Biodegradable synthetic high-density lipoprotein nanoparticles for atherosclerosis

Atherosclerosis remains one of the most common causes of death in the United States and throughout the world because of the lack of early detection. Macrophage apoptosis is a major contributor to the instability of atherosclerotic lesions. Development of an apoptosis targeted high-density lipoprotein (HDL)-mimicking nanoparticle (NP) to carry contrast agents for early detection of vulnerable plaques and the initiation of preventative therapies that exploit the vascular protective effects of HDL can be attractive for atherosclerosis. Here, we report the construction of a synthetic, biodegradable HDL-NP platform for detection of vulnerable plaques by targeting the collapse of mitochondrial membrane potential that occurs during apoptosis. This HDL mimic contains a core of biodegradable poly(lactic-co-glycolic acid), cholesteryl oleate, and a phospholipid bilayer coat that is decorated with triphenylphosphonium (TPP) cations for detection of mitochondrial membrane potential collapse. The lipid layer provides the surface for adsorption of apolipoprotein (apo) A-I mimetic 4F peptide, and the core contains diagnostically active quantum dots (QDs) for optical imaging. In vitro uptake, detection of apoptosis, and cholesterol binding studies indicated promising detection ability and therapeutic potential of TPP-HDL-apoA-I-QD NPs. In vitro studies indicated the potential of these NPs in reverse cholesterol transport. In vivo biodistribution and pharmacokinetics indicated favorable tissue distribution, controlled pharmacokinetic parameters, and significant triglyceride reduction for i.v.-injected TPP-HDL-apoA-I-QD NPs in rats. These HDL NPs demonstrate excellent biocompatibility, stability, nontoxic, and nonimmunogenic properties, which prove to be promising for future translation in early plaque diagnosis and might find applications to prevent vulnerable plaque progression

Formulation and Optimization of Mitochondria-Targeted Polymeric Nanoparticles

Targeted delivery of therapeutics to the mitochondria of cells without alteration of drug properties can be a vital technique in the treatment of a variety of mitochondrial-dysfunction-related diseases. Herein, we describe a detailed protocol for synthesis and characterization of a functionalized polymer to build mitochondria-targeted nanoparticles (NPs). The block polymer was decorated with a lipophilic triphenylphosphonium (TPP) cation for mitochondrial trafficking of payload-loaded polymeric NPs. TPP-based lipophilic cations have the ability to cross the mitochondrial membrane. A mitochondria-targeted block copolymer poly(dl-lactide-co-glycolide)-b-polyethylene glycol-TPP and a nontargeted poly(dl-lactide-co-glycolide)-b-polyethylene glycol polymer were synthesized and their NPs were prepared. A nanoprecipitation method combined with polymer blending technology was adopted in order to get suitable size and charged NPs for efficient mitochondrial trafficking

Immune Stimulating Photoactive Hybrid Nanoparticles for Metastatic Breast Cancer

A therapeutic technology that combines the phototoxic and immune-stimulating ability of photodynamic therapy (PDT) with the widespread effectiveness of the immune system can be very promising to treat metastatic breast cancer. We speculated that the knowledge of molecular mechanisms of existing multi-component therapies could provide clues to aid the discovery of new combinations of an immunostimulant with a photosensitizer (PS) using a nanoparticle (NP) delivery platform. Therapeutic challenges when administering therapeutic combinations include the choice of dosages to reduce side effects, the definitive delivery of the correct drug ratio, and exposure to the targets of interest. These factors are very difficult to achieve when drugs are individually administered. By combining controlled release polymer-based NP drug delivery approaches, we were able to differentially deliver zinc phthalocyanine (ZnPc) based PS to metastatic breast cancer cells along with CpG-ODN, a single-stranded DNA that is a known immunostimulant to manage the distant tumors in a temporally regulated manner resulting in more effective management of deadly metastatic breast cancer. We encapsulated ZnPc which is a long-wavelength absorbing PS within a polymeric NP core made up of poly(D,L-lactic- co -glycolic acid)- b -poly(ethylene glycol) (PLGA- b -PEG). After coating the outside of the polymeric core with gold NPs (AuNPs), we further modified the AuNP surface with CpG-ODN. In vitro cytotoxicity using 4T1 metastatic mouse breast carcinoma cells shows significant photocytotoxicity of the hybrid NPs containing both ZnPc and CpG-ODN after irradiation with a 660 nm LASER light and this activity was remarkably better than either treatment alone. Treatment of mouse bone marrow derived dendritic cells with the PDT-killed 4T1 cell lysate shows that the combination of PDT with a synergistic immunostimulant in a single NP system results in significant immune response, which can be used for the treatment of metastatic cancer

