L-asparaginase immobilization in supramolecular nanogels of PEG-grafted poly HPMA and bis(α-cyclodextrin) to enhance pharmacokinetics and lower enzyme antigenicity.

L-asparaginase (ASNase) enzyme has limited therapeutic use due to its poor pharmacokinetics and immunogenicity. To overcome these obstacles, we immobilized ASNase in biocompatible poly hydroxypropyl methacrylamide (P(HPMA))-based nanogels simply formed through the host-guest inclusion complex of ASNase-conjugated random copolymer of HPMA and polyethylene glycol (PEG) acrylate (P(HPMA-MPEGA)) and α-cyclodextrin dimer (bisCD) using cystamine as a linker. The effects of bisCD and polymer concentrations on particle size, gelation time, and recovery of enzyme activity were investigated. The ASNase-conjugated bisCD nanogels were discrete, homogeneous, and spherical with a mean projected diameter of 148 ± 41 nm. ASNase immobilized in the bisCD nanogels caused cytotoxicity on HL-60 cell line with IC50 of 3 IU/ml. In-vivo rat study revealed that the immobilized ASNase reduced the enzyme antigenicity and resulted in 8.1 folds longer circulation half-life than the native enzyme. Conclusively, immobilization of ASNase in P(HPMA-MPEGA) and bisCD supramolecular nanogels could enhance the therapeutic value of ASNase in cancer chemotherapy.

PEGylated nanohydrogels delivering anti-MicroRNA-21 suppress ovarian tumor-associated angiogenesis in matrigel and chicken chorioallantoic membrane models.

Introduction: Recently, MicroRNAs have gained increasing popularity as a novel nucleic acid-mediated medicine to regulate cancer-related protein expression. MicroRNA-21 (miR-21) is known as an oncogenic microRNA which is overexpressed in almost all cancers, including ovarian carcinoma that causes cisplatin (cis-Pt) resistance and vascular endothelial growth factor (VEGF) upregulation. So, miRNA-based therapy can be regarded as knocking down miR-21 expression, inducing tumor cell apoptosis, and suppressing tumor-associated angiogenesis. Methods: PEG5k-carboxymethylated polyethyleneimine nanohydrogels (PEG5k-CMPEI) were loaded with AntagomiR-21 (As-21) at different ratios of nitrogen to phosphorus (N/P). Particle size and ζ potential were determined for the As-21 loaded nanohydrogels. In the cellular experiments, miR-21 expression, cytotoxicity, and cis-Pt sensitivity were studied on A2780 ovarian cancer cell lines. Finally, tumor cell apoptosis and tumor cell-associated angiogenesis were explored in vitro and in vivo. Results: The nanohydrogels, featuring homogeneous size distribution and redox-responsiveness, were steadily loaded by As-21 at the optimum N/P ratio of 5 without any aggregation as determined by transmission electron microscopy (TEM). As-21-loaded nanohydrogels caused sequence-specific suppression of miR-21 expression and provoked apoptosis through ROS generation and caspase 3 activation. Cisplatin cytotoxicity was remarkably enhanced in A2780R as compared to A2780S following co-incubation with As-21-loaded nanohydrogels. Interestingly, the condition of the medium derived from As-21 nanohydrogel-treated A2780R cells inhibited VEGF suppression in human umbilical vein endothelial cells (HUVECs) and the formation of tubes in Matrigel. Moreover, the condition medium caused angiogenesis inhibition in the chicken chorioallantoic membrane (CAM) model. Conclusion: These results suggest that nanohydrogel-based delivery of As-21 can be a promising neoadjuvant therapy for treating resistant tumors via apoptosis induction and angiogenesis suppression.

Biotin receptor-targeting nanogels loaded with methotrexate for enhanced antitumor efficacy in triple-negative breast cancer in vitro and in vivo models.

High-dose methotrexate (MTX) chemotherapeutic applications confront drug specificity and pharmacokinetic challenges, which can be overcome by utilizing targeted drug delivery systems. In the present study, biotin-PEG conjugated nanogels of carboxymethyl polyethyleneimine (Biotin-PEG-CMPEI) were developed for active targeted delivery of MTX in triple negative breast cancer (TNBC). TEM and DLS analyses revealed uniform, discrete, and spherical particles with a mean hydrodynamic diameter of about 100 nm and ζ-potential of + 15 mV (pH = 7.4). Biotin-PEG-CMPEI nanogels exhibited a zero-order MTX release kinetics at pH = 7.5 and a swelling-controlled release at pH = 5.5. In 4 T1 cells treated with the MTX-loaded Biotin-PEG-CMPEI, the IC50 was reduced by about 10 folds compared to the free drug, while the unloaded nanogels showed no significant toxicity. In the model mice, the group treated with the MTX-loaded Biotin-PEG-CMPEI had a lower tumor volume and mortality rate animal model when compared to free drug. Additionally, histopathological analyses showed that the group treated with the MTX-loaded nanogels had less lung metastasis and glomerular damage caused by MTX. Overall, the MTX-loaded Biotin-PEG-CMPEI targeted directly against overexpressed biotin receptors in TNBC have been shown to improve the MTX safety and therapeutic efficacy.

