Author ORCID Identifier

0000-0002-9791-8904

Date of Graduation

5-2020

Document Type

Dissertation (PhD)

Program Affiliation

Cancer Biology

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Curtis R. Pickering

Committee Member

Jeffrey N. Myers

Committee Member

Michael Andreeff

Committee Member

Andrew G. Sikora

Committee Member

Faye M. Johnson

Abstract

Caspase-8 (CASP8) is one of the most frequently mutated genes in Head and Neck Squamous Carcinomas (HNSCC), and mutations of CASP8 are associated with poor overall survival. The distribution of these mutations in HNSCC suggests that they are likely to be inactivating. Inhibition of CASP8 has been reported to sensitize cancer cells to necroptosis, a unique cell death mechanism. Here, we evaluated how CASP8 regulates necroptosis in HSNCC using cell line models and syngeneic mouse xenografts. In vitro, knockdown of CASP8 rendered HNSCCs susceptible to necroptosis induced by a second mitochondria-derived activator of caspase (SMAC) mimetic, Birinapant, when combined with pan-caspase inhibitors zVAD-FMK or emricasan. Strikingly, inhibition of CASP8 function via knockdown or emricasan treatment was associated with enhanced radiation killing by Birinapant through induction of necroptosis. In a syngeneic mouse model of oral cancer, Birinapant, particularly when combined with radiation delayed tumor growth and enhanced survival under CASP8 loss. Exploration of the molecular underpinnings of necroptosis sensitivity confirmed that the level of functional receptor-interacting serine/threonine-protein kinase-3 (RIP3), a key enzyme in the necroptosis pathway was crucial in determining susceptibility to this mode of death. Although an in vitro screen revealed that many HNSCC cell lines were resistant to necroptosis due to low levels of RIP3, patient tumors maintain RIP3 expression and should therefore remain sensitive. Collectively, these results suggest that targeting the necroptosis pathway with SMAC mimetics, especially in combination with radiation, may be a relevant therapeutic approach in HNSCC with compromised CASP8 status, provided that RIP3 function is maintained.

Keywords

HNSCC, oral cancer, Caspase-8, Necroptosis, Radiation, RIP1, RIP3, MLKL, Birinapant, Z-VAD-FMK, Necrostatin-1s

Available for download on Saturday, May 01, 2021

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