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Grant Abstract: Trace element supplementation to enhance calcium-mediated suppression of human colon adenoma growth

Grant Number: 5R21CA201782-02
PI Name: Varani
Project Title: Trace element supplementation to enhance calcium-mediated suppression of human colon adenoma growth

Abstract: For the parent studies, we are utilizing human colon polyp tissue maintained in enteroid culture. We have developed a culture system to grow the premalignant human colon tissue using a minimally supplemented culture medium containing only epidermal growth factor and pituitary extract as growth promoters. Adenoma tissue in enteroid culture from 23 different cultures (representing 21 subjects) is now “banked away” as frozen stocks. DNA sequencing and bioinformatic data identifying the presence of mutations thought to influence colon polyp growth are available for each established culture. Phenotypic information (immuno-histochemical marker data) is also available for each of the premalignant tumors. In the parent grant studies, enteroid cultures are maintained under control conditions or exposed to increasing concentrations of calcium – either alone or in conjunction with additional trace elements as part of a natural product supplement (Aquamin®). The effects of each intervention is being evaluated over time by assessing effects of treatment on growth and differentiation. RNA sequencing and proteomic display differences are also being used to identify global changes induced by each intervention. The use of colon enteroid cultures provides a way to determine directly if human colon polyp growth can be more effectively modulated by calcium in conjunction with additional trace elements than by calcium alone. By using cultures established from tumors of several different subjects, we are in position to determine the variability in responsiveness to each intervention as a first step toward understanding the basis for variation in response among individuals.

When the parent grant was funded, the focus was on premalignant (adenoma) tissue. With support from the Administrative Supplement, we will carry out parallel studies with normal colonic mucosa. A number of advancements have made the proposed studies possible. First, we have developed culture conditions for the establishment and expansion of normal colonic mucosa in enteroid culture. Now that normal colonic tissue can be maintained, we have the capacity for a direct comparison of effects of the two interventions on normal colonic tissue responses versus those occurring in adenoma. Second, the clinical portion of our overall studies is now “open for recruitment” and subjects are being enrolled. Biopsies for enteroid culture will come from 12 subjects enrolled in the biomarker study. The biopsies will be from those taken before starting on treatment. Although the study is “blinded,” subjects are enrolled in cohorts; thus, the 12 subjects from whom biopsies will be taken for enteroid culture will consist of 4 subjects each who will subsequently receive calcium alone, Aquamin® or placebo as treatment during the 90-day intervention. Once established in culture, the normal mucosa enteroids will be treated with a range of concentrations of calcium or Aquamin® (exactly as being done with adenoma enteroids) and analyzed in exactly the same manner.

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