Abstract 2044: Development of stable and brain-penetrating disulfiram nanoparticles, characterization and efficacy in glioma cell culture and xenograft models
Authors: Hanumantha Rao Madala, Surendra R. Punganuru, Francis Ali-Osman, Kalkunte S. Srivenugopal
Journal: Cancer Research (2017)

Abstract Blood brain-barrier entry, tumor heterogeneity, need to eliminate the tumorigenic stem cells and inhibit the DNA repair protein MGMT are all essential considerations in glioma treatment. Previously, we showed that alcohol aversion drug disulfiram (DSF) shares all these properties [Carcinogenesis 35, 692, 2014; Oncotarget 4,502, 2013]. DSF was shown to inhibit the MGMT activity in the same way as ALDH by conjugating with the active-site cysteine 145 and sensitize brain tumors to alkylating agents. Although DSF exerts significant anticancer effects in vitro, the drug undergoes rapid decomposition and metabolism. To overcome this problem, we engineered DSF nanoparticles by encapsulating the drug in PEG: PLGA polymer by solvent evaporation method. The formulation was optimized for various process formulation variables. The optimized nano-formulation (NP) ranged from 70-80 nm in particle size as confirmed using dynamic light scattering and transition electron microscopy. Drug loading, encapsulation efficiency, in vitro release were also optimal and the particles showed excellent BBB permeability in vitro. The time course of internalization of particles using a cyanine dye (HITC-1) showed a lysosomal accumulation followed by mitochondria. Live animal imaging after IV injections of HITC-labeled DSF-NPs revealed a selective accumulation in the brain and subcutaneous tumors. Further, the cytotoxicity elicited by DSFNPs against SF188, DAOY, UW18, UW28 and T98G brain tumor cells was extended and greater than obtained with the free drug. When combined with DSF-NPs, temozolomide showed an 8-10-fold increased cytotoxicity against glioma cell lines. The DSFNPs also induced significant degradation of MGMT and other redox-sensitive proteins such as the NF-κB, MDR1 and GSTP1 in a concentration-dependent manner in glioma cells. ROS induction by the encapsulated drug was confirmed through DCF-DA and DHE staining of live cells; co-staining these cells with the mitochondrial membrane potential markers DiOC6 and DHE confirmed the altered integrity. GSTP1 inhibition and ROS induction by DSFNPs led to an activation of JNK/MAPK pathway, culminating in autophagy and apoptosis as verified by western blotting, FACS and immunofluorescence of appropriate markers. Finally, the antitumor efficacy of DSFNPs was tested in intracranial glioblastoma developed in nude mice by injecting luciferase-expressing SF-188 GBM cells. A superior tumor regression was evident in DSFNP-administered animals compared with the free drug after IVIS bioluminescence and histochemical staining of brain tissue. Collectively, our efforts provide strong evidence that DSF nanoparticles can cross the BBB and offer a sustained drug supply to eliminate CNS cancers. Further refinement of DSFNPs by incorporating brain targeting strategies is underway (supported by CPRIT grants RP130266 &amp; RP170207 to KSS). Citation Format: Hanumantha Rao Madala, Surendra R. Punganuru, Francis Ali-Osman, Kalkunte S. Srivenugopal. Development of stable and brain-penetrating disulfiram nanoparticles, characterization and efficacy in glioma cell culture and xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2044. doi:10.1158/1538-7445.AM2017-2044