Titania-Graphene Oxide Nanocomposite-Based Philadelphia-Positive Leukemia Therapy.

Philadelphia-positive (Ph+) leukemia is a type of blood cancer also known as acute lymphoblastic leukemia (ALL), affecting 20-30% of adults diagnosed worldwide and having an engraved prognosis as compared to other types of leukemia. The current treatment regimens mainly rely on tyrosine kinase inhibitors (TKIs) and bone marrow transplants. To date, several generations of TKIs have been developed due to associated resistance and frequent relapse, with cardiovascular system anomalies being the most devastating complication. Nanotechnology has the potential to address these limitations by the targeted drug delivery and controlled release of TKIs. This study focused on the titanium dioxide (TiO2) and graphene oxide (GO) nanocomposite employment to load nilotinib and ponatinib TKIs for therapy of Ph+ leukemia cell line (K562) and Ba/F3 cells engineered to express BCR-ABL oncogene. Meanwhile, after treatment, the oncogene expressing fibroblast cells (Rat-1 P185) were evaluated for their colony formation ability under 3D conditions. To validate the nanocomposite formation, the TiO2-GO nanocomposites were characterized by scanning electron microscope, DLS, XRD, FTIR, zeta potential, EDX, and element mapping. The TKI-loaded TiO2-GO was not inferior to the free drugs after evaluating their effects by a cell viability assay (XTT), apoptosis induction, and colony formation inhibition. The cell signaling pathways of the mammalian target of rapamycin (mTOR), signal transducers and activators of transcription 5 (STAT5), and extracellular signal-regulated kinase (Erk1/2) were also investigated by Western blot. These signaling pathways were significantly downregulated in the TKI-loaded TiO2-GO-treated groups. Based on the findings above, we can conclude that TiO2-GO exhibited excellent drug delivery potential that can be used for Ph+ leukemia therapy in the future, subject to further investigations.

Lyophilization Based Isolation of Exosomes.

Exosomes are nanoscale extracellular vesicles which regulate intercellular communication. They have great potential for application in nanomedicine. However, techniques for their isolation are limited by requirements for advanced instruments and costly reagents. In this study, we developed a lyophilization-based method for isolating exosomes from cultured cells. The isolated exosomes were characterized for protein content using Bradford assay, and for size distribution and shape using scanning electron microscopy (SEM) and nanoparticles tracking analysis (NTA). In addition, CD63, CD9, CD81, HSP70 and TSG101 were evaluated as essential exosomal surface markers using Western blot. Drug loading and release studies were performed to confirm their drug delivery properties using an in vitro model. Exosomes were also loaded with commercial dyes (Cy5, Eosin) for the evaluation of their drug delivery properties. All these characterizations confirmed successful exosome isolation with measurements of less than 150 nm, having a typical shape, and by expressing the known exosome surface protein markers. Finally, tyrosine kinase inhibitors (dasatinib and ponatinib) were loaded on the exosomes to evaluate their anticancer effects on leukemia cells (K562 and engineered Ba/F3-BCR-ABL) using MTT and Annexin-PI assays. The expression of MUC1 protein on the exosomes isolated from MCF-7 cells also indicated that their potential diagnostic properties were intact. In conclusion, we developed a new method for exosome isolation from cultured cells. These exosomes met all the essential requirements in terms of characterization, drug loading and release ability, and inhibition of proliferation and apoptosis induction in Ph+ leukemia cells. Based on these results, we are confident in presenting the lyophilization-based exosome isolation method as an alternative to traditional techniques for exosome isolation from cultured cells.

Exosomes based strategies for brain drug delivery.

Exosome application has emerged as a promising nanotechnology discipline for various diseases therapeutics and diagnoses. Owing to the natural properties of efficient drug delivery, higher biocompatibility, facile traversing of physiological barriers, and subtle side effects, exosomes shorten their way to clinical translation. Exosomes are nanoscale membrane-bound vesicles primarily involved in intercellular communication and exhibit natural blood-brain barrier (BBB) traversing ability, which enables their application as drug delivery vehicles for brain diseases treatment. Herein, we highlight recent exosome-based drug delivery endeavors for neurodegenerative diseases and brain cancer therapy, summarize the obstacles and future directions in clinical translation.