<i>Ex Vivo</i> Programming of Dendritic Cells by Mitochondria-Targeted Nanoparticles to Produce Interferon-Gamma for Cancer Immunotherapy

One of the limitations for clinical applications of dendritic cell (DC)-based cancer immunotherapy is the low potency in generating tumor antigen specific T cell responses. We examined the immunotherapeutic potential of a mitochondria-targeted nanoparticle (NP) based on a biodegradable polymer and zinc phthalocyanine (ZnPc) photosensitizer (T-ZnPc-NPs). Here, we report that tumor antigens generated from treatment of breast cancer cells with T-ZnPc-NPs upon light stimulation activate DCs to produce high levels of interferon-gamma, an important cytokine considered as a product of T and natural killer cells. The remarkable <i>ex vivo</i> DC stimulation ability of this tumor cell supernatant is a result of an interleukin (IL)-12/IL-18 autocrine effect. These findings contribute to the understanding of how <i>in situ</i> light activation amplifies the host immune responses when NPs deliver the photosensitizer to the mitochondria and open up the possibility of using mitochondria-targeted-NP-treated, light-activated cancer cell supernatants as possible vaccines

Mito-DCA: A Mitochondria Targeted Molecular Scaffold for Efficacious Delivery of Metabolic Modulator Dichloroacetate

Tumor growth is fueled by the use of glycolysis, which normal cells use only in the scarcity of oxygen. Glycolysis makes tumor cells resistant to normal death processes. Targeting this unique tumor metabolism can provide an alternative strategy to selectively destroy the tumor, leaving normal tissue unharmed. The orphan drug dichloroacetate (DCA) is a mitochondrial kinase inhibitor that has the ability to show such characteristics. However, its molecular form shows poor uptake and bioavailability and limited ability to reach its target mitochondria. Here, we describe a targeted molecular scaffold for construction of a multiple DCA loaded compound, Mito-DCA, with three orders of magnitude enhanced potency and cancer cell specificity compared to DCA. Incorporation of a lipophilic triphenylphosphonium cation through a biodegradable linker in Mito-DCA allowed for mitochondria targeting. Mito-DCA did not show any significant metabolic effects toward normal cells but tumor cells with dysfunctional mitochondria were affected by Mito-DCA, which caused a switch from glycolysis to glucose oxidation and subsequent cell death <i>via</i> apoptosis. Effective delivery of DCA to the mitochondria resulted in significant reduction in lactate levels and played important roles in modulating dendritic cell (DC) phenotype evidenced by secretion of interleukin-12 from DCs upon activation with tumor antigens from Mito-DCA treated cancer cells. Targeting mitochondrial metabolic inhibitors to the mitochondria could lead to induction of an efficient antitumor immune response, thus introducing the concept of combining glycolysis inhibition with immune system to destroy tumor

