Cell-Penetrating and Enzyme-Responsive Peptides for Targeted Cancer Therapy: Role of Arginine Residue Length on Cell Penetration and In Vivo Systemic Toxicity.

For the improved delivery of cancer therapeutics and imaging agents, the conjugation of cell-penetrating peptides (CPPs) increases the cellular uptake and water solubility of agents. Among the various CPPs, arginine-rich peptides have been the most widely used. Combining CPPs with enzyme-responsive peptides presents an innovative strategy to target specific intracellular enzymes in cancer cells and when combined with the appropriate click chemistry can enhance theranostic drug delivery through the formation of intracellular self-assembled nanostructures. However, one drawback of CPPs is their high positive charge which can cause nonspecific binding, leading to off-target accumulation and potential toxicity. Hence, balancing cell-specific penetration, toxicity, and biocompatibility is essential for future clinical efficacy. We synthesized six cancer-specific, legumain-responsive RnAANCK peptides containing one to six arginine residues, with legumain being an asparaginyl endopeptidase that is overexpressed in aggressive prostate tumors. When conjugated to Alexa Fluor 488, R1-R6AANCK peptides exhibited a concentration- and time-dependent cell penetration in prostate cancer cells, which was higher for peptides with higher R values, reaching a plateau after approximately 120 min. Highly aggressive DU145 prostate tumor cells, but not less aggressive LNCaP cells, self-assembled nanoparticles in the cytosol after the cleavage of the legumain-specific peptide. The in vivo biocompatibility was assessed in mice after the intravenous injection of R1-R6AANCK peptides, with concentrations ranging from 0.0125 to 0.4 mmol/kg. The higher arginine content in R4-6 peptides showed blood and urine indicators for the impairment of bone marrow, liver, and kidney function in a dose-dependent manner, with instant hemolysis and morbidity in extreme cases. These findings underscore the importance of designing peptides with the optimal arginine residue length for a proper balance of cell-specific penetration, toxicity, and in vivo biocompatibility.

Tumor-tropic Trojan horses: Using mesenchymal stem cells as cellular nanotheranostics.

Various classes of nanotheranostics have been developed for enhanced tumor imaging and therapy. However, key limitations for a successful use of nanotheranostics include their targeting specificity with limited off-site tissue accumulation as well as their distribution and prolonged retention throughout the entire tumor. Due to their inherent tumor-tropic properties, the use of mesenchymal stem cells (MSCs) as a "Trojan horse" has recently been proposed to deliver nanotheranostics more effectively. This review discusses the current status of "cellular nanotheranostics" for combined (multimodal) imaging and therapy in preclinical cancer models. Emphasis is placed on the limited knowledge of the signaling pathways and molecular mechanisms of MSC tumor-tropism, and how such information may be exploited to engineer MSCs in order to further improve tumor homing and nanotheranostic delivery using image-guided procedures.

A Smart Intracellular Self-Assembling Bioorthogonal Raman Active Nanoprobe for Targeted Tumor Imaging.

Inspired by the principle of in situ self-assembly, the development of enzyme-activated molecular nanoprobes can have a profound impact on targeted tumor detection. However, despite their intrinsic promise, obtaining an optical readout of enzyme activity with high specificity in native milieu has proven to be challenging. Here, a fundamentally new class of Raman-active self-assembling bioorthogonal enzyme recognition (nanoSABER) probes for targeted tumor imaging is reported. This class of Raman probes presents narrow spectral bands reflecting their vibrational fingerprints and offers an attractive solution for optical imaging at different bio-organization levels. The optical beacon harnesses an enzyme-responsive peptide sequence, unique tumor-penetrating properties, and vibrational tags with stretching frequencies in the cell-silent Raman window. The design of nanoSABER is tailored and engineered to transform into a supramolecular structure exhibiting distinct vibrational signatures in presence of target enzyme, creating a direct causality between enzyme activity and Raman signal. Through the integration of substrate-specific for tumor-associated enzyme legumain, unique capabilities of nanoSABER for imaging enzyme activity at molecular, cellular, and tissue levels in combination with machine learning models are shown. These results demonstrate that the nanoSABER probe may serve as a versatile platform for Raman-based recognition of tumor aggressiveness, drug accumulation, and therapeutic response.