Biomimetic Dp44mT-nanoparticles selectively induce apoptosis in Cu-loaded glioblastoma resulting in potent growth inhibition.

Selective targeting of elevated copper (Cu) in cancer cells by chelators to induce tumor-toxic reactive oxygen species (ROS) may be a promising approach in the treatment of glioblastoma multiforme (GBM). Previously, the Cu chelator di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT) attracted much interest due to its potent anti-tumor activity mediated by the formation of a highly redox-active Cu-Dp44mT complex. However, its translational potential was limited by the development of toxicity in murine models of cancer reflecting poor selectivity. Here, we overcame the limitations of Dp44mT by incorporating it in new biomimetic nanoparticles (NPs) optimized for GBM therapy. Biomimetic design elements enhancing selectivity included angiopeptide-2 functionalized red blood cell membrane (Ang-M) camouflaging of the NPs carrier. Co-loading Dp44mT with regadenoson (Reg), that transiently opens the blood-brain-barrier (BBB), yielded biomimetic Ang-MNPs@(Dp44mT/Reg) NPs that actively targeted and traversed the BBB delivering Dp44mT specifically to GBM cells. To further improve selectivity, we innovatively pre-loaded GBM tumors with Cu. Oral dosing of U87MG-Luc tumor bearing mice with diacetyl-bis(4-methylthiosemicarbazonato)-copperII (Cu(II)-ATSM), significantly enhanced Cu-level in GBM tumor. Subsequent treatment of mice bearing Cu-enriched orthotopic U87MG-Luc GBM with Ang-MNPs@(Dp44mT/Reg) substantially prevented orthotopic GBM growth and led to maximal increases in median survival time. These results highlighted the importance of both angiopeptide-2 functionalization and tumor Cu-loading required for greater selective cytotoxicity. Targeting Ang-MNPs@(Dp44mT/Reg) NPs also down-regulated antiapoptotic Bcl-2, but up-regulated pro-apoptotic Bax and cleaved-caspase-3, demonstrating the involvement of the apoptotic pathway in GBM suppression. Notably, Ang-MNPs@(Dp44mT/Reg) showed negligible systemic drug toxicity in mice, further indicating therapeutic potential that could be adapted for other central nervous system disorders.

Heme Oxygenase-1 targeting exosomes for temozolomide resistant glioblastoma synergistic therapy.

Glioblastoma (GBM) is a highly fatal and recurrent brain cancer without a complete prevailing remedy. Although the synthetic nanotechnology-based approaches exhibit excellent therapeutic potential, the associated cytotoxic effects and organ clearance failure rest major obstacles from bench to clinics. Here, we explored allogeneic bone marrow mesenchymal stem cells isolated exosomes (BMSCExo) decorated with heme oxygenase-1 (HMOX1) specific short peptide (HSSP) as temozolomide (TMZ) and small interfering RNA (siRNA) nanocarrier for TMZ resistant glioblastoma therapy. The BMSCExo had excellent TMZ and siRNA loading ability and could traverse the blood-brain barrier (BBB) by leveraging its intrinsic brain accumulation property. Notably, with HSSP decoration, the TMZ or siRNA encapsulated BMSCExo exhibited excellent TMZ resistant GBM targeting ability both in vitro and in vivo due to the overexpression of HMOX1 in TMZ resistant GBM cells. Further, the HSSP decorated BMSCExo delivered the STAT3 targeted siRNA to the TMZ resistant glioma and restore the TMZ sensitivity, consequently achieved the synergistically drug resistant GBM treatment with TMZ. Our results showed this biomimetic nanoplatform can serve as a flexible, robust and inert system for GBM treatment, especially emphasizing the drug resistant challenge.

