Role of RNA-Binding Protein RBM3 in Prostate Cancer Progression
Abstract
This application addresses the Early Investigator Research Award in Prostate Cancer Research Program (FY20 PCRP). Prostate cancer (PCa) is a deadly disease with 21% of diagnosed cases in 2020, which also equates to a 1-in-5 diagnosis rate of cancer in men. It also will result in 33,330 deaths in 2020. The biggest problems with PCa is the overtreatment of benign disease and inadequate therapies for metastatic and castrate resistant disease. Therefore, it is important to understand the biology of prostate cancer to find new targets for progressive disease. In this application, we are addressing the critical overarching challenge of defining the biology of lethal prostate cancer (PCa) to reduce death. We will use in vitro and in vivo approaches to understand how RNA binding protein RBM3 induces stemness and the effect of knocking out the gene on PCa growth and development. We and others have shown that RNA binding protein RBM3 is upregulated in PCa as compared to normal tissues. RNA binding protein RBM3 binds to AU-rich sequences located in various mRNAs including cytokines and proto-oncogenes and enhances their translation thereby increasing protein levels. We have also demonstrated that RBM3 increases stemness of cancer cells. Stem cells are the biggest problem because they can propagate, give rise to various cells in the tumor, and interact with the environment to ensure survival. To begin understanding the role of RBM3, we have proposed three independent aims. The first aim is designed is to understand the biology of RBM3 in the PCa cells in culture. We will take three cell lines, human LNCaP, its metastatic variant C4-2B, and mouse PTEN-CaP9 cells and increase or knock out RBM3 expression using two different CRISPR technologies. In preliminary studies, we have observed that RBM3 is significantly higher in C4-2B cells compared to LNCaP cells. Once we have confirmed that the protein expression has changed, we will determine what this does to the cells in terms of growth and ability to invade matrigel (a surrogate for invasion in tissue). Also, we will determine whether there is changes to stemness and expression of known stem cell markers CD133, CD44, ALDH1, and CD117. In addition, we have discovered a novel marker DCLK1. We will determine whether RBM3 increases gene transcription of the stem cell markers and also enhances translation of these proteins. We will also determine additional gene expression changes by performing a technique called Ribo-seq. For the second aim, we have generated a mouse where we have floxed RBM3 gene. Now, we can cross the mice with a mouse that expresses the Cre recombinase in prostate epithelial cells, macrophages and T cells. We will use specific mice for each of these and knockout RBM3 in each compartment. With the epithelial knockout, we will also knock out PTEN. These mice get spontaneous tumors, but we believe that loss of RBM3 will reduce tumor incidence and growth. With macrophage and T cell knockout, we will induce orthotopic tumors by injecting PTEN-CaP9 cells. Tumor-associated macrophages enhance tumor growth and metastasis and induce drug resistance. Activated T cells also suppress immune responses to the tumors. We predict that loss of RBM3 in these compartments will also stunt tumor growth. Finally, in the third aim, we will obtain tissues from the PCBN network and correlate the expression of RBM3 and the various stem cell markers with clinical and pathological features. This is a basic science research wherein we are proposing to understand the role of RBM3 in mediating stemness in cancer cells as well as macrophages and T cells in the tumor microenvironment. Stemness is a significant problem with cancers, as they can survive harsh conditions and reemerge as a new tumor. Hence identifying and eliminating these cells is a top priority. While the near-term goals are to understand the biology of the disease, the long-term implications of underst
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Dec 05, 2021
- Source ID
- W81XWH2110157
Entities
People
- Afreen Sayed
Organizations
- United States Army
- University of Kansas Medical Center