Targeted Enzyme Activity Imaging with Quantitative Phase Microscopy.

Quantitative phase imaging (QPI) is a powerful optical imaging modality for label-free, rapid, and three-dimensional (3D) monitoring of cells and tissues. However, molecular imaging of important intracellular biomolecules such as enzymes remains a largely unexplored area for QPI. Herein, we introduce a fundamentally new approach by designing QPI contrast agents that allow sensitive detection of intracellular biomolecules. We report a new class of bio-orthogonal QPI-nanoprobes for in situ high-contrast refractive index (RI) imaging of enzyme activity. The nanoprobes feature silica nanoparticles (SiO2 NPs) having higher RI than endogenous cellular components and surface-anchored cyanobenzothiazole-cysteine (CBT-Cys) conjugated enzyme-responsive peptide sequences. The nanoprobes specifically aggregated in cells with target enzyme activity, increasing intracellular RI and enabling precise visualization of intracellular enzyme activity. We envision that this general design of QPI-nanoprobes could open doors for spatial-temporal mapping of enzyme activity with direct implications for disease diagnosis and evaluating the therapeutic efficacy.

Tumor acidic environment directs nanoparticle impacts on cancer cells.

Despite impressive progress in nanotechnology-based cancer therapy being made by in vitro research, few nanoparticles (NPs) have been translated into clinical trials. The wide gap between in vitro results and nanomedicine's clinical translation might be partly due to acidic microenvironment of cancer cells being ignored in in vitro studies. To check this hypothesis, we studied the biological impacts of two different structures of NPs on cancer cells (MDA-MB 231) at acidic (pH: 6.5) low (pH: 7) and physiological pH (pH: 7.4). We uncovered that a slight change in the pH of the cancer cell microenvironment affects the cellular uptake efficacy and toxicity mechanism of nanographene sheets and SPION@silica nanospheres. Both nanostructures exerted more substantial toxic impacts (e. g. apoptosis, necrosis, membrane disruption, and oxidative stress induction) against cancer cells at physiological pH compared to acidic niche. They also differently slowed or arrested phases of the cell cycle at different pH (S and G2/M at normal pH while G0/G1 at acidic/low pH). More specifically, cancer cells expressed higher levels of interleukins involved in cancer cell resistance at acidic pH than those incubated at physiological pH. This study revealed that a slight change in extracellular pH of cancer cells could strongly affect the therapeutic/toxic impact of nanomaterials and therefore, it should be considered in the future cancer nanomedicine research.

Photodynamic therapy-mediated extirpation of cutaneous-resistant dermatophytosis with Ag@ZnO nanoparticles: an efficient therapeutic approach for onychomycosis.

Aim: The aim of this study was to determine whether photodynamic therapy of resistant onychomycosis with Ag@ZnO nanoparticles can promote the treatment procedure and extirpates the recurrence of fungal infection. Methods: Ag@ZnO nanoparticles (NPs) under UVB-radiation were applied to treat T. rubrum and T. mentagrophytes in vitro through photodynamic therapy. In vivo therapeutic efficacy, biocompatibility and biodistribution of Ag@ZnO NPs were studied. Results: 40 µg/ml of UVB-activated Ag@ZnO NPs showed 100% antifungal activity against dermatophytosis in vitro and in vivo followed by complete growth prevention by degeneration of spores and mycelium after 180 days, while posed biocompatibility. Conclusion: This study showed the superiority of photodynamic therapy with Ag@ZnO NPs followed by proper regeneration of the skin with Zinc ion of the shell.

Hybrid PCL/chitosan-PEO nanofibrous scaffolds incorporated with A. euchroma extract for skin tissue engineering application.

