In vitro and quantitative phosphoproteomic modeling of castration resistant prostate cancer.
cancer is a highly prevalent disease in elderly men, which is treatable at early stages,
yet causes morbidity and death upon progression to a hormone resistant, metastatic
stage. Given prostate cancer's propensity to metastasize to the bone marrow and escape
from androgen deprivation, the role of the bone marrow microenvironment in enabling
castration resistant growth was ... read moreexamined using human bone marrow derived mesenchymal
stem cells (hMSCs). The effect of bone marrow derived extracellular matrix (BM-ECM)
secreted by hMSCs was examined on LNCaPs and MDa-PCa-2bs, and found to cause cell
survival in both cell lines in androgen depleted conditions as well as chemoresistance
in LNCaPs. Phosphoproteomic analysis identified p-ERK as increased in LNCaP cells grown
on BM-ECM. The use of the specific MEK inhibitor U0126 was able to reverse the survival
advantage LNCaPs grown on BM-ECM in androgen depleted conditions. Finally, human primary
prostate tumors and bone metastases were examined for p-ERK levels and significantly
more p-ERK in bone metastases than primary tumors was found. In the second part of this
work, it was hypothesized that quantitatively measuring the activation of cell signaling
pathways in response to ligand treatments would enable understanding how cell signaling
causes castration resistant growth. Three cell lines (PC3, LNCaP, and MDA-PCa-2b) were
treated with 6 treatments (EGF, IGF1, IL6, TNFa, dihydrotestosterone, and docetaxel) and
the alterations in 8 key phosphoproteins were measured across three time points. LNCaPs
were also treated in combination with the targeted kinase inhibitors against MEK, PI3K,
mTOR, IKK, and p38 resulting in 3400 phosphoprotein measurements. For each treatment the
corresponding cell survival in androgen depleted conditions was assessed. A regression
model correlating phosphoprotein measurements to survival data was able to account for
the majority of the variation in cell survival. The effect of androgen on the
phosphoproteome was also observed and seen to increase PI3K related proteins, and the
innate signaling differences between the cell lines were observed. This regression
analysis was also employed to successfully describe the polarization of macrophages to
the M2 (tumor associated macrophage) versus M1 phenotype based on cell signaling
alterations in response to ligand treatment.
Thesis (Ph.D.)--Tufts University, 2013.
Submitted to the Dept. of Biomedical Engineering.
Advisor: David Kaplan.
Committee: Catherine Kuo, Paul Mathew, and Judith Clements.
Keywords: Biomedical engineering, Cellular biology, and Molecular biology.read less