A Multifunctional Nanozyme with NADH Dehydrogenase-Like Activity and Nitric Oxide Release under Near-Infrared Light Irradiation as an Efficient Therapeutic for Antimicrobial Resistance Infection and Wound Healing.

In recent years, antimicrobial resistance (AMR) has become one of the greatest threats to human health. There is an urgent need to develop new antibacterial agents to effectively treat AMR infection. Herein, a novel nanozyme platform (Cu,N-GQDs@Ru-NO) is prepared, where Cu,N-doped graphene quantum dots (Cu,N-GQDs) are covalently functionalized with a nitric oxide (NO) donor, ruthenium nitrosyl (Ru-NO). Under 808 nm near-infrared (NIR) light irradiation, Cu,N-GQDs@Ru-NO demonstrates nicotinamide adenine dinucleotide (NADH) dehydrogenase-like activity for photo-oxidizing NADH to NAD+ , thus disrupting the redox balance in bacterial cells and resulting in bacterial death; meanwhile, the onsite NIR light-delivered NO effectively eradicates the methicillin-resistant Staphylococcus aureus (MRSA) bacterial and biofilms, and promotes wound healing; furthermore, the nanozyme shows excellent photothermal effect that enhances the antibacterial efficacy as well. With the combination of NADH dehydrogenase activity, photothermal therapy, and NO gas therapy, the Cu,N-GQDs@Ru-NO nanozyme displays both in vitro and in vivo excellent efficacy for MRSA infection and biofilm eradication, which provides a new therapeutic modality for effectively treating MRSA inflammatory wounds.

Cellular membrane-targeting ruthenium complexes as efficient photosensitizers for broad-spectrum antibacterial activity.

A ruthenium complex [Ru(phen)2(phen-5-amine)-C14] (Ru-C14) with broad-spectrum antibacterial activity was designed and synthesized; positively charged Ru-C14 could target bacteria via electrostatic interactions and showed high binding effectiveness to cell membranes. In addition, Ru-C14 could act as a photosensitizer. Under 465 nm light irradiation, Ru-C14 could generate 1O2, thus disrupting the bacterial intracellular redox balance and leading to bacterial death. Ru-C14 also exhibited minimum inhibitory concentration values of 6.25 µM against Escherichia coli and 3.125 µM against Staphylococcus aureus; these values are lower than those of streptomycin and methicillin. This work combined the merits of cell membrane targeting and photodynamic therapy for antibacterial activity. The findings might open up a new avenue for effective anti-infection treatment and other medical applications.

Ruthenium photosensitizer anchored gold nanorods for synergistic photodynamic and photothermal therapy.

Ruthenium polypyridyl complexes have been widely used as bioprobes and photosensitizers. However, several disadvantages including slow cellular uptake, nonspecific binding with biomolecules and toxicity limit their applications. In this study, a nanocarrier of human serum albumin coated gold nanorods was developed to deliver a ruthenium photosensitizer for PDT/PTT combination therapy. The HSA coating endowed the nanodrug with high biocompatibility and stability under physiological conditions. Ru-GNR-HSANPs generate 1O2 and hydroxyl radicals to kill cancer cells under blue light irradiation, and exhibit excellent photothermal anticancer effects under 808 nm light irradiation. Significant synergistic anticancer effects were achieved by combined PDT/PTT therapy. Importantly, Ru-GNR-HSANPs can have the synergistic PDT/PTT functions with no need of drug release from the carrier.

Metal complexes against breast cancer stem cells.

With the highest incidence, breast cancer is the leading cause of cancer deaths among women in the world. Tumor metastasis is the major contributor of high mortality in breast cancer, and the existence of cancer stem cells (CSCs) has been proven to be the cause of tumor metastasis. CSCs are a small proportion of tumor cells, and they are associated with self-renewal and tumorigenic potential. Given the significance of CSCs in tumor initiation, expansion, relapse, resistance, and metastasis, studies should investigate and discover effective anticancer agents that can not only inhibit the proliferation of differentiated tumor cells but also reduce the tumorigenic capability of CSCs. Thus, new therapies must be discovered to treat and prevent this severely hazardous disease of human beings. The success of platinum complexes in cancer treatment has laid the basic foundation for the utilization of metal complexes in the treatment of malignant cancers, in particular the highly aggressive triple-negative breast cancer. Importantly, metal complexes currently have diverse and versatile competences in the therapeutic targeting of CSCs. The anti-CSC properties provide a strong impetus for the development of novel metal-based compounds for the targeting of CSCs and treatment of chemotherapy-resistant and relapsed tumors. In this review, we provide the latest advances in metal complexes including platinum, ruthenium, osmium, iridium, manganese, cobalt, nickel, copper, zinc, palladium, and tin complexes against breast CSCs obtained over the past decade, with pertinent literature including those published until 2021.

