M2pep-Modified Cyclodextrin-siRNA Nanoparticles Modulate the Immunosuppressive Tumor Microenvironment for Prostate Cancer Therapy.

Prostate cancer (PCa) is the most prevalent cause of cancer deaths in men. Conventional strategies, such as surgery, radiation, or chemotherapy, face challenges including poor prognosis and resistance. Therefore, the development of new improved strategies is vital to enhance patient outcomes. Recently, immunotherapy has shown potential in the treatment of a range of cancers, including PCa. Tumor-associated macrophages (TAMs) play an important role in the tumor microenvironment (TME) and reprogramming of TAMs is associated with remodeling the TME. The colony-stimulating factor-1/colony stimulating factor-1 receptor (CSF-1/CSF-1R) signaling pathway is closely related to the polarization of TAMs. The downregulation of CSF-1R, using small interfering RNA (siRNA), has been shown to achieve the reprogramming of TAMs, from the immunosuppressive M2 phenotype to the immunostimulatory M1 one. To maximize specific cellular delivery an M2 macrophage-targeting peptide, M2pep, was formulated with an amphiphilic cationic ß-Cyclodextrin (CD) incorporating CSF-1R siRNA. The resulting nanoparticles (NPs) increased M2 macrophage targeting both in vitro and in vivo, promoting the release of M1 factors and simultaneously downregulating the levels of M2 factors through TAM reprogramming. The subsequent remodeling of the TME resulted in a reduction in tumor growth in a subcutaneous PCa mouse model mainly mediated through the recruitment of cytotoxic T cells. In summary, this M2pep-targeted CD-based delivery system demonstrated significant antitumor efficacy, thus presenting an alternative immunotherapeutic strategy for PCa treatment.

Sialic acid-targeted cyclodextrin-based nanoparticles deliver CSF-1R siRNA and reprogram tumour-associated macrophages for immunotherapy of prostate cancer.

Prostate cancer remains a serious condition threatening the health of men. Due to the complicated nature of the tumour microenvironment (TME), conventional treatments face challenges including poor prognosis and tumour resistance, therefore new therapeutic strategies are urgently needed. Small interfering RNA (siRNA), a double-stranded non-coding RNA, regulates specific gene expression through RNA interference. Tumour-associated macrophages (TAMs) are a potential therapeutic target in cancer immunotherapy. Colony stimulating factor-1/colony stimulating factor-1 receptor (CSF-1/CSF-1R) signaling pathway plays a crucial role in the polarization of the immunosuppressive TAMs, M2 macrophages. Downregulation of CSF-1R is known to reprogram the immunosuppressive TAMs, M2 macrophages, to the immunostimulatory phenotype, M1 macrophages. Sialic acid is a ligand for Siglec-1 (CD169) which is overexpressed on M2 macrophages with little expression in other phenotypes. Therefore, a sialic acid-targeted cyclodextrin-based nanoparticle was developed to specifically deliver CSF-1R siRNA to M2 macrophages. The nanoparticles were studied in vitro using both human and mouse prostate cancer cell lines. Results show that the targeted nanoparticles achieved cell specific delivery to M2 macrophages via the sialic acid-CD169 axis. The expression of CSF-1R was significantly downregulated in M2 macrophages (29.64% for targeted vs 19.31% for non-targeted nanoparticles in THP-1-derived M2 macrophages and 38.94% for targeted vs 18.51% for non-targeted nanoparticles in RAW 264.7-derived M2 macrophages, n = 4, p < 0.01). The resulting reprograming of M2 macrophages to M1 enhanced the level of apoptosis in the prostate cancer cells in a Transwell model (49.17% for targeted vs 37.68% for non-targeted nanoparticles in PC-3 cells and 69.15% for targeted vs 44.73% for non-targeted nanoparticles in TRAMP C1 cells, n = 3, p < 0.01). Thus, this targeted cyclodextrin-based siRNA drug delivery system provides a potential strategy for prostate cancer immunotherapy.

Co-Formulation of Amphiphilic Cationic and Anionic Cyclodextrins Forming Nanoparticles for siRNA Delivery in the Treatment of Acute Myeloid Leukaemia.

Non-viral delivery of therapeutic nucleic acids (NA), including siRNA, has potential in the treatment of diseases with high unmet clinical needs such as acute myeloid leukaemia (AML). While cationic biomaterials are frequently used to complex the nucleic acids into nanoparticles, attenuation of charge density is desirable to decrease in vivo toxicity. Here, an anionic amphiphilic CD was synthesised and the structure was confirmed by Fourier-transform infrared spectroscopy (FT-IR), Nuclear Magnetic Resonance (NMR), and high-resolution mass spectrometry (HRMS). A cationic amphiphilic cyclodextrin (CD) was initially used to complex the siRNA and then co-formulated with the anionic amphiphilic CD. Characterisation of the co-formulated NPs indicated a significant reduction in charge from 34 ± 7 mV to 24 ± 6 mV (p < 0.05) and polydispersity index 0.46 ± 0.1 to 0.16 ± 0.04 (p < 0.05), compared to the cationic CD NPs. Size was similar, 161−164 nm, for both formulations. FACS and confocal microscopy, using AML cells (HL-60), indicated a similar level of cellular uptake (60% after 6 h) followed by endosomal escape. The nano co-formulation significantly reduced the charge while maintaining gene silencing (21%). Results indicate that blending of anionic and cationic amphiphilic CDs can produce bespoke NPs with optimised physicochemical properties and potential for enhanced in vivo performance in cancer treatment.

