Date of Graduation

5-2015

Document Type

Thesis (MS)

Program Affiliation

Experimental Therapeutics

Degree Name

Masters of Science (MS)

Advisor/Committee Chair

VARSHA GANDHI

Committee Member

HESHAM AMIN

Committee Member

KUMUDHA BALAKRISHNAN

Committee Member

JAN BURGER

Committee Member

GEORGE CALIN

Abstract

Class I phosphatidylinositol 3-kinase isoforms (α, β, δ, and γ) play a major role in cancer cell growth and survival. PI3K α and β are most studied. PI3K pathway is highly dysregulated in many cancers and aberrant PI3K signaling is associated with oncogene mutations and disease progression in solid tumors and in hematologic malignancies.

Chronic lymphocytic leukemia (CLL) is driven by B-cell receptor (BCR) signaling that promotes B-cell proliferation and survival. PI3K is a critical node in BCR pathway and PI3Kδ has a pivotal role in B-cell development and maintenance and this isoform is over-expressed in many B-cell malignancies, including CLL.

Idelalisib is a FDA approved small molecule PI3Kδ inhibitor. Idelalisib promotes apoptosis in CLL by disrupting molecular pathways related to BCR signaling, leukemic migration and signals from the microenvironment. Importantly, idelalisib inhibits BCR-derived PI3K signaling, dampening survival signals. We hypothesized that similar to inhibition of α and β isoforms, attenuation of PI3Kδ will repress transcription, reduce short-lived anti-apoptotic proteins, induce DNA damage and repair responses, leading to enhanced apoptosis of malignant CLL cells.

Idelalisib treatment induced moderate levels of apoptosis in CLL lymphocytes. Idelalisib treatment with IgM stimulation decreased phosphorylation of AKT, a downstream signaling molecule. We observed a significant decrease in global RNA synthesis and a decline in Mcl-1 transcript and protein levels, with no changes to Bcl-2 protein and mRNA expression. Interestingly, we observed that idelalisib induced γH2AX, a hallmark for DNA damage and repair response.

Bendamustine is a FDA approved alkylating agent for CLL therapy. We hypothesized that idelalisib-mediated decline in Mcl-1and bendamustine-induced DNA damage will sensitize B-CLL cells and this mechanism-based combination will lead to a synergistic interaction.

At clinically relevant concentrations, bendamustine and idelalisib as single agents induced moderate level of apoptosis; however, combination treatment resulted in enhanced CLL cell death. Combination index assessment demonstrated that idelalisib and bendamustine couplet resulted in synergistic cytotoxicity. Mechanistic investigations suggest that the synergy maybe due to modulation of Mcl-1 protein levels and DNA damage and repair responses in CLL. Collectively, the emerging role of PI3K inhibitor in combination with bendamustine provides a unique modality for CLL therapy.

Keywords

PI3K inhibitors, Bendamustine, Chronic Lymphocytic Leukemia, combination treatment, isoform-specific inhibition, apoptotic proteins

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