Targeted biocompatible Zn-metal-organic framework nanocomposites for intelligent chemotherapy of breast cancer cells.

Finding a novel drug delivery system (DDS) represents one of the most challenging endeavors in cancer therapy. Hence, in this study, we developed a new biocompatible and biodegradable zinc-based nanoscale metal-organic framework (Zn-NMOF) coated with folic acid (FA) functionalized chitosan (CS) to facilitate targeted delivery of doxorubicin (D), a standard chemotherapeutic agent, into breast cancer cells. The synthesis of the NMOF-CS-FA-D nanocomposite preceded its comprehensive characterization via FT-IR, DLS, XRD, SEM, and TEM analyses. Subsequent in vitro studies were conducted on MCF-7 breast cancer cells and HFF-1 normal cells, encompassing assessments of cell viability, expression levels of apoptotic and autophagy genes, cell cycle arrest, and apoptotic analyses. The size of the NMOF-CS-FA-D particles was determined to be less than 80 nm, with a drug loading efficiency of 72 ± 5%. The release kinetics of DOX from the nanocomposite were investigated, revealing controlled release behavior at pH 7.4 and accelerated release at pH 5.0, which is conducive to drug delivery into cancer cells. In vitro results indicated a 17.39% ± 6.34 cell viability after 24 h of treatment with a 40 nM concentration of the NMOF-CS-FA-D nanocomposite. Furthermore, the expression levels of Caspase-9 and BAX, key apoptotic genes, along with BECLIN1, an autophagy gene, were found to increase by two-fold, four-fold, and two-fold, respectively, following 5 h of treatment with the nanocomposite. Additionally, analysis of cell cycle distribution revealed 15.4 ± 2% of cells in the sub-G1 phase, indicative of apoptotic cells, and 31.9% of cells undergoing early and late apoptosis in MCF-7 cells. Collectively, these findings underscore the potential of the NMOF-CS-FA-D nanocomposite in inhibiting cancer cell proliferation with low side effects.

Bifunctional folic acid targeted biopolymer Ag@NMOF nanocomposite [{Zn2 (1,4-bdc) 2 (DABCO)} n] as a novel theranostic agent for molecular imaging of colon cancer by SERS.

Without a doubt, cancer and its negative impact on human health have created many hurdles for people across the world since conventional approaches have not offered a reliable ability in the eradication of cancer. As a result, finding novel approaches, like using bimodal nanoparticles as a potential nanocarrier in molecular imaging and cancer therapy, is remarkably required these days. In the present study, ex-situ (Ge) and in-situ (Gi) green synthesized silver (Ag) nanoparticles entrapped in metal-organic framework nanocomposites (NMOF) coated with folic acid (FA) targeted chitosan (CS) was successfully developed as a novel bifunctional nanocarrier for detection and treatment of colon cancer cells. Then nanocarriers, such as NMOF-CS-FA, Ge-Ag@NMOF-CS-FA, Gi-Ag@NMOF-CS-FA, and C-Ag@NMOF-CS-FA, were characterized via FT-IR, DLS, SERS, TEM, and SEM and results have potentially confirmed the quality and quantity of synthesized nanocomposites. The hydrodynamic diameters of NMOF-CS, Ge-Ag@NMOF-CS, Gi-Ag@NMOF-CS, and C-Ag@NMOF-CS specimens were measured at around 99.7 ± 10 nm, 110 ± 10 nm, 118 ± 10 nm, 115 ± 10 nm, respectively. Also, the PDI values less than 0.2 confirm the reliable distribution of these nanocomposites. Afterward, the cell viability assay was conducted on HCT116 and HGF cell lines for evaluating biocompatibility and targeting efficiency of nanocomposites; FA functionalized nanocomposites have intensively indicated better performance in cancer cells targeting and their inhibition, and IC50 was attained for 10 ng/mL of Ge-Ag@NMOF-CS-FA while non-targeted nanocarriers did not have toxicity more than 20 % on HCT116 colon cancer cells. Moreover, according to the results, the cell viability of HGF normal cells was at least 85 % after being exposed to different concentrations of nanocomposites for 24 h. This indicates that the synthesized nanocomposites do not have significant toxic effects on normal cells. The results indicate that this novel nanocomposite has the potential to effectively deliver drugs to cancer cells.

Chitosan-Albumin Nanocomposite as a Promising Nanocarrier for Efficient Delivery of Fluconazole Against Vaginal Candidiasis.

