We will determine molecular pathways that allow expression of estrogen receptors (ER) in ER negative cells by performing a whole genome analysis on a rare patient with ER negative breast tissue.
General overview: Patients with breast cancer have tumors that eithercontain estrogen receptor alpha (ERα) or do not contain this receptor. Inindividuals with estrogen receptors, hormone therapy with either anti-estrogensor aromatase inhibitors are effective. It would be beneficial to be able toinduce estrogen receptors in patients with estrogen receptor negative breastcancer via some means. The understanding of the enhancers and promoters whichstimulate increased production of estrogen receptor in breast tissue are poorlyunderstood. A prismatic case of a patient with juvenile gigantomastia hasenabled us to postulate the possibility of finding a key enhancer mutationwhich stimulates the estrogen receptor content in breast. If we can characterizethis, we have the potential target for a therapy for breast cancer patients. Juvenile gigantomastia is a condition in whicha pre-or early pubertal girl develops massive breast tissue. In our patient andanother patient reported in the literature, the percentage of breast ductalcells with estrogen receptor has been very high in the range of 80 to 90%. Innormal women this percentage is between 1 to 2%. We plan to perform whole genomesequencing and subsequent analyses on the patient and her parents as well as ona previously reported juvenile gigantomastia patient with 90% estrogen receptorpositivity. We will then determine the functionality of mutations found using CRISPRtechnology and in vitro studies. IRB approval for this has been submitted andis under consideration.
Statement of work: We will perform whole genome sequencing and analyseson the gigantomastia patient, her mother and father, and on the patient’sbreast tissue. Mutations in the breast tissue which are not found in the mother,father or the patient’s blood will be considered candidates for study. Furtherselection of candidate genes will be those which could putatively alterenhancers or promoters of the estrogen receptor or epigenetic methylation orde-methylation of these same enhancers. We anticipate performing CRISPR/Caspase9 analysis on these candidate genes to determine which will increase estrogenreceptor expression in breast tissue. Forthe latter studies, we will use MCF 10 ER negative breast cancer cells whichare grown in vitro. We will then utilize cDNA array analyses to determine thedownstream signaling molecules which contribute to the mediation of the stimulationof estrogen receptor in breast tissue
Project and criteria for funding: This patient has been evaluatedwith clinical information over the years but we have not undertaken extensivegenetic testing to determine the precise cause of the juvenile gigantomastia. Prismaticcases such as the one presented in this proposal allow for a unique opportunityto link genotypes to phenotypes in humans and evaluate.
Desired outcomes of the project: Determine the molecular pathways thatregulate the expression of estrogen receptor in normal breast tissue. Once wehave identified key pathways, we will utilize the MCF 10 cells to demonstratethat we can turn on estrogen receptor in the cells. Ultimately the goal will be to developtargets that could be used in patients with breast cancer who have estrogenreceptor negative tumors. In those patients our goal would be to enhance theexpression of the estrogen receptor and accordingly the potentialresponsiveness of the tissue to anti-estrogens or aromatase inhibitors. Thisintensive genetic study of a prismatic case may be a very efficient way toidentify genes and pathways which regulate the estrogen receptor
We plan to utilize the funding for support of a graduate student in the research phase of their program or a postdoctoral fellow. This student will become familiar with whole genome sequencing and RNA-Seq, specifically examining pathways responsible for estrogen receptor expression and degradation. The student will be directly supervised by Dr. Felder(PI) and assisted by Dr. Wei Yue since collectively they have over 60 years of direct experience with discovery of novel cellular receptor pathways as well as cancer biology at the cellular and molecular level.