Evaluation of nanoparticle delivered cisplatin in beagles

Intracranial neoplasia is a significant cause of morbidity and mortality in both human and veterinary patients, and is difficult to treat with traditional therapeutic methods. Cisplatin is a platinum (Pt)-containing chemotherapeutic agent approved by the Food and Drug Administration; however, substantial limitations exist for its application in canine brain tumor treatment due to the difficulty in crossing the blood-brain barrier (BBB), development of resistance, and toxicity. A modified Pt(iv)-prodrug of cisplatin, Platin-M, was recently shown to be deliverable to the brain via a biocompatible mitochondria-targeted lipophilic polymeric nanoparticle (NP) that carries the drug across the BBB and to the mitochondria. NP mediated controlled release of Platin-M and subsequent reduction of this prodrug to cisplatin allowed cross-links to be formed with the mitochondrial DNA, which have no nucleotide excision repair system, forcing the overactive cancer cells to undergo apoptosis. Here, we report in vitro effects of targeted Platin-M NPs (T-Platin-M-NPs) in canine glioma and glioblastoma cell lines with results indicating that this targeted NP formulation is more effective than cisplatin. In both the cell lines, T-Platin-M-NP was significantly more efficacious compared to carboplatin, another Pt-based chemotherapy, which is used in the settings of recurrent high-grade glioblastoma. Mitochondrial stress analysis indicated that T-Platin-M-NP is more effective in disrupting the mitochondrial bioenergetics in both the cell types. A 14-day distribution study in healthy adult beagles using a single intravenous injection at 0.5 mg kg(-1) (with respect to Platin-M) of T-Platin-M-NPs showed high levels of Pt accumulation in the brain, with negligible amounts in the other analyzed organs. Safety studies in the beagles monitoring physical, hematological, and serum chemistry evaluations were within the normal limits on days 1, 7, and 14 after injection of either 0.5 mg kg(-1) or 2 mg kg(-1) or 2.2 mg kg(-1) (with respect to Platin-M) of T-Platin-M-NPs. At all doses over the 14-day period, no neurotoxicity was observed based upon periodic neurological examinations and cerebrospinal fluid analysis. These studies demonstrated the translational nature of T-Platin-M-NPs for applications in the treatment of brain tumors

Nanocarriers for tracking and treating diseases

Site directed drug delivery with high efficacy is the biggest challenge in the area of current pharmaceuticals. Biodegradable polymer-based controlled release nanoparticle platforms could be beneficial for targeted delivery of therapeutics and contrast agents for a myriad of important human diseases. Biodegradable nanoparticles, which can be engineered to load multiple drugs with varied physicochemical properties, contrast agents, and cellular or intracellular component targeting moieties, have emerged as potential alternatives for tracking and treating human diseases. In this review, we will highlight the current advances in the design and execution of such platforms for their potential application in the diagnosis and treatment of variety of diseases ranging from cancer to Alzheimer's and we will provide a critical analysis of the associated challenges for their possible clinical translation

Abstract 1025: Optimized DZNep exposure presensitizes pancreatic cancer cells to anticancer nucleoside analogues: potential clinical implications

Abstract We evaluated the potential of a histone methylation reversal agent 3-deazaneplanocin A (DZNep) in improving the chemosensitivity of pancreatic cancer to nucleoside analogs (i.e., gemcitabine). DZNep brought delayed but selective cytotoxicity to pancreatic cancer cells without affecting normal human pancreatic ductal epithelial (HPDE) cells. Co-exposure of DZNep and gemcitabine induced cytotoxic additivity or synergism in both well- and poorly-differentiated pancreatic cell lines. In contrast, DZNep exerted antagonism with gemcitabine against HPDE cells with significant reduction in cytotoxicity compared with the gemcitabine-alone regimen. DZNep marginally depended on purine nucleoside transporters for its cytotoxicity, but the transport dependence was circumvented by acyl derivatization. Drug exposure studies revealed that a short priming with DZNep followed by gemcitabine treatment rather than co-treatment of both agents produced a maximal chemosensitization response in both gemcitabine-sensitive and gemcitabine-resistant pancreatic cancer cells. DZNep rapidly and reversibly decreased trimethylation of histone H3 lysine 27 but increased trimethylation of lysine 9 in an EZH2- and JMJD1A/2C-dependent manner, respectively. However, DZNep potentiation of nucleoside analog chemosensitization was found to be temporally coupled to trimethylation changes in lysine 27 and not lysine 9. Polymeric nanoparticles engineered to chronologically release DZNep followed by gemcitabine produced pronounced chemosensitization and dose-lowering effects. Together, our results identify that an optimized DZNep exposure can presensitize pancreatic cancer cells to anticancer nucleoside analogs and emphasize the promising clinical utilities of histone methylation reversal agents in future pancreatic cancer combination therapies. Citation Format: Sau Wai Hung, Hardik Mody, Sean Marrache, Yangzom D. Bhutia, Franklin Davis, Jong Hyun Cho, Shanta Dhar, Chung K. Chu, Rajgopal Govindarajan. Optimized DZNep exposure presensitizes pancreatic cancer cells to anticancer nucleoside analogues: potential clinical implications. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1025. doi:10.1158/1538-7445.AM2013-1025</jats:p