Brain-Targeted Codelivery of Bcl-2/Bcl-xl and Mcl-1 Inhibitors by Biomimetic Nanoparticles for Orthotopic Glioblastoma Therapy.

Glioblastoma (GBM) is among the most treatment-resistant solid tumors and often recurrs after resection. One of the mechanisms through which GBM escapes various treatment modalities is the overexpression of anti-apoptotic Bcl-2 family proteins (e.g., Bcl-2, Bcl-xl, and Mcl-1) in tumor cells. Small-molecule inhibitors such as ABT-263 (ABT), which can promote mitochondrial-mediated cell apoptosis by selectively inhibiting the function of Bcl-2 and Bcl-xl, have been proven to be promising anticancer agents in clinical trials. However, the therapeutic prospects of ABT for GBM treatment are hampered by its limited blood-brain barrier (BBB) penetration, dose-dependent thrombocytopenia, and the drug resistance driven by Mcl-1, which is overexpressed in GBM cells and further upregulated upon treatment with ABT. Herein, we reported that the Mcl-1-specific inhibitor A-1210477 (A12) can act synergistically with ABT to induce potent cell apoptosis in U87 MG cells, drug-resistant U251 cells, and patient-derived GBM cancer stem cells. We further designed a biomimetic nanomedicine, based on the apolipoprotein E (ApoE) peptide-decorated red blood cell membrane and pH-sensitive dextran nanoparticles, for the brain-targeted delivery of ABT and A12. The synergistic anti-GBM effect was retained after encapsulation in the nanomedicine. Additionally, the obtained nanomedicine possessed good biocompatibility, exhibited efficient BBB penetration, and could effectively suppress tumor growth and prolong the survival time of mice bearing orthotopic GBM xenografts without inducing detectable adverse effects.

Correction: SERS-based nanostrategy for rapid anemia diagnosis.

Correction for 'SERS-based nanostrategy for rapid anemia diagnosis' by Pir Muhammad et al., Nanoscale, 2020, 12, 1948-1957, DOI: 10.1039/C9NR09152A.

Biocompatible exosomes nanodrug cargo for cancer cell bioimaging and drug delivery.

Therapy against cancer remains a daunting issue for human health, despite remarkable innovations in many areas of pathology. In situ biosynthesized nanoclusters bestow a novel remedy for carcinogenic cell imaging. Exosomes have received special attention as an efficient tool for the diagnosis of various diseases, including cancers. All types of cells (healthy or diseased) generate exosomes, making them significantly unique for relevant disease diagnosis and treatment. In this contribution, we exploit the possibility of utilizing the exosomes to facilitate chemotherapeutics, viz. the combination of doxorubicin (Dox) and biosynthesized silver nanoclusters in cancer cells. Our study showed a new facile way for bioimaging of cancer cells using biosynthesized silver-DNA nanoclusters, and thus further targeting cancer cells using the relevant cancer exosomes as drug delivery cargo. After isolating exosomes from neoplastic cells, i.e. HeLa, loaded with the drug, and treating other neoplastic cells with cargo-loaded isolated exosomes, we found that cargo-loaded isolated exosomes can readily enter into the targeted cancer cells and efficiently kill these neoplastic cells. This raises the possibility of acting as a novel facile modality for target cancer theranostics with high efficiency and biocompability.

Specific Oxide Nanoclusters Enhance Intracellular Reactive Oxygen Species for Cancer-Targeted Therapy.

Bio-nanotechnology based cancer therapeutics exponentially increase every year. A therapeutic strategy to induce intracellular reactive oxygen species (ROS) has received promising success in oncotherapy. In this study, the new strategy has been exploited by the treatment of iridium (Ir) and Fe2+ ions with cancer cells to biosynthesize the biocompatible fluorescent iridium oxide (IrO2) and iron oxide nanoclusters (NCs) under the specific redox heterogeneous microenvironment of these diseased cells and tumors. The hydroxyl radical produced by the presence of Fe2+ and H2O2 in cancer cells apparently increased the ROS level in cancer cells during the process of biosynthesized NCs and, hence, simultaneously instigated apoptosis of relevant cells. Therefore, intracellular ROS-mediated in situ biosynthesis of IrO2 and iron oxide NCs may also act as anticancer agents and provide a promising pathway for targeted cancer therapy.