Currently, the high incidence of fungal infections among females has resulted in outstanding problems. Candida species is related with multidrug resistance and destitute clinical consequences. Chitosan-albumin derivatives with more stability exhibit innate antifungal and antibacterial effects that boost the activity of the drug without inflammatory impact. The stability and sustained release of Fluconazole in mucosal tissues can be ensured by encapsulating in protein/polysaccharide nanocomposites. Thus, we developed chitosan-albumin nanocomposite (CS-A) loaded with Fluconazole (Flu) antifungals against vaginal candidiasis. Various ratios of CS/Flu (1:1, 1:2, 2:1) were prepared. Thereafter, the CS-A-Flu nanocomposites were qualified and quantified using FT-IR, DLS, TEM, and SEM analytical devices, and the size range from 60 to 100 nm in diameter was attained for the synthesized nanocarriers. Afterward, the antifungal activity, biofilm reduction potency, and cell viability assay were performed for biomedical evaluation of formulations. The minimum inhibitory concentration) and minimum fungicidal concentration on Candida albicans were attained at 125 ng/µL and 150 ng/µL after treatment with a 1:2 (CS/Flu) ratio of CS-A-Flu. The biofilm reduction assay indicated that biofilm formation was between 0.05 and 0.1% for CS-A-Flu at all ratios. The MTT assay also exhibited excellent biocompatibility for samples, about 7 to 14% toxicity on human HGF normal cells. These data have indicated that CS-A-Flu would be a promising candidate against Candida albicans.

Novel Chimeric Endolysin Conjugated Chitosan Nanocomplex as a Potential Inhibitor Against Gram-Positive and Gram-Negative Bacteria.

Resistance to antimicrobial agents has created potential problems in finding efficient treatments against bacteria. Thus, using new therapeutics, such as recombinant chimeric endolysin, would be more beneficial for eliminating resistant bacteria. The treatment ability of these therapeutics can be further improved if they are used with biocompatible nanoparticles like chitosan (CS). In this work, covalently conjugated chimeric endolysin to CS nanoparticles (C) and non-covalently entrapped endolysin in CS nanoparticles (NC) were effectively developed and, consequently, qualified and quantified using analytical devices, including FT-IR, dynamic light scattering, and TEM. Eighty to 150 nm and 100 nm to 200 nm in diameter were measured for CS-endolysin (NC) and CS-endolysin (C) using a TEM, respectively. The lytic activity, synergistic interaction, and biofilm reduction potency of nano-complexes were investigated on Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and Pseudomonas aeruginosa (P. aeruginosa) strains. The outputs revealed a good lytic activity of nano-complexes after 24 h and 48 h of treatment, especially in P. aeruginosa (approximately 40% cell viability after 48 h of treatment with 8 ng/mL), and potential biofilm reduction performance was attained in E. coli strains (about 70% reduction after treatment with 8 ng/mL). The synergistic interaction between nano-complexes and vancomycin was exhibited in E. coli, P. aeruginosa, and S. aureus strains at 8 ng/mL concentrations, while the synergistic effects of pure endolysin and vancomycin were not remarkable in E. coli strains. These nano-complexes would be more beneficial in suppressing the bacteria with a high level of antibiotic resistance.

Development of Novel Chimeric Endolysin Conjugated with Chitosan-Zn-Metal-Organic Framework Nanocomposites with Antibacterial Activity.

Bacterial diseases have been considered the most crucial issue and are threatening human health all around the world. Also, resistance to antimicrobial drugs has become a big hurdle against efficient therapy. As a result, recombinant chimeric endolysin was produced in E. coli host to use as a potential antibacterial agent against bacteria resistance and replacement to conventional antibiotics in this study. Then, chitosan (C)-coated nanoscale metal-organic frameworks (CS-NMOFs) nanocomposite was synthesized as a novel nano delivery system to further improve the antibacterial activity of endolysin. After characterization of nanocomposite with analytical devices such as FT-IR, DLS, and TEM and determining the nanometric size of samples (30 nm to 90 nm), endolysin was covalently (endolysin-CS-NMOFs (C)) and non-covalently (endolysin-CS-NMOFs (NC)) conjugated to nanocomposite. Thereafter, the lytic ability, synergistic interaction, and biofilm reduction manner of endolysin-containing CS-NMOF nanocomposites were evaluated on E. coli, S. aureus, and P. aeruginosa strains. The results depicted an excellent lytic ability of nanocomposites after 24 h and 48 h of treatment, especially endolysin-CS-NMOFs (NC) on E. coli and P. aeruginosa strains. The synergistic interaction between nanocomposite and vancomycin did not attain for P. aeruginosa strain whereas the reverse was true for E. coli and S. aureus strains at 8 ng/mL concentration. Next, nanocomposites demonstrated potential biofilm reduction activities in various strains, especially in S. aureus and P. aeruginosa. Ultimately, our outputs demonstrate an efficient performance of the synthesized nanocomposite as an appropriate substitution for conventional antibiotics against bacteria.