Ruthenium Complexes as Promising Candidates against Lung Cancer.

Lung cancer is one of the most common malignancies with the highest mortality rate and the second-highest incidence rate after breast cancer, posing a serious threat to human health. The accidental discovery of the antitumor properties of cisplatin in the early 1960s aroused a growing interest in metal-based compounds for cancer treatment. However, the clinical application of cisplatin is limited by serious side effects and drug resistance. Therefore, other transition metal complexes have been developed for the treatment of different malignant cancers. Among them, Ru(II/III)-based complexes have emerged as promising anticancer drug candidates due to their potential anticancer properties and selective cytotoxic activity. In this review, we summarized the latest developments of Ru(II/III) complexes against lung cancer, focusing mainly on the mechanisms of their biological activities, including induction of apoptosis, necroptosis, autophagy, cell cycle arrest, inhibition of cell proliferation, and invasion and metastasis of lung cancer cells.

Diatom-like silica-protein nanocomposites for sustained drug delivery of ruthenium polypyridyl complexes.

Inspired by the unique glass cell wall of diatom, we design a new nanostructure of human serum albumin nanoparticle (HSANP) coated with silica (HSA/SiO2), which consists of a core-satellite assembly of small silica nanoparticles on a single HSANP. The HSA/SiO2 nanoparticles are used for delivering ruthenium polypyridyl complexes into cells. The silica coating increases the Ru loading efficiency, and prevents the burst release of Ru from HSA/SiO2. The Ru release rate can be controlled by adjusting the amount of coated silica on HSANP, affording a drug delivery system with controlled drug release rate. The Ru-HSA/SiO2 nanoparticles show high stability in physiological condition, and significantly increase the Ru uptake into cells, which proceeds via clathrin-mediated endocytosis into the lysosomes. The silica coating takes no effect on the fluorescence intensity and ROS generation of loaded Ru in HSA/SiO2. Furthermore, Ru4-HSA/SiO2 exhibit weak cytotoxicity in dark, however, the nanodrug can be activated by light irradiation and generate ROS to damage cells, thus achieving an excellent photodynamic therapy efficiency. Therefore, the diatom-like nanostructure can function as sustained drug delivery nanocarrier of ruthenium polypyridyl complex and can be used for bioimaging and photodynamic therapy.

Recent development and application of cyclometalated iridium(III) complexes as chemical and biological probes.

Iridium complexes have been widely applied as molecular sensors because of their rich photophysical properties, including large Stokes shifts, long emission lifetimes, environment-sensitive emissions, and high luminescence quantum yields. In this paper, we review the recent development and application of iridium complexes as probes for ions, anions, gaseous species, organic molecules, small biomolecules, biomacromolecules, and subcellular organelles. Our outlook for iridium-based probes is also discussed.

A Dual Functional Nanoreactor for Synergistic Starvation and Photodynamic Therapy.

The combination of photodynamic therapy (PDT) and enzyme therapy is a highly desirable approach in malignant tumor therapies as it takes advantage of the spatial-controlled PDT and the effective enzyme-catalyzed bioreactions. However, it is a challenge to co-encapsulate hydrophilic enzymes and hydrophobic photosensitizers, and these two agents often interfere with each other. In this work, a protocell-like nanoreactor (GOx-MSN@MnPc-LP) has been designed for synergistic starvation therapy and PDT. In this nanoreactor, the hydrophilic glucose oxidase (GOx) is loaded in the pore of mesoporous silica nanoparticles (MSNs), while the hydrophobic manganese phthaleincyanide (MnPc) is loaded in the membrane layer of liposome. This spatial separation of two payloads protects GOx and MnPc from the cellular environment and avoids interference with each other. GOx catalyzes the oxidation of glucose, which generates hydrogen peroxide and gluconic acid, leading to the starvation therapy via glucose consumption in cancer cells, as well as the disruption of cellular redox balance. MnPc produces cytotoxic singlet oxygen under 730 nm laser irradiation, achieving PDT. The antitumor effects of the nanoreactor have been verified on tumor cells and tumor-bearing mice models. GOx-MSN@MnPc-LP efficiently inhibits tumor growth in vivo with a single treatment, indicating the robust synergy of starvation therapy and PDT treatment. This work also offers a versatile strategy for delivering hydrophilic enzymes and hydrophobic photosensitizers using a protocell-like nanoreactor for effective cancer treatment.

