Graduation Date

Spring 5-5-2018

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Cancer Research

First Advisor

Michael R. Green

Second Advisor

Javeed Iqbal

Abstract

Diffuse Large B-cell Lymphoma (DLBCL) is the most common subtype of lymphoma. Despite a cure rate of 40% with standard R-CHOP therapy, patients that refract or relapse are subject to a dismal prognosis. Cases of DLBCL can be classified by their molecular expression phenotype, with the GCB-like subtype aligning with the profile of a germinal center B-cell and the ABC-like subtype aligning to that of an activated B-cell. Aggressive disease is often characterized by high levels of B-cell Receptor (BCR) signaling. This pathway engages downstream kinases responsible for stimulating proliferation and survival that play a key role under the normal circumstances of B-cell development. A comprehensive study aimed at delineating sources of inhibitor insensitivity within the BCR signaling pathway was conducted in order to identify novel drivers of disease and improve clinical outcome. A cohort of 39 aggressive lymphoma cell lines was assayed for sensitivity to Ibrutinib, a BTK inhibitor, and Umbralisib, a PI3Kδ inhibitor. Combined with intracellular phosphoflow measurements, these results revealed that higher levels of proximal BCR kinase (SYK, LYN, BTK, BLNK) and AKT (downstream of PI3K) signaling were highly linked and predictive of inhibitor insensitivity. Simultaneous inhibition of these pathways with Ibrutinib and Umbralisib consequently revealed a synergistic relationship. Following these results, a DNA copy number analysis of 673 DLBCL patient profiles was performed alongside 249 matching gene expression profiles to uncover the genomic drivers responsible for higher signaling. These results identified an enrichment of genes with transcription factor activity within regions of significant DNA copy number gain and matching transcript gain. The TCF4 transcription factor was identified within the most significant gain peak at chromosome 18q21 and led to increased transcript and protein translation. TCF4 gain was associated with the aggressive ABC-like phenotype, poor survival, and increased transcription of key BCR signaling component targets, such as BLNK, BTK, PIK3CA (PI3Kα), and the IgM heavy chain constant region. Collectively, these results identified sources of inhibitor insensitivity within DLBCL, and TCF4 was characterized as a driving force behind aggressive BCR signaling.

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