Targeted PEGylated Chitosan Nano-complex for Delivery of Sodium Butyrate to Prostate Cancer: An In Vitro Study.

Introduction: Cancer remains a challenging issue against human health throughout the world; As a result, introducing novel approaches would be beneficial for cancer treatment. In this research, sodium butyrate (Sb) is one of the effective anti-cancer therapeutics (also a potent survival factor for normal cells) that was used for prostate cancer suppression in the platform of modified chitosan (CS) nano-complex (polyethylene glycol (PEG)-folic acid (FA)-Sb-CS). Methods: Different analytical devices including Fourier transform infrared, dynamic light scattering, high-performance liquid chromatography, scanning electron microscopy, and transmission electron microscopy were applied for the characterization of synthetics. On the other hand, biomedical tests including cell viability assay, molecular and functional assay of apoptosis/autophagy pathways, and cell cycle arrest analysis were potentially implemented on human PC3 (folate receptor-negative prostate cancer) and DU145 (folate receptor-positive prostate cancer) and HFF-1 normal cell lines. Results: The quality of the syntheses was effectively verified, and the size range from 140 to 170 nm was determined for the PEG-CS-FA-Sb sample. Also, 75 ± 5% of drug entrapment efficiency with controlled drug release manner (Sb release of 54.21% and 74.04% for pHs 7.4 and 5.0) were determined for nano-complex. Based on MTT results, PEG-CS-FA-Sb has indicated 72.07% and 33.53% cell viability after 24 h of treatment with 9 mM on PC3 and DU145 cell lines, respectively, which is desirable anti-cancer performance. The apoptotic and autophagy genes overexpression was 15-fold (caspase9), 2.5-fold (BAX), 11-fold (ATG5), 2-fold (BECLIN1), and 3-fold (mTORC1) genes in DU145 cancer cells. More than 50% of cell cycle arrest and 45.05% of apoptosis were obtained for DU145 cancer cells after treatment with nano-complex. Conclusion: Hence, the synthesized Sb-loaded nano-complex could specifically suppress prostate cancer cell growth and induce apoptosis and autophagy in the molecular and cellular phases.

Margetuximab conjugated-PEG-PAMAM G4 nano-complex: a smart nano-device for suppression of breast cancer.

Abstract: Breast cancer due to its high incidence and mortality is the second leading cause of death among females. On the other hand, nanoparticle-based drug delivery is one of the most promising approaches in cancer therapy, nowadays. Hence, margetuximab- and polyethylene glycol-conjugated PAMAM G4 dendrimers were efficiently synthesized for targeted delivery of quercetin (therapeutic agent) to MDA-MB-231 breast cancer cells. Synthesized nano-complexes were characterized using analytical devices such as FT-IR, TGA, DLS, Zeta potential analyzer, and TEM. The size less than 40 nm, - 18.8 mV surface charge, efficient drug loading capacity (21.48%), and controlled drug release (about 45% of drug release normal pH after 8 h) were determined for the nano-complex. In the biomedical test, the cell viability was obtained 14.67% at 24 h of post-treatment for 800 nM concentration, and IC50 was ascertained at 100 nM for the nano-complex. The expression of apoptotic Bax and Caspase9 genes was increased by more than eightfolds and more than fivefolds after treatment with an optimal concentration of nanocarrier. Also, more than threefolds of cell cycle arrest was observed at the optimal concentration synthetics, and 27.5% breast cancer cell apoptosis was detected after treatment with 100 nM nano-complex. These outputs have been indicating the potential capacity of synthesized nano-complex in inhibiting the growth of breast cancer cells.

Chitosan-coated Zn-metal-organic framework nanocomposites for effective targeted delivery of LNA-antisense miR-224 to colon tumor: in vitro studies.

