While immunotherapy has become a highly promising paradigm for cancer treatment in recent years, it has long been recognized that photodynamic therapy (PDT) has the ability to trigger antitumor immune responses. However, conventional PDT triggered by visible light has limited penetration depth, and its generated immune responses may not be robust enough to eliminate tumors. Herein, upconversion nanoparticles (UCNPs) are simultaneously loaded with chlorin e6 (Ce6), a photosensitizer, and imiquimod (R837), a Toll-like-receptor-7 agonist. The obtained multitasking UCNP-Ce6-R837 nanoparticles under near-infrared (NIR) irradiation with enhanced tissue penetration depth would enable effective photodynamic destruction of tumors to generate a pool of tumor-associated antigens, which in the presence of those R837-containing nanoparticles as the adjuvant are able to promote strong antitumor immune responses. More significantly, PDT with UCNP-Ce6-R837 in combination with the cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) checkpoint blockade not only shows excellent efficacy in eliminating tumors exposed to the NIR laser but also results in strong antitumor immunities to inhibit the growth of distant tumors left behind after PDT treatment. Furthermore, such a cancer immunotherapy strategy has a long-term immune memory function to protect treated mice from tumor cell rechallenge. This work presents an immune-stimulating UCNP-based PDT strategy in combination with CTLA-4 checkpoint blockade to effectively destroy primary tumors under light exposure, inhibit distant tumors that can hardly be reached by light, and prevent tumor reoccurrence via the immune memory effect.Keywords: checkpoint blockade; immune memory; immunotherapy; photodynamic therapy; upconversion nanoparticles;

Given the complexity of tumors, several nanomaterial-based treatment modalities like chemotherapy (CT), photodynamic therapy (PDT) and photothermal therapy (PTT) have been developed for combating cancers. However, it is still unclear which strategy is better or how to select optimal approaches for combination treatment since each strategy has been investigated under different conditions. Inspired by its good payload capacity and unique near-infrared absorption, reduced graphene oxide (rGO) was selected in this study as the carrier for loading of doxorubicin (DOX), a chemotherapy drug, and chlorin e6 (Ce6), a photosensitizer. The therapeutic efficacies of PTT, CT and PDT were systematically investigated in vitro using 2D culture and multicellular tumor spheroid (3D) models. Interestingly, while all three types of therapies delivered by rGO appeared to be effective in the conventional 2D cell culture model, only PTT but not CT and PDT showed great treatment efficacy in the 3D tumor spheroid model at the tested concentrations. Such a difference is due to the fact that heat diffusion is much more efficient than the diffusion of therapeutic molecules inside the tumor. Furthermore, in vivo evidence also confirmed the unique advantage of PTT compared to the other two treatment modalities using the TdT-mediated dUTP nick end labeling (TUNEL) staining assay. This study highlights the unique advantages of nanomedicine-based photothermal therapy among these three modalities in cancer treatment in terms of killing tumor cells located far from tumor blood vessels.

Whether graphene and graphene oxide (GO) would affect the activities of bacteria has been under debate. Nevertheless, how graphene derivatives with biocompatible coatings interact with microorganisms and the underlying mechanisms are important issues for nanobiotechnology, and remain to be further explored. Herein, three new types of nano-GOs functionalized with polyethylene glycol (nGO-PEGs) were synthesized by varying the PEGylation degree, and their effects on Escherichia coli (E. coli) were carefully investigated. Interestingly, nGO-PEG (1:1), the one with relatively lower PEGylation degree, could significantly stimulate bacterial growth, whereas as-made GO and the other two nGO-PEGs showed no effect. Further analysis revealed that nGO-PEG (1:1) treatment significantly accelerated FtsZ-ring assembly, shortening Phase 1 in the bacterial cell cycle. Both DNA synthesis and extracellular polymeric substance (EPS) secretion were also dramatically increased. This unique phenomenon suggests promising potentials in microbial engineering as well as in clinical detection of bacterial pathogens. As a proof-of-concept, nGO-PEG (1:1) treatment could remarkably enhance (up to 6-fold) recombinant protein production in engineered bacteria cells. To our best knowledge, this is the first demonstration of functionalized GO as a novel, positive regulator in microbial engineering. Moreover, our work highlights the critical role of surface chemistry in modulating the interactions between nanomaterials and microorganisms.