A HCBP1 peptide conjugated ruthenium complex for targeted therapy of hepatoma.

An HCBP1 peptide-ruthenium conjugate (Ru-ß-Ala-FQHPSFI) as a potential candidate for targeted therapy of hepatoma was synthesized. Ru-ß-Ala-FQHPSFI shows drastically enhanced cytotoxicity and high selectivity for hepatoma cells versus noncancer liver cells. Raman imaging shows that this peptide-based drug can be taken up well by the hepatoma cells compared with the bare ruthenium complex (Ru) and the opposite sequence peptide-ruthenium conjugate (Ru-ß-Ala-IFSPHQF). This study presents a new strategy for the construction of tumor-targeting metal-based anticancer therapeutics.

Modular design of nanobody-drug conjugates for targeted-delivery of platinum anticancer drugs with an MRI contrast agent.

A multifunctional nanobody-drug conjugate (NDC) was constructed in this work for the targeted delivery of a platinum prodrug and an MRI contrast agent. The NDC can be specifically internalized into EGFR positive cancer cells, resulting in higher therapeutic effect and lower side-effects relative to cisplatin. The Gd-binding domain enables the in situ detection of the drug distribution in vivo.

Reaction of Histone H1 with trans-Platinum Complexes and the Effect on DNA Platination.

Transplatin is an inactive platinum drug; however, a number of analogues, such as trans-EE and trans-PtTz, demonstrate promising antitumor activity in vitro and in vivo. Although the ultimate target is nuclear DNA, increasing evidence indicate that proteins also play important roles in the display of antitumor activity. The linker histone H1 is situated by the portal between the unwrapped DNA and the nucleosome core. Our recent study revealed that H1 can readily react with cisplatin, and the adducts tend to form ternary complexes with DNA. In this work, we have investigated the reaction of histone H1 with two antitumor-active trans-oriented complexes, trans-EE and trans-PtTz, and the effect of H1 upon the platination of DNA. The results show that trans-platinum drugs are much more reactive than cisplatin toward H1. Interestingly, in addition to the expected bidentate adducts (by displacement of the two labile chlorido ligands), also a tridentate adduct can be formed by displacement of one nonlabile carrier ligand of trans-EE or trans-PtTz. The trans-Pt/H1 adducts can then react with DNA and generate protein-Pt-DNA ternary complexes. Additionally, platinum can be transferred from trans-Pt/H1 adducts to DNA, generating binary trans-Pt/DNA complexes. Such a transfer of the platinum agent to DNA was not observed in the reaction of cisplatin. Furthermore, the detailed investigation carried out on a model peptide indicates that H1 promotes the DNA platination by trans- EE, while it reduces that of trans-PtTz and cisplatin. These results suggest that H1 can play a key role in the DNA platination and modulate the efficacy of different platinum agents.

Design of ruthenium-albumin hydrogel for cancer therapeutics and luminescent imaging.

Improving cell uptake of metal compounds has became an important goal in the field of metal-based anticancer agents. This may combat platinum resistance and side effects seen commonly in current anticancer chemotherapy regimes. Here, we explore a novel degradable ruthenium-albumin hydrogel, which shows strong luminescence for cell imaging and high selectivity for cancer cells versus non-cancer cells. This is an early indication of the possibility of reducing unwanted side effects of metals by using bovine serum albumin hydrogel as a delivery strategy. This work provides a strong basis for development of a new class of metal-based cancer therapeutic agents.