Nowadays, nano-compartments are considered as an effective drug delivery system (DDS) for cancer therapy. Targeted delivery of therapeutic agents is an advantageous approach by which cancer cells can be targeted without harming normal cells, and eliminates the negative effects of conventional therapies such as chemotherapy. In this research, a novel zinc-based nanoscale metal-organic framework (Zn-NMOF) coated with folic acid (FA) functionalized chitosan (CS) has been constructed and applied as efficient delivery of LNA (locked nucleic acid)-antisense miR-224 to colon cancer cell lines. LNA-antisense miR-224 as a therapeutic sequence was able to considerably block highly expressed miR-224 and downregulated cancer cell growth. The prepared nano-complex was characterized by analytical devices such as FT-IR, UV-Vis spectrophotometry, DLS, TEM, and XRD. The size range of NMOF-CS-FA-LNA-antisense miR-224 (MCFL224) nano-complex was obtained nearly at 200 nm. The entrapment efficiency of LNA-antisense miR-224 was calculated 72 ± 5% and a significant release profile of LNA-antisense miR-224 was observed at first 6 h (about 50%). Then, in vitro assays were implemented on HCT116 (folic acid receptor-positive colon cancer cell line) and CRL1831 (normal colon cell line) to evaluate the therapeutic efficiency of the MCFL224 nano-complex. In these investigations, decreased cell viability (14.22 ± 0.3% after 72 h treatment), increased apoptotic and autophagy-related genes expression level (BECLIN1: 34-folds, BAX: 36-folds, mTORC1: 10-folds, and Caspase-9: 9-folds more than control), higher cell cycle arrest in sub-G1 phase (19.53% of cells in sub-G1 phase), and more apoptosis analyses (late apoptosis: 67.7%) were evaluated in colon cancer cells treated with MCFL224 nano-complex. Results remarkably indicate the inhibited growth of colon cancer cells and induced cell apoptosis which suggests MCFL224 as a promising nanocomposite for colon cancer therapy.

Anti-HER2 VHH Targeted Fluorescent Liposome as Bimodal Nanoparticle for Drug Delivery and Optical Imaging.

OBJECTIVES: The aim of this study was to formulate fluorescent-labeled targeted immunoliposome to visualize the delivery and distribution of drugs in real-time. METHODS: In this study, fluorescent-labeled liposomes were decorated with anti-HER2 VHH or Herceptin to improve the monitoring of intracellular drug delivery and tumor cell tracking with minimal side effects. The conjugation efficiency of antibodies was analyzed by SDS-PAGE silver staining. In addition, the physicochemical characterization of liposomes was performed using DLS and TEM. Finally, confocal microscopy visualized nanoparticles in the target cells. RESULTS: Quantitative and qualitative methods characterized the intracellular uptake of 110±10 nm particles with near 70% conjugation efficiency. In addition, live-cell trafficking during hours of incubation was monitored by wide-field microscopy imaging. The results show that the fluorescent- labeled nanoparticles can specifically bind to HER2-positive breast cancer with minimal off-target delivery. CONCLUSION: These nanoparticles can have several applications in personalized medicine, especially drug delivery and real-time visualization of cancer therapy. Moreover, this method also can be applied in the targeted delivery of contrast agents in imaging and thermotherapy.

Design and synthesis of novel 4-thiazolidinone derivatives with promising anti-breast cancer activity: Synthesis, characterization, in vitro and in vivo results.

Novel lead compounds as anticancer agents with the ability to circumvent emerging drug resistance have recently gained a great deal of interest. Thiazolidinones are among such compounds with well-established biological activity in the field of oncology. Here, we designed, synthesized and characterized a series of thiazolidinone structures (8a-8k). The results of anti-proliferative assay led to the discovery of compound 8j with a high potent cytotoxic effect using colon, liver and breast cancer cells. Furthermore, MDA-MB-231 and 4T1 cell lines were used to represent triple negative breast cancer (TNBC). Next, a number of in vitro and in vivo evaluations were carried out to demonstrate the potential activity against TNBC and also elucidate the possible mechanism of cell death induction. Our in vitro outcomes exhibited an impressive anticancer activity for compound 8j toward MDA-MB-231 cells through inducing apoptosis and a remarkable anti-metastatic feature via suppressing MMP-9 expression as well. Consistently, the in vivo and immunohistopathologic evaluations demonstrated that this compound significantly inhibited the 4T1 induced tumor growth and its metastasis to the lung. Altogether, among numerous thiazolidinone derivatives, compound 8j might represent a promising anticancer agent for TNBC, which is a major concern in the developed and developing countries.

Liposomal nanoparticle armed with bivalent bispecific single-domain antibodies, novel weapon in HER2 positive cancerous cell lines targeting.

