Supplement Intervention Study will be asked to complete 3 additional blood draws at Week
      0, 4 and 8 of the Supplement Intervention Study (N=64).

      The 64 Supplement Intervention study participants will be randomly chosen from the eligible
      participants in the first part of the study (n=400). Sample size calculations were performed
      by Dr. Vernon Chinchilli in order to estimate the number of participants that the
      investigators would need in order to detect differences in their vitamin D levels. The
      investigators accounted for loss-to-follow-up and normal withdrawal from the study.

      NOTE:

      The total participants that will get screened has been changed (from the original AICR grant)
      and revised in the budget from 300 to 400 participants. The investigators made this revision
      because the investigators decided that loss to follow-up may be higher than the originally
      estimated 10%. The investigators then estimated that loss to follow-up may actually be closer
      to 20%. This proportional difference was calculated from the investigators' original estimate
      of 52 to 64 (which is about 24% increase in participants) and was revised also to increase
      the amount of participants that the investigators will screen from the original 300 to an
      estimated 372 (determined proportionally with a 24% increase). The investigators went ahead
      and assumed that the upper limit may include 400 participants so that the investigators can
      end up with 64 participants in the supplementation trial.

      The investigators will conduct an intention-to-treat analysis to analyze differences in the
      serum level of vitamin D metabolites and vitamin D metabolite ratios within supplement and
      placebo group pairs at times 0, 1 month and 2 months. Differences in the pairwise log
      concentration of metabolites and log metabolite ratios will be tested by Wilcoxon Signed-Rank
      Tests and linear mixed-effect models (SAS Proc Mixed) using restricted maximum likelihood
      estimation to account for one time point as well as repeated measures of vitamin D
      metabolites. Potential confounding and effect modification by dietary intake of vitamin D
      will be assessed.
    

Intervention name(s): 800 IU Vitamin D; Placebo
Intervention type(s): Dietary Supplement; Dietary Supplement
Intervention description(s): Among the cross-sectional study participants, a subset is randomly selected and matched based on genetic ancestry, and randomized to the intervention group.; Among the cross-sectional study participants, a subset is randomly selected and matched based on genetic ancestry, and randomized to the placebo group
Other intervention name(s): Vital Nutrients; Vital Nutrients Placebo
Intervention MeSH terms: Vitamin D
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Triple (Participant, Care Provider, Outcomes Assessor)
Primary purpose: Basic Science