Nanomedicine-based immunotherapy for central nervous system disorders.

Central nervous system (CNS) disorders represent a broad spectrum of brain ailments with short- and long-term disabilities, and nanomedicine-based approaches provide a new therapeutic approach to treating CNS disorders. A variety of potential drugs have been discovered to treat several neuronal disorders; however, their therapeutic success can be limited by the presence of the blood-brain barrier (BBB). Furthermore, unique immune functions within the CNS provide novel target mechanisms for the amelioration of CNS diseases. Recently, various therapeutic approaches have been applied to fight brain-related disorders, with moderate outcomes. Among the various therapeutic strategies, nanomedicine-based immunotherapeutic systems represent a new era that can deliver useful cargo with promising pharmacokinetics. These approaches exploit the molecular and cellular targeting of CNS disorders for enhanced safety, efficacy, and specificity. In this review, we focus on the efficacy of nanomedicines that utilize immunotherapy to combat CNS disorders. Furthermore, we detailed summarize nanomedicine-based pathways for CNS ailments that aim to deliver drugs across the BBB by mimicking innate immune actions. Overview of how nanomedicines can utilize multiple immunotherapy pathways to combat CNS disorders.

In situ self-assembled Ag-Fe3O4 nanoclusters in exosomes for cancer diagnosis.

Recently, exosomes have gained attention as an effective tool for early cancer detection. Almost all types of cells release exosomes, making them substantially important for disease diagnosis. In this study, we have utilized HepG2 cancer cells for the in situ biosynthesis of silver and iron oxide nanoclusters (NCs) from their respective salts (i.e., AgNO3 and FeCl2, respectively) in the presence of glutathione (GSH). The self-assembled biosynthesized silver and iron NCs were readily loaded on exosomes as payloads and secreted into the cell culture medium. The cargo loaded exosomes were then isolated and characterized by electron microscopy for nano-silver and iron oxide NC confirmation. Ag NCs have potential as a fluorescent probe and Fe3O4 NCs as a contrast agent for CT and MRI. Furthermore, these isolated exosomes from HepG2 cancer cells have a significant influence on cellular uptake and cell viability when exposed to both HepG2 and U87 cancer cells. These findings demonstrate that the biocompatible nature of these self-assembled NCs loaded on exosomes could be utilized to bioimage cancer in the initial stages through fluorescence imaging.

SERS-based nanostrategy for rapid anemia diagnosis.

Iron detection is one of the critical markers to diagnose multiple blood-related disorders that correspond to various biological dysfunctions. The currently available anemia detection approach can be used only for pre-treated blood samples that interfere with the actual iron level in blood. Real-time detection approaches with higher sensitivity and specificity are certainly needed to cope with the commercial level clinical analyses. Herein, we presented a novel strategy to determine the blood iron that can be easily practiced at commercial levels. The blend of well-known iron-cyanide chemistry with nanotechnology is advantageous with ultrahigh sensitivity in whole blood analysis without any pre-treatments. This approach is a combined detection system of the conventional assay (UV-visible spectroscopy) with surface-enhanced Raman scattering (SERS). Organic cyanide modified silver nanoparticles (cAgNPs) can selectively respond to Fe3+ ions and Hb protein with a detection limit of 10 fM and 0.46 µg mL-1, respectively, without being affected by matrix interfering species in the complex biological fluid. We confirmed the clinical potential of our new cAgNPs by assessing iron-status in multiple anemia patients and normal controls. Our SERS-based iron quantitation approach is highly affordable for bulk-samples, cheap, quick, flexible, and useful for real-time clinical assays. Such a method for metal-chelation has extendable features of therapeutics molecular tracking within more complex living systems at cellular levels.