Single-chain antibody-decorated Au nanocages@liposomal layer nanoprobes for targeted SERS imaging and remote-controlled photothermal therapy of melanoma cancer cells.

The development of theranostic platforms combining surface-enhanced Raman spectroscopy (SERS) imaging with NIR-stimulated photothermal therapy (PTT) is of utmost importance for the precise diagnosis and selective treatment of cancers, especially in superficial solid tumors. For this purpose, a versatile theranostic nanoprobe of liposomal layer-coated Au nanocages (AuNCs) was decorated with an anti-MUC18 single-chain antibody (scFv). 4-mercapto benzoic acid (p-MBA)-labeled AuNCs (p-AuNCs) were coated by a liposomal layer (p-AuNCs@lip), followed by conjugating anti-MUC18 scFv via post-insertion method to form immuno-liposomal layer-coated AuNCs (p-AuNCs@scFv-lip). Physicochemical characterizations of the p-AuNCs@scFv-lip were investigated by transmission electron microscopy (TEM) and UV-vis and Raman spectroscopy. Furthermore, the targeting ability and theranostic efficiency of the nanoprobe were evaluated for specific diagnosis and treatment of cancerous melanoma cells by flow cytometry, SERS mapping, and live/dead assay. The formation of lipid layer on p-AuNCs surface was confirmed by TEM imaging. After decorating the liposomal layer with scFv, a relevant red shift was observed in the UV-vis spectrum. Moreover, p-AuNCs@lip presented characteristic peaks in the Raman spectrum, which exhibited only a minor change after scFv conjugation (p-AuNCs@scFv-lip). Interestingly, the cellular uptake of AuNCs@scFv-lip by A375 cell line (MUC18+) showed a 24-fold enhancement compared with SKBR3 cells (MUC18-). AuNCs@scFv-lip specifically identified A375 cells from SKBR cells via SERS mapping and effectively killed A375 cells through the PTT mechanism. Taken together, this theranostic platform can provide a promising tool for both in situ diagnosis and remote-controlled thermal ablation of cancer cells.

Redox-sensitive, PEG-shielded carboxymethyl PEI nanogels silencing MicroRNA-21, sensitizes resistant ovarian cancer cells to cisplatin.

A series of branched polyethylenimine (PEI) modifications including PEGylation (PEG2k-PEI) for steric shielding, redox-sensitive crosslinking for synthesis PEG2k-PEI-ss nanogels and subsequent carboxymethylation (PEG2k-CMPEI-ss) for modulation of the polymer pka have been introduced for cellular delivery of Anti-miR-21. The synthesis was characterized using 1H NMR, FTIR, TNBS, potentiometric titration, particle size and ζ potential. Loading of Anti-miR-21 at various N/P ratios was investigated by gel retardation, ethidium bromide dye exclusion, heparin sulfate competition and DNase I digestion experiments. The miR-21 silencing was measured by stem-loop RT PCR in A2780 ovarian cancer cell lines whether it is sensitive to resistant to cisplatin. It has been shown that PEG2k-CMPEI-ss was well suited for delivery of Anti-miR-21 in terms of nucleic acid loading, preservation against extracellular matrix and nucleases and sequence-specific silencing of miRNA-21 in vitro. Moreover, it has been demonstrated that pre-treating cells with Anti-miR-21 loaded nanogels can sensitize them to cis-Pt even at non-toxic concentraions. The results indicate that PEG2k-CMPEI-ss mediated microRNA delivery can be considered as a novel strategy for ovarian cancer therapy.

Progresses in microRNA Delivery Using Synthetic Nanovectors in Cancer Therapy.

MicroRNAs are small noncoding RNAs with key roles in gene expression. It has been revealed that aberrant expression of microRNAs is related to gene expression abnormality, and they have the potential to be used as anti-cancer drugs. However, the delivery of microRNAs is limited due to barriers, such as low uptake and insufficient endosomal release, intracellular nucleases degradation, phagocytic elimination, and renal filtration. To overcome these issues, novel delivery systems are developed for improving the efficiency of microRNAs therapy ranging from viral to synthetic; some are further developed with targeted ligands for active targeting purposes. Such delivery systems provide efficient cellular uptake and endosomal release as well as low cytotoxicity and minimum unwanted host immune response. Nevertheless, more complementary studies are warranted before being applied in human studies. This review deals with recent updates on the challenges and achievements of the various nanotechnology-based gene delivery vehicles with a special emphasis on the miRNA delivery in cancer therapy. In addition, we attempted to categorize the designed delivery systems based on miRNA therapeutic molecule. The related cellular signaling pathways and pharmacological action against cancer promotion have also been highlighted.