Skin tissue engineering is an advanced method to repair and regenerate skin injuries. Recent research is focused on the development of scaffolds that are safe, bioactive, and cytocompatible. In this work, a new hybrid nanofibrous scaffold composed of polycaprolactone/chitosan-polyethylene oxide (PCL/Cs-PEO) incorporated with Arnebia euchroma (A. euchroma) extract were synthesized by the two-nozzle electrospinning method. Then the synthesized scaffold was characterized for morphology, sustainability, chemical structure and properties. Moreover, to verify their potential in the burn wound healing process, biodegradation rate, contact angle, swelling properties, water vapor permeability, mechanical properties, antibacterial activity and drug release profile were measured. Furthermore, cytotoxicity and biocompatibility tests were performed on human dermal fibroblasts cell line via XTT and LDH assay. It is shown that the scaffold improved and increased proliferation during in-vitro studies. Thus, results confirm the efficacy and potential of the hybrid nanofibrous scaffold for skin tissue engineering.

Epinephrine-entrapped chitosan nanoparticles covered by gelatin nanofibers: A bi-layer nano-biomaterial for rapid hemostasis.

Uncontrolled hemorrhage accounts for significant death risk both in trauma and surgery. Various bleeding control techniques have been emerged to augment hemostasis, which still has several limitations and drawbacks. In this study, epinephrine-entrapped chitosan nanoparticles were electrosprayed on a base pad and covered by a gelatin nanofiber layer (E-CS-Gl. Physico-chemical characteristics, hemocompatibility, cytotoxicity, and blood coagulation tests were studied in-vitro, and blood coagulation and hemostasis potential tests were performed in-vivo. The in-vitro results showed that the prepared nano-biomaterial is cytocompatible against HuGu cells. Also, hemocompatibility studies showed that PT and aPTT times did not change in comparison with the controls. Further blood coagulation study indicated that E-CS-Gl provides an ultimate interface to induce red blood cell absorption and aggregation, resulting in augmented blood coagulation. E-CS-Gl also caused rapid clotting in rat models of ruptured femoral artery and liver compared to controls. Findings exhibited that E-CS-Gl is a safe and effective hemostatic agent and provides a new approach for fast and safe hemorrhage control.

Colloidal synthesis of tunably luminescent AgInS-based/ZnS core/shell quantum dots as biocompatible nano-probe for high-contrast fluorescence bioimaging.

Tremendous demands for simultaneous imaging of biological entities, along with the drawback of photobleaching in fluorescent dyes, have encouraged scientists to apply novel and non-toxic colloidal quantum dots (QDs) in biomedical researches. Herein, a novel aqueous-phase approach for the preparation of multicomponent In-based QDs is reported. Absorption and photoluminescence emission spectra of the as-prepared QDs were tuned by alteration of QDs' composition as Zn-Ag-In-S/ZnS, Ag-In-S/ZnS and Cu-Ag-In-S/ZnS core/shell QDs. In order to reach reproducibly intense and tunable light-emissive colloidal QDs with green, amber, and red color, various optimization steps were carefully performed. The structural characterizations such as EDX, ICP-AES, XRD, TEM and FT-IR measurements were also carried out to demonstrate the success of the present method to prepare extremely quantum-confined QDs capped with functional groups. Then, to ensure their promising biomedical applications, the generated intracellular reactive oxygen species (ROS) by QDs were quantitatively and qualitatively measured in dark conditions and under 405 nm laser irradiation. Our results verified an enhancement in the generation of reactive oxygen species (ROS) and cytotoxic effects in the presence of laser irradiation while their muted toxic effects in dark conditions confirmed biocompatible properties of un-excited In-based QDs. Moreover, bioimaging analysis revealed strong merits of the suggested synthetic route to achieve ideal fluorescent QDs as bright/multi-color optical nano-probes in imaging and transporting pumps in the cell membrane. This further emphasized the potential ability of the present AgInS-based/ZnS QDs in obtaining required results as theranostic agents for simultaneous treatment and imaging of cancer. The harmonized advantages in simplicity and effectiveness of synthesis procedure, excellent structural/optical properties enriched with confirmed biomedical merits in high contrast imaging and potential treatment highlight the present work.

