NYMC Student Theses and Dissertations

Date of Award

5-19-2020

Document Type

Doctoral Dissertation - Open Access

Degree Name

Doctor of Philosophy

Department

Microbiology and Immunology

First Advisor

Dr. Raj Tiwari

Second Advisor

Dr. Debabrata Banerjee

Third Advisor

Dr. Jan Geliebter

Abstract

Breast cancer remains the second leading cause of cancer death among women in the U.S. Although organ-confined disease is curable, metastatic disease remains incurable and an unmet clinical need. Hence, the development of new therapeutic targets and further understanding of the genesis of malignant progression is needed and to this end analysis of the tumor microenvironment is imperative for novel therapeutic targets in breast cancer. We and others have demonstrated that bone marrow derived endothelial progenitor cells (EPCs) incorporate in the neovasculature around implanted tumors supporting their growth and metastasis. The incorporation of EPCs from the marrow to the tumor is estrogen responsive. We evaluated the effect of estrogen and a naturally occurring anti-estrogen 3, 3’-diindolylmethane (DIM) on precursor progenitor cells of angiogenesis, EPCs, at the cellular and molecular levels using CD133+ /CD34+ stem cells that were isolated by positive selection from human umbilical cord blood as an in vitro model of vasculogenesis. Moreover, we evaluated the secretome of the EPCs and their role in breast cancer progression. Using flow cytometry, we show that the highly proliferative EPCs can differentiate into functional endothelial cells. We concluded that EPCs are estrogen sensitive, and estradiol mediated enhancement of the function of EPCs is abrogated by the anti-estrogen DIM. We further elucidated the significance and function of EPCs in the breast cancer environment using in vitro models and attempted to define the interacting determinants. We examined the cell-cell interaction by evaluating the effect of conditioned media from EPCs on breast cancer cell and vice versa. We further isolated secretory exosomes from EPCs and breast cancer and examined their mutual interaction using end point measurements such as cell proliferation, invasion, and migration. Further, we xviii examined the contents of the exosomes in term of their microRNAs, and the cytokines in the tumor microenvironment. We report that exosomes of EPCs are enriched with oncogenic miRNAs including hsa-miR-16-5p, hsa-miR-17-5p, hsa-miR-181a-5p, hsa-miR-21-5p, hsa-miR-142-3p, and their miRNAs are also significant modulators of breast cancer transformation and metastasis. Our studies are consistent with the hypothesis that secretory exosomes and paracrine cytokines are shuttles of cell-cell communication and important targets of novel breast cancer therapy. The exosomes and the microRNAs they carry are the communicating determinants between breast cancer cells and EPCs in the tumor microenvironment and presumably help evolution of breast cancer subsets such as the triple negative breast cancer (TNBC) with gain of metastatic phenotype. Disruption of the cell-cell communication can lead to novel TNBC therapeutic and fulfill a much-needed clinical need.

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