Title

Cytotoxicity, Synergism and Apoptosis Induction: Effects of Resveratrol Derivatives and Bortezomib on Human Multiple Myeloma Cells

Date of Award

2022

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Xinyu (Eric) Wang, PhD

Second Advisor

Harold Komiskey, PhD

Third Advisor

Srujana Rayalam, DVM, PhD

Fourth Advisor

Lori Redmond, PhD

Abstract

Multiple myeloma (MM) is a cancer of plasma cells in the bone marrow characterized by bone lesions, hypercalcemia, anemia, and renal failure. Bortezomib (BTZ), a common treatment for MM, is a proteasome inhibitor that induces apoptosis in MM cells. However, high doses of BTZ can be toxic, signifying a need for a synergistic drug combination. Resveratrol (RES) is a phenolic compound found in grapes shown to inhibit MM cell growth and induce apoptosis in these cells. We hypothesize that combining BTZ with a RES derivative at certain concentrations is synergistic in reducing viability and inducing apoptosis in MM cells.

To address this hypothesis, we performed cell viability assays to assess the effects of increasing concentrations of RES derivatives and BTZ on human MM RPMI 8226 cell viability over time (24, 48, 72 hours) and western blots to assess these compounds’ effects on proapoptotic protein expression in MM cells following 24 hour treatment. Viability assay results for individual and combination drug treatments were analyzed by CompuSyn software to identify synergistic drug combinations. One synergistic combination was selected for further analysis, and MM cells were treated for 24 hours with this combination. For these treated cells, western blots were used to assess proapoptotic protein expression and flow cytometry was used to determine the percentages of cells in apoptotic stages.

We found that BTZ as well as RES and its derivatives pinostilbene (PIN), piceatannol (PIC), and pterostilbene (PTS) decreased MM cell viability in a dose- and time-dependent manner and increased expression of cleaved proapoptotic proteins poly(ADP-ribose) polymerase 1 (PARP1) and caspase-3 in a dose-dependent manner. In analyzing different combinations of RES derivatives with BTZ with CompuSyn software, we selected the combination of 5 nM BTZ and 5 μM PIN for further assessment as a synergistic treatment. MM cells treated with the combination for 24 hours showed increased cleaved PARP1 and capase-3 expression and higher percentages of apoptotic cells versus cells treated with the individual compounds alone.

Our results suggest that a combination of 5 nM BTZ and 5 μM PIN decreases MM cell viability more than would be expected from the effects of each drug alone. This may be due to differences in how these compounds target the MM cells. Mechanistically, this combination seems to synergistically decrease MM cell viability through the induction of apoptosis.

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