Nano in nano: Biosynthesized gold and iron nanoclusters cargo neoplastic exosomes for cancer status biomarking.

Timely detection is crucial for successful treatment of cancer. The current study describes a new approach that involves utilization of the tumor cell environment for bioimaging with in-situ biosynthesized nanoscale gold and iron probes and subsequent dissemination of Au-Fe nanoclusters from cargo exosomes within the circulatory system. We have isolated the Au-Fe cargo exosomes from the blood of the treated murine models after in situ biosyntheses from their respective pre-ionic solutions (HAuCl4, FeCl2), whereas Na2SeO3 supplementation added into Au lethal effect. The microarray data of various differentially expressed genes revealed the up-regulated tumor ablation and metal binding genes in SGC-7901 cell lines after treatment with Au-Fe-Se triplet ionic solution. The isolation of Au-Fe nanoclusters cargo exosomes (nano in nano) after secretion from deeply seated tumors may help in early diagnosis and reveal the tumor ablation status during and after the relevant treatment like radio-chemo therapies et al.

Real-Time Multimodal Bioimaging of Cancer Cells and Exosomes through Biosynthesized Iridium and Iron Nanoclusters.

Multimodal bioimaging is a powerful tool for visualizing the abnormal state at the target site of the related disease. In this study, we used multimodal imaging techniques such as computed tomography, fluorescence, and magnetic resonance imaging to improve early and precise diagnosis of tumor. Herein, we reported the facile in situ biosynthesis of iridium and iron oxide nanoclusters (NCs) in cancer cells or tumor tissue. These NCs are used as a multimodal bioimaging probe to improve the image sensitivity and specificity toward the tumor. These NCs are applied for the in vivo multimodal imaging in the form of an imaging probe capable of enhancing the sensitivity of the image and specificity toward the tumor tissue. Our observation demonstrates that highly luminescent and magnetic NCs are not only biocompatible but also tumor-targeted because NC formation does not take place in normal cells and tissues. In addition, we isolated exosomes and the biosynthesized NCs internalized within exosomes, and these exosomes can be used as cancer biomarkers.

Mammalian cells: a unique scaffold for in situ biosynthesis of metallic nanomaterials and biomedical applications.

Production of nanoscale materials often requires the use of toxic chemicals and complex synthetic procedures. A new scaffold has been explored for in situ synthesis of nanomaterials that utilizes natural biological systems in the form of plants, bacteria, fungi, algae and redox-imbalanced mammalian cells and systems. The latter approach has become popular in recent years and has shown some promising results in bioimaging of cancer, as well as inflammatory and neurodegenerative maladies. Biosynthesis of nanoclusters in redox-imbalanced mammalian cells is facile, cost-effective and environmentally friendly with higher biocompatibility and target specificity and lower adverse effects than traditional synthetic approaches. Herein, we describe recent advances in mammalian green in situ biosynthesis for biomedical applications, especially in cancer and neurodegenerative disease theranostics.

In Vivo Biosynthesized Zinc and Iron Oxide Nanoclusters for High Spatiotemporal Dual-Modality Bioimaging of Alzheimer's Disease.

Alzheimer's disease is still incurable and neurodegenerative, and there is a lack of detection methods with high sensitivity and specificity. In this study, by taking different month old Alzheimer's mice as models, we have explored the possibility of the target bioimaging of diseased sites through the initial injection of zinc gluconate solution into Alzheimer's model mice post-tail vein and then the combination of another injection of ferrous chloride (FeCl2) solution into the same Alzheimer's model mice post-stomach. Our observations indicate that both zinc gluconate solution and FeCl2 solution could cross the blood-brain barrier (BBB) to biosynthesize the fluorescent zinc oxide nanoclusters and magnetic iron oxide nanoclusters, respectively, in the lesion areas of the AD model mice, thus enabling high spatiotemporal dual-modality bioimaging (i.e., including fluorescence bioimaging (FL) and magnetic resonance imaging (MRI)) of Alzheimer's disease for the first time. The result presents a novel promising strategy for the rapid and early diagnosis of Alzheimer's disease.