Core hydrophobicity tuning of a self-assembled particle results in efficient lipid reduction and favorable organ distribution

Atherosclerosis, the deadliest disease in the United States, arises due to the build up of plaques in the arteries as a result of excessive cholesterol deposition and an impaired cholesterol removal process. High density lipoproteins (HDL), popularly known as “good cholesterol”, are naturally occurring nano-sized particles that, along with apolipoproteins, are deployed to maintain cholesterol homeostasis in the body. Both cholesterol efflux, from the fat-laden macrophages in the arteries, and intracellular lipid transport, to deliver cholesterol to the mitochondria of liver cells for metabolism, hold key responsibilities to maintain healthy lipid levels inside the body. We designed a library of nine mitochondria targeted polymer–lipid hybrid nanoparticles (NPs), comprised of completely synthetic yet biodegradable components, that are capable of performing HDL-like functions. Using this library, we optimized a superior mitochondria targeted NP candidate, which can show favourable organ distribution, therapeutic potential, and non-toxic properties. Two targeted NP formulations with optimum NP size, zeta potential, and cholesterol binding and release properties were identified. Lipid reduction and anti-oxidative properties of these two NPs demonstrated cholesterol removal ability.In vivotherapeutic evaluation of the targeted-NP formulations in apolipoprotein E knockout (apoE−/−) mice indicated lipid reduction and anti-inflammatory properties compared to non-targeted NPs. This synthetic targeted NP with potential abilities to participate in both extra- and intracellular cholesterol transport might potentiate therapeutic interventions for heart diseases

Pharmacological reversal of histone methylation presensitizes pancreatic cancer cells to nucleoside drugs: in vitro optimization and novel nanoparticle delivery studies

We evaluated the potential of an investigational histone methylation reversal agent, 3-deazaneplanocin A (DZNep), in improving the chemosensitivity of pancreatic cancer to nucleoside analogs (i.e., gemcitabine). DZNep brought delayed but selective cytotoxicity to pancreatic cancer cells without affecting normal human pancreatic ductal epithelial (HPDE) cells. Co-exposure of DZNep and gemcitabine induced cytotoxic additivity or synergism in both well- and poorly-differentiated pancreatic cell lines by increased apoptosis. In contrast, DZNep exerted antagonism with gemcitabine against HPDE cells with significant reduction in cytotoxicity compared with the gemcitabine-alone regimen. DZNep marginally depended on purine nucleoside transporters for its cytotoxicity, but the transport dependence was circumvented by acyl derivatization. Drug exposure studies revealed that a short priming with DZNep followed by gemcitabine treatment rather than co-treatment of both agents to produce a maximal chemosensitization response in both gemcitabine-sensitive and gemcitabine-resistant pancreatic cancer cells. DZNep rapidly and reversibly decreased trimethylation of histone H3 lysine 27 but increased trimethylation of lysine 9 in an EZH2- and JMJD1A/2C-dependent manner, respectively. However, DZNep potentiation of nucleoside analog chemosensitization was found to be temporally coupled to trimethylation changes in lysine 27 and not lysine 9. Polymeric nanoparticles engineered to chronologically release DZNep followed by gemcitabine produced pronounced chemosensitization and dose-lowering effects. Together, our results identify that an optimized DZNep exposure can presensitize pancreatic cancer cells to anticancer nucleoside analogs through the reversal of histone methylation, emphasizing the promising clinical utilities of epigenetic reversal agents in future pancreatic cancer combination therapies