Charge-Selective Delivery of Proteins Using Mesoporous Silica Nanoparticles Fused with Lipid Bilayers.

Efficient and safe intracellular delivery of proteins is highly desired in the development of protein therapeutics. Current methods of protein delivery commonly suffer from low loading efficiency, low stability in serum, and lack of versatility for different proteins. Here, we developed a platform for efficient protein delivery using mesoporous silica nanoparticles (MSN) with lipid fusion. By different surface modifications on MSN, the positively charged MSN (MSN+) and the negatively charged MSN (MSN-), were generated for loading different proteins. The cargo proteins, based on the surface charges, can be selectively loaded in very high efficiency. The protein-loaded MSNs were fused with liposomes to form a protocell-like delivery system (MSN-LP) in order to prevent burst release of proteins. The lipid fusion significantly increases the stability of the nanosystem in physiological conditions, and the MSN-LP protocell can efficiently deliver proteins into cells. The cargo proteins can be released in cells in a sustained manner. Fifteen different proteins, including two protein complexes, were tested using this delivery system. Further analyses indicate that the proteins can maintain their functions after delivery into cells. Fluorescent proteins, GFP, and KillerRed show fluorescence in cells, indicating the correct folding of proteins during encapsulation and delivery. Protein activity analysis shows that KillerRed protein can generate ROS in cells, while SOD can eliminate ROS in cells. Hence, the proteins delivered by this system remain their structure and function in cells. This work provides a versatile strategy for charge-selective delivery of proteins with high loading efficiency and high stability.

Transglutaminase mediated PEGylation of nanobodies for targeted nano-drug delivery.

Targeted delivery of anticancer drugs that selectively accumulate in malignant cells could enhance drug efficacy and reduce side effects of conventional chemotherapy. In this work, we designed a single domain antibody (nanobody) based drug delivery system for targeted delivery of anticancer drugs. An anti-EGFR nanobody (Nb) was constructed with a C3-tag and a Q-tag for site specific modifications under physiological conditions. The site specific PEGylation of the nanobody was achieved via a transglutaminase catalyzed reaction through the coupling of the Q-tag with PEG-NH2. As a proof of concept, the PEGylated nanobody was tethered to HSA coated upconversion nanoparticles (UCNPs) through the C3-tag, and an anticancer drug, doxorubicin (DOX), was loaded. Results showed that the Nb-conjugated drug delivery system exhibits superior specificity to the EGFR positive tumor cells. The drug delivery system is highly accumulated in the EGFR positive tumor cells (A431), whereas there was no detectable accumulation in the EGFR negative cells (MCF-7). Consequently, the drug loaded particles demonstrated significantly higher anti-proliferation to A431 cells than to MCF-7 cells. This work provides an effective approach for site-specific modification of nanobodies for the construction of targeted drug delivery systems.

Co-delivery of all-trans-retinoic acid enhances the anti-metastasis effect of albumin-bound paclitaxel nanoparticles.

Co-delivery of all-trans-retinoic acid and paclitaxel using albumin-bound nanoparticles demonstrated a significantly improved anti-metastatic effect to breast cancer both in vitro and in vivo. Notably, the co-delivery nanoparticles exhibited more pronounced therapeutic effects than the combination of two free drugs or two HSA loaded single drugs.

Asplatin enhances drug efficacy by altering the cellular response.

Aspirin, a widely used anti-inflammatory drug, has been shown to be effective for the prevention and remission of cancers (Science, 2012, 337(21) 1471-1473). Asplatin, a Pt(iv) prodrug of cisplatin with the ligation of aspirin (c,c,t-[PtCl2(NH3)2(OH)(aspirin)]), demonstrates significantly higher cytotoxicity than cisplatin towards tumor cells and almost fully overcomes the drug resistance of cisplatin resistant cells. In this work, we have studied the molecular mechanism of asplatin by investigating the cellular response to this compound in order to understand the prominent inhibitory effect on the proliferation of cancer cells. The apoptosis analyses and the related gene expression measurements show that aspirin released from asplatin significantly modulates the cellular response to the platinum agent. Asplatin promotes the apoptosis via the BCL-2 associated mitochondrial pathway. The down-regulation of BCL-2 along with the up-regulation of BAX and BAK enhances the mitochondrial outer membrane permeability, resulting in the cytochrome c release from mitochondria into the cytosol. This event promotes the apoptosis by activation of caspase processing. Consequently, the ligation of aspirin significantly enhances the drug efficacy of the platinum complex in the low micromolar range. The alteration of the cellular response is probably responsible for the circumvention of the cisplatin resistance by asplatin. These results provide an insight into the mechanism of asplatin and provide information for designing new classic platinum drugs.