Breast cancer is the leading cause of mortality among all cancers. HER2, human epidermal growth factor receptors type 2, a receptor tyrosine kinase that induces interminable cell proliferation, is overexpressed in 20-25 percent of breast cancers. In spite of significant progress in nanomedicine in the past decade, being subjected to genetic drift that hides many paramount epitopes has rendered targeting HER2 as a big challenge. In the present study, we developed monovalent and bivalent monospecific along with bivalent bispecific VHH targeting different epitopes on HER2, and showed that bivalent bispecific VHH has the highest affinity among other tested modalities. Then we covalently coupled VHHs to the fluorescent labeled liposomal nanoparticle to produce targeted liposomes. Based on flow cytometry results, bivalent bispecific VHH targeted liposomes showed the highest fluorescent intensity, on HER2 breast cancer cells. Liposomes conjugated to bivalent monospecific VHH exhibited enhanced affinity toward HER2 positive cell lines compared to monovalent targeted liposomes, with bivalent bispecific liposomes appearing as the most robust probe.

Anti-HER2 VHH Targeted Magnetoliposome for Intelligent Magnetic Resonance Imaging of Breast Cancer Cells.

The combination of liposomes with magnetic nanoparticles, because of their strong effect on T2 relaxation can open new ways in the innovative cancer therapy and diagnosis. In order to design an intelligent contrast agent in MRI, we chose anti-HER2 nanobody the smallest fully functional antigen-binding fragments evolved from the variable domain, the VHH, of a camel heavy chain-only antibody. These targeted magnetoliposomes bind to the HER2 antigen which is highly expressed on breast and ovarian cancer cells so reducing the side effects as well as increasing image contrast and effectiveness. Cellular iron uptake analysis and in vitro MRI of HER2 positive cells incubated with targeted nanoparticles show specific cell targeting. In vitro MRI shows even at the lowest density (200 Cells/µl), dark spots corresponding to labeled cells which were still detectable. These results suggest that this new type of nanoparticles could be effective antigen-targeted contrast agents for molecular imaging.

The effect of superparamagnetic iron oxide nanoparticles surface engineering on relaxivity of magnetoliposome.

The purpose of this work is evaluating the effect of ultra small superparamagnetic iron oxide nanoparticles (USPIONs) coatings on encapsulation efficiency in liposomes and cellular cytotoxicity assay. Moreover, we assessed the effects of surface engineering on the relaxivity of magnetoliposome nanoparticles in order to create a targeted reagent for the intelligent diagnosis of cancers by MRI. For estimating the effect of nanoparticle coatings on encapsulation, several kinds of USPIONs coated by dextran, PEG5000 and citrate were used. All kinds of samples are monodispersed and below 100 ± 10 nm and the coatings of USPIONs have no significant effect on magnetoliposome diameter. The coating of USPIONs could have effect on percentage of encapsulation. The dextran coated USPIONs have more stability and quality accordingly the encapsulation increased up to 92%, then the magnetoliposome nano particles have been targeted by Herceptin and anti-HER2 VHH, separately. Over storage period of four weeks the resulting particles were stable and physico-chemical properties such as size and zetapotential did not show any significant changes. The relaxivity of contrast agents was measured using a 1.5 T MRI. The r2/r1 ratio was more than two for all samples which demonstrate the negative contrast enhancing of all SPION embedded specimens. The high ratio of r2/r1 as well as high r2 is the best combination of a negative contrast agent as it is obtained for pure magnetite. The value of r2/r1 for all other samples including Herceptin targeted magnetoliposome, anti-HER2 VHH targeted magnetoliposome and non-targeted magnetoliposome were between ~21 to ~28, which show the magnetite embedded samples have enough negative contrast to be detectable by MRI. Therefore the HER2 targeted magnetoliposomes are a good and stable candidate as contrast agents in clinical radiology and biomedical research with minimal cytotoxicity and biocompatibility effects. Copyright © 2016 John Wiley & Sons, Ltd.

A caspase 8-based suicide switch induces apoptosis in nanobody-directed chimeric receptor expressing T cells.

In accordance with the two-step hypothesis of T cell activation and the observation that stimulation through the T cell receptor (TCR) alone may lead to anergy, we focused on the introduction of co-stimulatory signaling to this type of receptors to achieve optimal activation. Enhanced mRNA and cell surface receptor expression via the co-stimulatory gene fragment (OX40) was confirmed by RT-PCR and flow cytometry. Inclusion of the OX40 co-stimulatory signaling region in series with the TCR led to enhanced antigen-induced IL-2 production after stimulation by MUC1-expressing cancer cell lines as compared to the chimeric receptor without OX40. Moreover, with the aim of maintaining high efficiency, while providing a means of controlling any possible unwanted proliferation in vivo, a regulation system was used. This controls the dimerization of a membrane-bound caspase 8 protein. Toward that goal, pFKC8 and CAR constructs were co-transfected into Jurkat cells, and the level of apoptosis was measured. 24 h after addition of the dimerizer, a 91% decrease in transfected cells was observed.