The toxicity of silver nanomaterials (NM 300K) is reduced when combined with N-Acetylcysteine: Hazard assessment on Enchytraeus crypticus.

The widespread production and use of silver nanomaterials (AgNMs) in consumer and medical products have been raising environmental concerns. Once in the environment, the soil is one of the major sinks of AgNMs due to e.g. sewage sludge applications, and invertebrates are directly exposed. In this study, we investigate the potential of N-acetylcysteine (NAC) to reduce the toxic effects of Ag NM300 K (and AgNO3) on the soil invertebrate Enchytraeus crypticus. Ag NM300 K induces mortality, reproduction impairment, and avoidance. The addition of NAC to the soil showed a remarkable reduction in the toxicity of Ag, indicating that NAC can act as a detoxifying agent for terrestrial organisms exposed to Ag materials. That the reduction in toxicity likely is caused by thiol groups, was confirmed by GSH and GSSH studies. Identifying the mechanisms and hence alternatives that allow the recovery of contaminated soils is an important mitigation measure to promote environmental safety and reduce the associated risks to human health. Further, it may inform on strategies to implement in safe-by-design industry development.

Soft Nanohydrogels Based on Laponite Nanodiscs: A Versatile Drug Delivery Platform for Theranostics and Drug Cocktails.

A new nanohydrogel drug delivery platform based on Laponite nanodiscs, polyacrylate, and sodium phosphate salts is described. The hybrid nanohydrogel is tailored to obtain soft and flexible nanohydrogels with G' around 3 kPa, which has been proposed as the ideal stiffness for drug delivery applications. In vitro studies demonstrate that the new nanohydrogels are biocompatible, biodegradable, nonswellable, pH-responsive, and noncytotoxic and are able to deliver antineoplastic drugs into cancer cells. The IC50 of nanohydrogels containing cisplatin, 4-fluorouracil, and cyclophosphamide is significantly lower than the IC50 of the free drugs. In vivo experiments suggest that the new nanomaterials are biocompatible and do not accumulate in crucial organs. The simple formulation procedure enables encapsulation of virtually any water-soluble molecule, without the need for chemical modification of the guests. These nanohydrogels are a versatile platform that enables the simultaneous encapsulation of several cancer drugs, yielding an efficient drug cocktail delivery system, which for instance presents a positive synergistic effect against MCF-7 cells.

Vascular endothelial growth factor increases during blood-brain barrier-enhanced permeability caused by Phoneutria nigriventer spider venom.

Phoneutria nigriventer spider accidental envenomation provokes neurotoxic manifestations, which when critical, results in epileptic-like episodes. In rats, P. nigriventer venom (PNV) causes blood-brain barrier breakdown (BBBb). The PNV-induced excitotoxicity results from disturbances on Na(+), K(+) and Ca(2+) channels and glutamate handling. The vascular endothelial growth factor (VEGF), beyond its angiogenic effect, also, interferes on synaptic physiology by affecting the same ion channels and protects neurons from excitotoxicity. However, it is unknown whether VEGF expression is altered following PNV envenomation. We found that adult and neonates rats injected with PNV showed immediate neurotoxic manifestations which paralleled with endothelial occludin, ß-catenin, and laminin downregulation indicative of BBBb. In neonate rats, VEGF, VEGF mRNA, and Flt-1 receptors, glutamate decarboxylase, and calbindin-D28k increased in Purkinje neurons, while, in adult rats, the BBBb paralleled with VEGF mRNA, Flk-1, and calbindin-D28k increases and Flt-1 decreases. Statistically, the variable age had a role in such differences, which might be due to age-related unequal maturation of blood-brain barrier (BBB) and thus differential cross-signaling among components of the glial neurovascular unit. The concurrent increases in the VEGF/Flt-1/Flk-1 system in the cerebellar neuron cells and the BBBb following PNV exposure might imply a cytokine modulation of neuronal excitability consequent to homeostatic perturbations induced by ion channels-acting PNV neuropeptides. Whether such modulation represents neuroprotection needs further investigation.

Expression of VEGF and Flk-1 and Flt-1 receptors during blood-brain barrier (BBB) impairment following Phoneutria nigriventer spider venom exposure.

Apart from its angiogenic and vascular permeation activity, the vascular endothelial growth factor (VEGF) has been also reported as a potent neuronal protector. Newborn rats with low VEGF levels develop neuron degeneration, while high levels induce protective mechanisms in several neuropathological conditions. Phoneutria nigriventer spider venom (PNV) disrupts the blood-brain barrier (BBB) and causes neuroinflammation in central neurons along with excitotoxic signals in rats and humans. All these changes are transient. Herein, we examined the expression of VEGF and its receptors, Flt-1 and Flk-1 in the hippocampal neurons following envenomation by PNV. Adult and neonatal rats were evaluated at time limits of 2, 5 and 24 h. Additionally, BBB integrity was assessed by measuring the expression of occludin, ß-catenin and laminin and neuron viability was evaluated by NeuN expression. VEGF, Flt-1 and Flk-1 levels increased in PNV-administered rats, concurrently with respective mRNAs. Flt-1 and Flk-1 immunolabeling was nuclear in neurons of hippocampal regions, instead of the VEGF membrane-bound typical location. These changes occurred simultaneously with the transient decreases in BBB-associated proteins and NeuN positivity. Adult rats showed more prominent expressional increases of the VEGF/Flt-1/Flk-1 system and earlier recovery of BBB-related proteins than neonates. We conclude that the reactive expressional changes seen here suggest that VEGF and receptors could have a role in the excitotoxic mechanism of PNV and that such role would be less efficient in neonate rats.