Supramolecular Insights into Domino Effects of Ag@ZnO-Induced Oxidative Stress in Melanoma Cancer Cells.

Recent studies suggest that cancer cell death accompanied by organelle dysfunction might be a promising approach for cancer therapy. The Golgi apparatus has a key role in cell function and may initiate signaling pathways to mitigate stress and, if irreparable, start apoptosis. It has been shown that Golgi disassembly and fragmentation under oxidative stress act as indicators for stress-mediated cell death pathways through cell cycle arrest in the G2/M phase. The present study shows that UV-induced reactive oxygen species (ROS) generation by Ag@ZnO nanoparticles (NPs) transform the Golgi structures from compressed perinuclear ribbons into detached vesicle-like structures distributed in the entire cytoplasm of melanoma cells. This study also demonstrates that Ag@ZnO NP-induced Golgi fragmentation cooccurs with G2 block of cell cycle progression, preventing cells from entering the mitosis phase. Additionally, the increased intracellular ROS production triggered by Ag@ZnO NPs upon UV exposure promoted autophagy. Taken together, Ag@ZnO NPs induce stress-related Golgi fragmentation and autophagy, finally leading to melanoma cell apoptosis. Intracellular oxidative stress generated by Ag@ZnO NPs upon UV irradiation may thus represent a targeted approach to induce cancer cell death through organelle destruction in melanoma cells, while fibroblast cells remained largely unaffected.

Molecular interaction of fibrinogen with zeolite nanoparticles.

Fibrinogen is one of the key proteins that participate in the protein corona composition of many types of nanoparticles (NPs), and its conformational changes are crucial for activation of immune systems. Recently, we demonstrated that the fibrinogen highly contributed in the protein corona composition at the surface of zeolite nanoparticles. Therefore, understanding the interaction of fibrinogen with zeolite nanoparticles in more details could shed light of their safe applications in medicine. Thus, we probed the molecular interactions between fibrinogen and zeolite nanoparticles using both experimental and simulation approaches. The results indicated that fibrinogen has a strong and thermodynamically favorable interaction with zeolite nanoparticles in a non-cooperative manner. Additionally, fibrinogen experienced a substantial conformational change in the presence of zeolite nanoparticles through a concentration-dependent manner. Simulation results showed that both E- and D-domain of fibrinogen are bound to the EMT zeolite NPs via strong electrostatic interactions, and undergo structural changes leading to exposing normally buried sequences. D-domain has more contribution in this interaction and the C-terminus of γ chain (γ377-394), located in D-domain, showed the highest level of exposure compared to other sequences/residues.

Luminescent, low-toxic and stable gradient-alloyed Fe:ZnSe(S)@ZnSe(S) core:shell quantum dots as a sensitive fluorescent sensor for lead ions.

In this paper, an aqueous-based approach is introduced for facile, fast, and green synthesis of gradient-alloyed Fe-doped ZnSe(S)@ZnSe(S) core:shell quantum dots (QDs) with intense and stable emission. Co-utilization of co-nucleation and growth doping strategies, along with systematic optimization of emission intensity, provide a well-controllable/general method to achieve internally doped QDs (d-dots) with intense emission. Results indicate that the alloyed ZnSe(S)@ZnSe(S) core:shell QDs have a gradient structure that consists of a Se-rich core and a S-rich shell. This gradient structure cannot only passivate the core d-dots by means of the wider band gap S-rich shell, but also minimizes the lattice mismatch between alloyed core-shell structures. Using this novel strategy and utilizing the wider band gap S-rich shell can obviously increase the cyan emission intensity and also drastically improve the emission stability against chemical and optical corrosion. Furthermore, the cytotoxicity experiments indicate that the obtained d-dots are nontoxic nanomaterials, and thus they can be considered as a promising alternative to conventional Cd-based QDs for fluorescent probes in biological fields. Finally, it is demonstrated that the present low-toxicity and gradient-alloyed core:shell d-dots can be used as sensitive chemical detectors for Pb2+ ions with excellent selectivity, small detection limit, and rapid response time.