Synthesis of highly stable and biocompatible gold nanoparticles for use as a new X-ray contrast agent.

This work reports a novel reduction procedure for the synthesis of Gum Arabic (GA) capped-gold nanoparticles (AuNPs) in glucosammonium formate as a new ionic liquid. The GA coated AuNPs show good stability in physiological media. The synthesized AuNPs were characterized by UV-Vis spectroscopy, transmission electron microscopy, dynamic light scattering and X-ray diffraction analysis. These stable AuNPs are introduced as a new contrast agent for X-ray Computed Tomography (X-ray CT). These nanoparticles have higher contrasting properties than the commercial contrast agent, Visipaque. The precursors used (Gum Arabic and glucose based-ionic liquid) for synthesis of AuNPs are biocompatible and non-toxic.

miR-21, An Oncogenic Target miRNA for Cancer Therapy: Molecular Mechanisms and Recent Advancements in Chemo and Radio-resistance.

In the past decade, miRNAs have been extensively attracted the scientist's attentions as tumor suppressors or oncogenes that have been implicated in tumor progression, metastasis and intrinsic resistance to various cancer therapies. microRNA-21 (miR-21) demonstrates a potential oncogenic function and targets tumor inhibitor proteins in almost all types of cancer. miR-21 overexpression has been studied in terms of cell proliferation, migration, invasion, metastasis, and apoptosis regulation. Inhibition of miRNA expression using antisense technology by various nanovectors of different sizes, shapes and compositions has been evolved progressively to overcome the barriers confronted by miRNA delivery. Application of miR-21 antisense oligonucleotides for treating cancerous cells has become a promising achievement for cancer therapy. Moreover, miR-21 can mediate resistance to radiation and chemotherapy. The expanding role of miR-21 functions in human cancers with an emphasis on its regulatory targets and mechanisms, miR-21 related achievements against cancer promotion have been discussed.

Fluorometric study of fluoxetine DNA binding.

Fluoxetine (FLX), a selective serotonin reuptake inhibitor (SSRI), is commonly prescribed to treat depression. The interaction between FLX antidepressant and calf thymus DNA (ctDNA) was investigated under simulated physiological conditions (Tris-HCl buffer at pH 7.4) using methylene blue [3,7-bis(dimethylamino) pheno-thrazin-5-ium chloride] (MB) dye as a probe using fluorescence spectroscopy. A strong fluorescence quenching reaction of DNA to FLX was observed. The corresponding numbers of binding sites (n) and binding constants (K(f)) of DNA with FLX at 281, 310 and 318K were calculated 2.1×10(5), 6.7×10(5) and 9.7×10(5) respectively. It can be concluded that FLX molecules could interact with ctDNA via outside, non-intercalative, binding as evidenced by quenching study with I(-), ionic strength with NaCl and competitive investigation with MB. Thermodynamic parameters, enthalpy (ΔH) and entropy (ΔS) changes were calculated according to Van't Hoff equation, which indicated that reaction is entropically driven. Furthermore, the interaction of FLX with poly A-T and poly C-G were carried out in order to comprehend the binding location of drug to DNA.

DNA-binding studies of fluoxetine antidepressant.

Fluoxetine is a selective serotonin reuptake inhibitor (SSRI) antidepressant that is widely prescribed. The DNA-binding behavior of fluoxetine antidepressant and calf thymus DNA was investigated in Tris-HCl buffer at physiological pH 7.4 with a series of techniques, including UV-Vis and circular dichroism spectroscopies, competitive study with Hoechst 33258, viscometry, and cyclic voltammetry. Fluoxetine molecules bind to DNA via groove mode as illustrated by hypochromism with no red shift in the UV absorption band of fluoxetine, decrease in Hoechst-DNA solution fluorescence, and no significant changes in viscosity of DNA. The CD spectra of DNA molecules show a little change in stacking mode of base pair but no modification changes in DNA conformation, for example, from B-DNA to A or C-DNA. The binding constant (K(b)) of DNA with fluoxetine was calculated to be 6.7 × 10(4) M(-1), which is in the range of reported and known groove binders, such as distamycin. All results showed the groove-binding mode of interaction of fluoxetine with DNA.