Biosynthesized Gold Nanoclusters and Iron Complexes as Scaffolds for Multimodal Cancer Bioimaging.

Cancer treatment has a far greater chance of success if the neoplasm is diagnosed before the onset of metastasis to vital organs. Hence, cancer early diagnosis is extremely important and remains a major challenge in modern therapeutics. In this contribution, facile and new method for rapid multimodal tumor bioimaging is reported by using biosynthesized iron complexes and gold nanoclusters via simple introduction of AuCl4- and Fe2+ ions. The observations demonstrate that the biosynthesized Au nanoclusters may act as fluorescent and computed tomography probes for cancer bioimaging while the iron complexes behave as effective contrast agent for magnetic resonance imaging. The biosynthesized iron complexes and gold nanoclusters are found biocompatible in vitro (MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay) and in vivo for all the vital organs of circulatory and excretory system. These observations raise the possibility that the biosynthesized probes may find applications in future clinical diagnosis for deep seated early neoplasms by multimodal imaging.

Protective effect of TiO2 nanowhiskers on Tetra Sulphonatophenyl Porphyrin (TSPP) complexes induced oxidative stress during photodynamic therapy.

BACKGROUND: Tetra Sulphonatophenyl Porphyrin (TSPP) is well known photosensitizer for photodynamic therapy; nevertheless, its well-known adverse effects hamper its potential use. Recently, nano TiO2's potential role in biomedical has been defined for various disease theranostics, including cancer and other infections. Thus, in this contribution we have explored the possibility of utilizing TiO2 nanowhiskers as novel strategy to lower TSPP adverse effects both in vitro, and in vivo. METHODS: Various concentrations of TSPP, TiO2-TSPP, and TiO2 were injected to three different rat groups, while fourth group was kept as control. Toxic effects were evaluated on excretory and circulatory system by using histopathology, fluorescent microscopy, complete blood cells count (CBC) and serum enzymes. RESULTS: In complete blood cells count, all cells were significantly (p<0.01) affected by the various concentration and treatment groups. The various dose concentrations and treatment also significantly (p<0.01) affected the serum enzyme parameters including AST, ALT, LDH, Creatinine and BUN level. The low concentration of TSPP-TiO2 was found to be the safest, on the bases of serum enzyme parameters, CBC, histopathology, and fluorescent microscopic analysis. The MTT assay was used to evaluate in vitro cytotoxicity, and the results demonstrated maximum viability in illuminated TSPP-TiO2 nanowhiskers group when compared with TSPP treated group. CONCLUSIONS: It was evident that increase in concentration of TSPP increased the toxic effects; however, the TiO2 nanowhiskers combination with TSPP decreased these adverse effects. Moreover, TSPP (0.1 mM) combined with TiO2 nanowhiskers (0.6 mM) was safer than TSPP (0.1 mM) alone.

Real-Time Evaluation of Live Cancer Cells by an in Situ Surface Plasmon Resonance and Electrochemical Study.

This work presents a new strategy of the combination of surface plasmon resonance (SPR) and electrochemical study for real-time evaluation of live cancer cells treated with daunorubicin (DNR) at the interface of the SPR chip and living cancer cells. The observations demonstrate that the SPR signal changes could be closely related to the morphology and mass changes of adsorbed cancer cells and the variation of the refractive index of the medium solution. The results of light microscopy images and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide studies also illustrate the release or desorption of HepG2 cancer cells, which were due to their apoptosis after treatment with DNR. It is evident that the extracellular concentration of DNR residue can be readily determined through electrochemical measurements. The decreases in the magnitudes of SPR signals were linearly related to cell survival rates, and the combination of SPR with electrochemical study could be utilized to evaluate the potential therapeutic efficiency of bioactive agents to cells. Thus, this label-free, real-time SPR-electrochemical detection technique has great promise in bioanalysis or monitoring of relevant treatment processes in clinical applications.