Intracellular ROS Induction by Ag@ZnO Core-Shell Nanoparticles: Frontiers of Permanent Optically Active Holes in Breast Cancer Theranostic.

In this study, we investigated whether ZnO coating on Ag nanoparticles (NPs) tunes electron flux and hole figuration at the metal-semiconductor interface under UV radiation. This effect triggers the photoactivity and generation of reactive oxygen species from Ag@ZnO NPs, which results in enhanced cytotoxic effects and apoptotic cell death in human breast cancer cells (MDA-MB231). In this context, upregulation of apoptotic cascade proteins (i.e., Bax/Bcl2 association, p53, cytochrome c, and caspase-3) along with activation of oxidative stress proteins suggested the occurrence of apoptosis by Ag@ZnO NPs in cancer cells through the mitochondrial pathway. Also, preincubation of breast cancer cells with Ag@ZnO NPs in dark conditions muted NP-related toxic effects and consequent apoptotic fate, highlighting biocompatible properties of unexcited Ag@ZnO NPs. Furthermore, the diagnostic efficacy of Ag@ZnO NPs as computed tomography (CT)/optical nanoprobes was investigated. Results confirmed the efficacy of the photoactivated system in obtaining desirable outcomes from CT/optical imaging, which represents novel theranostic NPs for simultaneous imaging and treatment of cancer.

Delivery of adapalene using a novel topical gel based on tea tree oil nano-emulsion: Permeation, antibacterial and safety assessments.

The aim of present study was to design and optimize 0.1% adapalene loaded nano-emulsion to improve the drug efficacy and increase its user compliance. Effect of type and concentration of surfactants was studied on size of 0.1% adapalene loaded nano-emulsion. Optimized formulation was then evaluated for particle size, polydispersity index, morphology, viscosity, and pH. Subsequently, 1% carbopol® 934 was incorporated to the optimized formulation for preparation of its gel form. The efficacy and safety of 0.1% adapalene loaded nano-emulsion gel was assessed compared to marketed gel containing 0.1% adapalene. In-vitro studies showed that adapalene permeation through the skin was negligible in both adapalene loaded nano-emulsion gel and adapalene marketed gel. Furthermore, drug distribution studies in skin indicated higher retention of adapalene in the dermis in adapalene loaded nano-emulsion gel compared with adapalene marketed gel. Antibacterial activity against Propionibacterium acnes showed that adapalene loaded nano-emulsion is effective in reducing minimum inhibitory concentration of the formulation in comparison with tea tree oil nano-emulsion, and pure tea tree oil. In vivo skin irritation studies showed absence of irritancy for adapalene loaded nano-emulsion gel. Also, blood and liver absorption of the drug, histological analysis of liver and liver enzyme activity of rats after 90 days' treatment were investigated. No drug was detected in blood/liver which in addition to an absence of any adverse effect on liver and enzymes showed the potential of adapalene loaded nano-emulsion gel as a novel carrier for topical delivery of adapalene.

Correction to: Promising Antibacterial Effects of Silver Nanoparticle-Loaded Tea Tree Oil Nanoemulsion: a Synergistic Combination Against Resistance Threat.

With regrets, there is an error in the name of one of the authors which has only been noticed after publication.

Promising Antibacterial Effects of Silver Nanoparticle-Loaded Tea Tree Oil Nanoemulsion: a Synergistic Combination Against Resistance Threat.