A dual-fluorescent nano-carrier for delivering photoactive ruthenium polypyridyl complexes.

A dual-fluorescent nanocarrier was synthesized by coating human serum albumin (HSA) on lanthanide-doped upconversion nanoparticles (UCNPs). The HSA coating makes the particles highly biocompatible and well dispersed in aqueous solutions. This nanocarrier demonstrates two types of fluorescence: the blue upconversion fluorescence under excitation with a 980 nm light of the UCNP core, and the green fluorescence under the excitation with a 450 nm light of the polymerized HSA layer. This dual-fluorescence property makes the material more applicable in bio-imaging. A photo-sensitive ruthenium complex ([Ru(bpy)2(dmbpy)2]Cl2, Ru-1) was loaded to generate the light-responsive Ru-HSA-UCNPs. This conjugate showed very low inhibitory effect on cell proliferation in the dark, while light irradiation significantly enhanced its cytotoxicity to cancer cells. Further investigations showed that irradiation activated Ru-1 and the product became highly reactive to DNA. This result suggests the potential application of this conjugate in the controlled release of active anticancer agents in tumor sites.

Oral delivery of a platinum anticancer drug using lipid assisted polymeric nanoparticles.

Self-assembled cholesterol-asplatin-incorporated nanoparticles (SCANs) were prepared for oral delivery of a Pt(IV) prodrug. SCANs exhibit high gastrointestinal stability, sustained drug release and enhanced cell uptake. The oral bioavailability of SCANs was 4.32-fold higher than that of free Pt(IV) prodrugs. The oral administration of SCANs efficaciously inhibits tumor growth with negligible toxicity.

Human Serum Albumin Conjugated Nanoparticles for pH and Redox-Responsive Delivery of a Prodrug of Cisplatin.

Platinum anticancer drugs are particularly in need of controlled drug delivery because of their severe side effects. Platinum(IV) agents are designed as prodrugs to reduce the side effects of platinum(II) drugs; however, premature reduction could limit the effect as a prodrug. In this work, a highly biocompatible, pH and redox dual-responsive delivery system is prepared by using hybrid nanoparticles of human serum albumin (HSA) and calcium phosphate (CaP) for the Pt(IV) prodrug of cisplatin. This conjugate is very stable under extracellular conditions, so that it protects the platinum(IV) prodrug in HSA. Upon reaching the acidic and hypoxic environment, the platinum drug is released in its active form and is able to bind to the target DNA. The Pt-HSA/CaP hybrid inhibits the proliferation of various cancer cells more efficiently than cisplatin. Different cell cycle arrests suggest different cellular responses of the Pt(IV) prodrug in the CaP nanocarrier. Interestingly, this delivery system demonstrates enhanced cytotoxicity to tumor cells, but not to normal cells.

A versatile pH-responsive platform for intracellular protein delivery using calcium phosphate nanoparticles.

Intracellular protein delivery has great biomedical applications; the safe and efficient delivery vectors are crucial for achieving this goal. Here we report a platform for efficient protein delivery using calcium phosphate (CaP) nanoparticles. The well-dispersed and highly-stable nanoparticles (∼100 nm) are prepared with the protein loading capacity of up to 29%. The nanoparticles are stable under serum conditions; however, after being internalized into cells, the particles quickly release protein in the weak acidic endosomes/lysosomes. The decomposition of CaP promotes the endo-lysosomal escape of proteins released from nanoparticles. The protein/CaP conjugates were prepared under mild conditions (aqueous solution, room temperature); hence the protein released from nanoparticles retained its folding and function. In addition, all materials used for the preparation are highly biocompatible. This method has been applied for the loading of three model proteins, BSA, GFP and KillerRed; similar loading properties were observed on these proteins. Therefore, this work offers a general approach for intracellular protein delivery, which could be applicable for therapeutic proteins.