Inhibitory effects and targeting mechanisms of caffeic acid phenethyl ester (CAPE) in human multiple myeloma cells
Location
Suwanee, GA
Start Date
7-5-2024 1:00 PM
End Date
7-5-2024 4:00 PM
Description
Background: Multiple Myeloma (MM) is a cancer of plasma cells in the bone marrow microenvironment which results in elevated levels of abnormal monoclonal proteins (M-proteins) in the blood. These M-proteins crowd out normally functioning antibodies, preventing them from fighting infection. In addition to immunosuppression, symptoms of MM include anemia, hypercalcemia, renal failure, and bone lesions. It is estimated that there will be 35,780 new cases in the United States in 2024 with a 5-year survival rate of 60%. Caffeic acid phenethyl ester (CAPE) is a phytochemical reported to have a plethora of bioactive properties including antitumor traits. We previously reported that CAPE inhibits growth of MM cells through induction of apoptosis and oxidative stress. The objective of this study is to evaluate the effects of novel CAPE derivatives on human MM cell growth.
Methods: CAPE derivatives were synthesized by coupling caffeic acid with derivatives of phenylalanine and tyrosine. MM cells were treated with CAPE and derivatives (0 – 40 μM) for 24, 48, or 72 hours, and effects on cell viability were measured using Presto Blue assays. Effects of CAPE derivatives (50 μM) on viability of the normal cell counterpart (human peripheral blood mononuclear cells) were also measured. Two potent derivatives (CAPE-7 and CAPE-9) were identified and assessed for their effects on apoptosis induction in MM cells through western blotting for apoptotic markers (caspase-3 and PARP-1) and flow cytometric analysis of apoptotic stages. Effects of these compounds on oxidative stress in MM RPMI 8226 cells were measured by assessing cell viability changes from pretreatment with antioxidant N-acetyl cysteine (NAC) or glutathione inhibitor buthionine sulfoximine (BSO). Reactive oxygen species (ROS) production was assessed with a dichlorodihydrofluorescein (DCFDA) assay. Effects on antioxidant expression in MM cells were assessed with western blotting for catalase, superoxide dismutase 1, and thioredoxin. In silico studies were conducted to predict pharmacokinetic profiles of the two derivatives by ADMET predictorTM software.
Results: CAPE and most of the 11 synthesized derivatives had the greatest cytotoxic effects in the MM RPMI 8226 cell line compared to U266 and NCI-H929. CAPE-7 and CAPE-9 were identified as the most potent derivatives across the MM cell lines and exhibited minimal toxicity in PBMCs. Interestingly, CAPE-7 and CAPE-9 did not significantly induce apoptosis in MM cells apart from CAPE-9 in U266 cells. Pretreatment of MM RPMI 8226 cells with BSO sensitized the cells to treatment by CAPE, CAPE-7, and CAPE-9, while NAC did not significantly affect viability. CAPE-7 and CAPE-9 reduced ROS production in RPMI 8226 cells similarly to CAPE, while significantly decreased protein expression of catalase and thioredoxin versus the control. ADMET predictions for these compounds indicated slight risk of mutagenicity but lower overall ADMET risk compared to CAPE.
Discussion: Several synthesized CAPE derivatives exhibit cytotoxic effects in human MM cells which may shed light on the structure activity relationship of CAPE. Our results suggest the more potent derivatives CAPE-7 and CAPE-9 target MM cells non-apoptotically and via oxidative stress by downregulating antioxidant expression.
Embargo Period
5-23-2024
Inhibitory effects and targeting mechanisms of caffeic acid phenethyl ester (CAPE) in human multiple myeloma cells
Suwanee, GA
Background: Multiple Myeloma (MM) is a cancer of plasma cells in the bone marrow microenvironment which results in elevated levels of abnormal monoclonal proteins (M-proteins) in the blood. These M-proteins crowd out normally functioning antibodies, preventing them from fighting infection. In addition to immunosuppression, symptoms of MM include anemia, hypercalcemia, renal failure, and bone lesions. It is estimated that there will be 35,780 new cases in the United States in 2024 with a 5-year survival rate of 60%. Caffeic acid phenethyl ester (CAPE) is a phytochemical reported to have a plethora of bioactive properties including antitumor traits. We previously reported that CAPE inhibits growth of MM cells through induction of apoptosis and oxidative stress. The objective of this study is to evaluate the effects of novel CAPE derivatives on human MM cell growth.
Methods: CAPE derivatives were synthesized by coupling caffeic acid with derivatives of phenylalanine and tyrosine. MM cells were treated with CAPE and derivatives (0 – 40 μM) for 24, 48, or 72 hours, and effects on cell viability were measured using Presto Blue assays. Effects of CAPE derivatives (50 μM) on viability of the normal cell counterpart (human peripheral blood mononuclear cells) were also measured. Two potent derivatives (CAPE-7 and CAPE-9) were identified and assessed for their effects on apoptosis induction in MM cells through western blotting for apoptotic markers (caspase-3 and PARP-1) and flow cytometric analysis of apoptotic stages. Effects of these compounds on oxidative stress in MM RPMI 8226 cells were measured by assessing cell viability changes from pretreatment with antioxidant N-acetyl cysteine (NAC) or glutathione inhibitor buthionine sulfoximine (BSO). Reactive oxygen species (ROS) production was assessed with a dichlorodihydrofluorescein (DCFDA) assay. Effects on antioxidant expression in MM cells were assessed with western blotting for catalase, superoxide dismutase 1, and thioredoxin. In silico studies were conducted to predict pharmacokinetic profiles of the two derivatives by ADMET predictorTM software.
Results: CAPE and most of the 11 synthesized derivatives had the greatest cytotoxic effects in the MM RPMI 8226 cell line compared to U266 and NCI-H929. CAPE-7 and CAPE-9 were identified as the most potent derivatives across the MM cell lines and exhibited minimal toxicity in PBMCs. Interestingly, CAPE-7 and CAPE-9 did not significantly induce apoptosis in MM cells apart from CAPE-9 in U266 cells. Pretreatment of MM RPMI 8226 cells with BSO sensitized the cells to treatment by CAPE, CAPE-7, and CAPE-9, while NAC did not significantly affect viability. CAPE-7 and CAPE-9 reduced ROS production in RPMI 8226 cells similarly to CAPE, while significantly decreased protein expression of catalase and thioredoxin versus the control. ADMET predictions for these compounds indicated slight risk of mutagenicity but lower overall ADMET risk compared to CAPE.
Discussion: Several synthesized CAPE derivatives exhibit cytotoxic effects in human MM cells which may shed light on the structure activity relationship of CAPE. Our results suggest the more potent derivatives CAPE-7 and CAPE-9 target MM cells non-apoptotically and via oxidative stress by downregulating antioxidant expression.