Abstract 3618: The mechanism of functional lipoprotein-like nanoparticle (FLip-NP) induced lymphoma cell death

Abstract

Abstract Introduction: Diffuse large B-cell lymphoma (DLBCL) cells have an increased demand for cholesterol and cholesteryl esters to maintain membrane anchored, pro-survival signaling pathways, such as B-cell receptor (BCR) signaling. We have previously reported that synthetic, biomimetic functional lipoprotein gold nanoparticles (FLip-NPs) induce lymphoma cell death in vitro and in vivo through targeted, receptor-mediated cholesterol depletion (Yang et al PNAS 2013) and that cellular cholesterol levels influence response to FLip-NPs (Rink et al Molecular Pharmaceutics 2017). Given the critical role of cholesterol in maintaining proper membrane organization to enable intracellular signaling, we hypothesized that FLip-NP-induced cholesterol depletion reorganizes the plasma membrane, leading to reduced membrane anchored signaling. Additionally, we hypothesized that FLip-NPs will synergize with small molecule inhibitors of BCR-related kinases, leading to enhanced lymphoma cell death. Methods: SUDHL4 [germinal center (GC) DLBCL] and Ramos [Burkitt’s lymphoma] cells were used for experiments. Confocal fluorescent microscopy was used to visualize changes in membrane organization following FLip-NP treatment. Changes in the localization and phosphorylation of signaling kinases (e.g. AKT, ERK1/2) following FLip-NP treatment were measured using immunoprecipitation, phospho-kinase arrays, and western blot assays. The Amplex Red Cholesterol Assay was used to measure total cellular cholesterol, while cell viability was quantified using the MTS assay. Results: In SUDHL4 and Ramos cells, FLip-NPs induced a reorganization of the plasma membrane, clustering of the HDL receptor, scavenger receptor type B1. This reorganization correlated with decreased phosphorylation of the signaling kinases AKT, ERK1/2, LCK and LYN, as well as a reduction in total cellular cholesterol. Given the decreases in pro-survival kinase phosphorylation, we investigated whether small molecule inhibitors of these kinases enhanced FLip-NP efficacy. FLip-NPs synergized with inhibitors of PI3K (idelalisib, duvelisib), SYK (R406), and AKT (GDC-0068), increasing lymphoma cell death. Finally, addition of a PTEN inhibitor, to prevent AKT de-phosphorylation, rescued Ramos and SUDHL4 from FLip-NP induced cell death. Conclusion: These data demonstrate that FLip-NPs reduce B-cell lymphoma cell cholesterol, inducing changes in the organization of the cell membrane and, subsequently, a series of changes to membrane-anchored pro-survival and pro-proliferative signaling pathways. Enhancing cholesterol starvation strategies, such as the combination of the targeted cholesterol depletion agent FLip-NPs with small molecule inhibitors of signaling kinases, represents a novel therapeutic strategy for DLBCL and may be applicable to other cholesterol dependent malignancies. Citation Format: Jonathan S. Rink, Shuo Yang, Adam Lin, Reem Karmali, Colby S. Thaxton, Leo I. Gordon. The mechanism of functional lipoprotein-like nanoparticle (FLip-NP) induced lymphoma cell death [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3618.