Temporal Dynamics of Caspase Activation in PPI-treated Cancer Cells

by Ashley Utz

Faculty Mentor: Professor Randall Reif

Cancer cells rely on glycolysis even under normoxic conditions. The use of this pathway results in measurable intracellular acidification, which is characterized as an early event in the apoptosis program. The pH is restored by activation of voltage- gated proton pumps, preventing acidification. Proton pump inhibitors (PPIs), such as omeprazole, inhibit the H+/K+-ATPase system found at the secretory surface of gastric parietal cells. Research has shown that omeprazole is also capable of inducing caspase-dependent apoptosis in Jurkat T-lymphocytes. However, the effects of PPIs on caspase activity remain largely unknown. The goal of this study was to determine the temporal dynamics of caspase activity in Jurkat cells treated with omeprazole, dexlansoprazole, or esomeprazole for six hours. After the incubation period, cells were held in place by anti- CD71 antibodies on the device’s affinity surface and fluorescence microcopy was used to monitor caspase activity in real time. Caspase activation was observed over a six-hour period with the fluorogenic caspase probe, L-bisaspartic acid rhodamine 110 (D2R). Elucidation of the intensity and timing of caspase activation will be beneficial for evaluating PPIs as potential cancer therapeutics.

Asymmetric Synthesis of DEHP

By Hannah Harris

Faculty Mentor: Professor Davis Oldham

Di(2-ethylhexyl) phthalate (DEHP) is a chiral molecule used as a plasticizer in many commercial products, and its metabolites have been linked to endocrine disruption and other adverse health effects in mice. Differences in the toxicity of the enantiomeric forms are not well studied. In order to synthesize (R,R)-DEHP, 2-ethyl-1-hexanol was reacted with vinyl acetate and Amano Lipase PS in dichloromethane at 0˚C for 48 hours to yield (R)-2-ethyl-1-hexanol (1) (60% yield, 75:25 e.r.) and 2-ethyl-1-hexyl acetate (2) (83% yield). (1) was refluxed with phthalic anhydride and pyridine at 120˚C for 3 hours yielding crude (R)-MEHP (3). After purification via column chromatography, the percent yield of pure (R)-MEHP, confirmed by 1H NMR, was 54% (75:25 e.r.). (3) was reacted with (R)-2-ethyl-1-hexanol (73:27 e.r.), N,N-dicyclohexylcarbodiimide, and 4-dimethylaminopyridine in dichloromethane at room temperature for 20 hours. The resulting (R,R)-DEHP (4) was purified via column chromatography (39% yield, approx. 75:25 e.r.) and confirmed by 1H NMR. (2) was hydrolyzed by potassium hydroxide in ethanol for 30 minutes at room temperature. (S)-2-ethyl-1-hexanol (5) was recovered (105% yield, 83:17 e.r.) and refluxed following the procedure previously described to produce (S)-MEHP (6), confirmed by 1H NMR in 15% yield after purification. Future work will optimize the enzymatic resolution of (1) and explore the reduction of 2-ethyl-2-hexenal by baker’s yeast as a more efficient method for synthesizing (2). Subsequently, the two other enantiomers of DEHP will be synthesized.