Highly resistant pathogens may be developed in patients with immune disorders after prolonged exposure to antibiotics, a growing threat worldwide. In order to overcome these problems, this study introduces a new class of engineered nanosystems comprising of tea tree oil nanoemulsion (TTO NE) loaded with Ag nanoparticles (NPs). Silver shows a strong toxicity towards a wide range of microorganisms. Also, TTO NE could be employed as a promising and safe antimicrobial agent for local therapies of bacterial infections. The nanosystem was prepared by low-energy method. Mean droplet size of the NE was found to be 17.7 nm. Results of the antibacterial assays showed promising ability of the designed nanosystem for eradication of Gram-positive and Gram-negative bacteria (95%). Also, it was shown that introducing colloidal Ag NPs to the TTO NE exerted a synergistic effect against Escherichia coli (FIC 0.48) while only an additive effect was observed against Staphylococcus aureus (FIC 0.75). The antibacterial effects of TTO NE+Ag NPs together with their compatibility with human cells can present them as a suitable candidate to fight against the antibacterial resistance threat.

Harnessing the Cancer Radiation Therapy by Lanthanide-Doped Zinc Oxide Based Theranostic Nanoparticles.

In this paper, doping of europium (Eu) and gadolinium (Gd) as high-Z elements into zinc oxide (ZnO) nanoparticles (NPs) was designed to optimize restricted energy absorption from a conventional radiation therapy by X-ray. Gd/Eu-doped ZnO NPs with a size of 9 nm were synthesized by a chemical precipitation method. The cytotoxic effects of Eu/Gd-doped ZnO NPs were determined using MTT assay in L929, HeLa, and PC3 cell lines under dark conditions as well as exposure to ultraviolet, X-ray, and γ radiation. Doped NPs at 20 µg/mL concentration under an X-ray dose of 2 Gy were as efficient as 6 Gy X-ray radiation on untreated cells. It is thus suggested that the doped NPs may be used as photoinducers to increase the efficacy of X-rays within the cells, consequently, cancer cell death. The doped NPs also could reduce the received dose by normal cells around the tumor. Additionally, we evaluated the diagnostic efficacy of doped NPs as CT/MRI nanoprobes. Results showed an efficient theranostic nanoparticulate system for simultaneous CT/MR imaging and cancer treatment.

Impacts of quantum dots in molecular detection and bioimaging of cancer.

INTRODUCTION: A number of assays have so far been exploited for detection of cancer biomarkers in various malignancies. However, the expression of cancer biomarker(s) appears to be extremely low, therefore accurate detection demands sensitive optical imaging probes. While optical detection using conventional fluorophores often fail due to photobleaching problems, quantum dots (QDs) offer stable optical imaging in vitro and in vivo. METHODS: In this review, we briefly overview the impacts of QDs in biology and its applications in bioimaging of malignancies. We will also delineate the existing obstacles for early detection of cancer and the intensifying use of QDs in advancement of diagnostic devices. RESULTS: Of the QDs, unlike the II-VI type QDs (e.g., cadmium (Cd), selenium (Se) or tellurium (Te)) that possess inherent cytotoxicity, the I-III-VI 2 type QDs (e.g., AgInS2, CuInS2, ZnS-AgInS2) appear to be less toxic bioimaging agents with better control of band-gap energies. As highly-sensitive bioimaging probes, advanced hybrid QDs (e.g., QD-QD, fluorochrome-QD conjugates used for sensing through fluorescence resonance energy transfer (FRET), quenching, and barcoding techniques) have also been harnessed for the detection of biomarkers and the monitoring of delivery of drugs/genes to the target sites. Antibody-QD (Ab-QD) and aptamer- QD (Ap-QD) bioconjugates, once target the relevant biomarker, can provide highly stable photoluminescence (PL) at the target sites. In addition to their potential as nanobiosensors, the bioconjugates of QDs with homing devices have successfully been used for the development of smart nanosystems (NSs) providing targeted bioimaging and photodynamic therapy (PDT). CONCLUSION: Having possessed great deal of photonic characteristics, QDs can be used for development of seamless multifunctional nanomedicines, theranostics and nanobiosensors.