Recently, conjugated polymers have been widely explored in the field of nanomedicine. Careful evaluations of their biological effects are thus urgently needed. Hereby, we systematically evaluated the biological effects of different types of conjugated polymers on macrophages and dendritic cells (DCs), which play critical roles in the innate and adaptive immune systems, respectively. While naked poly-(3,4-ethylenedioxythiophene):poly(4-styrenesulfonate) (PEDOT:PSS) exhibits a high level of cytotoxicity, polyethylene glycol (PEG) modified PEDOT:PSS (PEDOT:PSS-PEG) shows greatly reduced toxicity to various types of cells. To our surprise, PEGylation of PEDOT:PSS could obviously enhance the cellular uptake of these nanoparticles, leading to subsequent immune stimulations of both macrophages and DCs. In contrast, another type of conjugated polymer, polypyrrole (PPy), is found to be an inert material with neither significant cytotoxicity nor noticeable immune-stimulation activity. Interestingly, utilizing ovalbumin (OVA) as a model antigen, it is further uncovered in our ex vivo experiment that PEDOT:PSS-PEG may serve as an adjuvant to greatly enhance the immunogenicity of OVA upon simple mixing. Our study on the one hand suggests the promise of developing novel nano-adjuvants based on conjugated polymers, and on the other hand highlights the importance of careful evaluations of the impacts of any new nanomaterials developed for nanomedicine on the immune systems.

A dendritic cell (DC) vaccine, which is based on efficient antigen delivery into DCs and migration of antigen-pulsed DCs to draining lymph nodes after vaccination, is an effective strategy in initiating CD8+ T cell immunity for immunotherapy. Herein, antigen-loaded upconversion nanoparticles (UCNPs) are used to label and stimulate DCs, which could be precisely tracked after being injected into animals and induce an antigen-specific immune response. It is discovered that a model antigen, ovalbumin (OVA), could be adsorbed on the surface of dual-polymer-coated UCNPs via electrostatic interaction, forming nanoparticle–antigen complexes, which are efficiently engulfed by DCs and induce DC maturation and cytokine release. Highly sensitive in vivo upconversion luminescence (UCL) imaging of nanoparticle-labeled DCs is successfully carried out, observing the homing of DCs to draining lymph nodes after injection. In addition, strong antigen-specific immune responses including enhanced T cell proliferation, interferon gamma (IFN-γ) production, and cytotoxic T lymphocyte (CTL)-mediated responses are induced by a nanoparticle-pulsed DC vaccine, which is promising for DC-based immunotherapy potentially against cancer.Keywords: DC vaccine; immunotherapy; sensitive tracking; UCNP;

Because circulating tumor cells (CTCs) have been proven to be an important clue of the tumor metastasis, their detection thus plays a pivotal role in the diagnosis and prognosis of cancer. Herein, we fabricate an electrochemical sensor by directly conjugating two cell-specific aptamers, TLS1c and TLS11a, which specifically recognize MEAR cancer cells, to the surface of a glassy carbon electrode (GCE) via the formation of amide bonds. The two aptamers are simultaneously conjugated to the GCE surface via precisely controlled linkers: TLS1c through a flexible linker (a single-stranded DNA T15; ss-TLS1c) and TLS11a through a rigid linker (a double-stranded DNA T15/A15; ds-TLS11a). It is found that such ss-TLS1c/ds-TLS11a dual-modified GCEs show greatly improved sensitivity in comparison with those modified with a single type of aptamer alone or ds-TLS1c/ds-TLS11a with both rigid linkers, suggesting that our optimized, rationally designed electrode–aptamer biosensing interface may enable better recognition and thus more sensitive detection of tumor cells. Through the utilization of this dual-aptamer-modified GCE, as few as a single MEAR cell in 109 whole blood cells can be successfully detected with a linear range of 1–14 MEAR cells. Our work demonstrates a rather simple yet well-designed and ultrasensitive tumor cell detection method based on the cell-specific aptamer-modified GCE, showing a promising potential for further CTC-related clinical applications.Keywords: biosensing interface; cell-specific aptamer; chemically modified electrode; circulating tumor cells; electrochemical detection;

Given the complexity of tumors, several nanomaterial-based treatment modalities like chemotherapy (CT), photodynamic therapy (PDT) and photothermal therapy (PTT) have been developed for combating cancers. However, it is still unclear which strategy is better or how to select optimal approaches for combination treatment since each strategy has been investigated under different conditions. Inspired by its good payload capacity and unique near-infrared absorption, reduced graphene oxide (rGO) was selected in this study as the carrier for loading of doxorubicin (DOX), a chemotherapy drug, and chlorin e6 (Ce6), a photosensitizer. The therapeutic efficacies of PTT, CT and PDT were systematically investigated in vitro using 2D culture and multicellular tumor spheroid (3D) models. Interestingly, while all three types of therapies delivered by rGO appeared to be effective in the conventional 2D cell culture model, only PTT but not CT and PDT showed great treatment efficacy in the 3D tumor spheroid model at the tested concentrations. Such a difference is due to the fact that heat diffusion is much more efficient than the diffusion of therapeutic molecules inside the tumor. Furthermore, in vivo evidence also confirmed the unique advantage of PTT compared to the other two treatment modalities using the TdT-mediated dUTP nick end labeling (TUNEL) staining assay. This study highlights the unique advantages of nanomedicine-based photothermal therapy among these three modalities in cancer treatment in terms of killing tumor cells located far from tumor blood vessels.