Immunoadjuvant-Modified Rhodobacter sphaeroides Potentiate Cancer Photothermal Immunotherapy.

Photothermal immunotherapy has become a promising strategy for tumor treatment. However, the intrinsic drawbacks like light instability, poor immunoadjuvant effect, and poor accumulation of conventional inorganic or organic photothermal agents limit their further applications. Based on the superior carrying capacity and active tumor targeting property of living bacteria, an immunoadjuvant-intensified and engineered tumor-targeting bacterium was constructed to achieve effective photothermal immunotherapy. Specifically, immunoadjuvant imiquimod (R837)-loaded thermosensitive liposomes (R837@TSL) were covalently decorated onto Rhodobacter sphaeroides (R.S) to obtain nanoimmunoadjuvant-armed bacteria (R.S-R837@TSL). The intrinsic photothermal property of R.S combined R837@TSL to achieve in situ near-infrared (NIR) laser-controlled release of R837. Meanwhile, tumor immunogenic cell death (ICD) caused by photothermal effect of R.S-R837@TSL, synergizes with released immunoadjuvants to promote maturation of dendritic cells (DCs), which enhance cytotoxic T lymphocytes (CTLs) infiltration for further tumor eradication. The photosynthetic bacteria armed with immunoadjuvant-loaded liposomes provide a strategy for immunoadjuvant-enhanced cancer photothermal immunotherapy.

Intracellular Gelation-Mediated Living Bacteria for Advanced Biotherapeutics in Mouse Models.

Despite its significant potential in various disease treatments and diagnostics, microbiotherapy is consistently plagued by multiple limitations ranging from manufacturing challenges to in vivo functionality. Inspired by the strategy involving nonproliferating yet metabolically active microorganisms, we report an intracellular gelation approach that can generate a synthetic polymer network within bacterial cells to solve these challenges. Specifically, poly(ethylene glycol dimethacrylate) (PEGDA, 700 Da) monomers are introduced into the bacterial cytosol through a single cycle of freeze-thawing followed by the initiation of intracellular free radical polymerization by UV light to create a macromolecular PEGDA gel within the bacterial cytosol. The molecular crowding resulting from intracytoplasmic gelation prohibits bacterial division and confers robust resistance to simulated gastrointestinal fluids and bile acids while retaining the ability to secrete functional proteins. Biocompatibility assessments demonstrate that the nondividing gelatinized bacteria are effective in alleviating systemic inflammation triggered by intravenous Escherichia coli injection. Furthermore, the therapeutic efficacy of gelatinized Lactobacillus rhamnosus in colitis mice provides additional support for this approach. Collectively, intracellular gelation indicates a universal strategy to manufacture next-generation live biotherapeutics for advanced microbiotherapy.

Autonomously Assembled Living Capsules by Microbial Coculture for Enhanced Bacteriotherapy of Inflammatory Bowel Disease.

Microorganism-mediated self-assembling of living formulations holds great promise for disease therapy. Here, we constructed a prebiotic-probiotic living capsule (PPLC) by coculturing probiotics (EcN) with Gluconacetobacter xylinus (G. xylinus) in a prebiotic-containing fermentation broth. Through shaking the culture, G. xylinus secretes cellulose fibrils that can spontaneously encapsulate EcN to form microcapsules under shear forces. Additionally, the prebiotic present in the fermentation broth is incorporated into the bacterial cellulose network through van der Waals forces and hydrogen bonding. Afterward, the microcapsules were transferred to a selective LB medium, which facilitated the colonization of dense probiotic colonies within them. The in vivo study demonstrated that PPLC-containing dense colonies of EcN can antagonize intestinal pathogens and restore microbiota homeostasis by showing excellent therapeutic performance in treating enteritis mice. The in situ self-assembly of probiotics and prebiotics-based living materials provides a promising platform for the treatment of inflammatory bowel disease.

Epigenetics-Based Tumor Cells Pyroptosis for Enhancing the Immunological Effect of Chemotherapeutic Nanocarriers.

Pyroptosis is a lytic and inflammatory form of programmed cell death and could be induced by chemotherapy drugs via caspase-3 mediation. However, the key protein gasdermin E (GSDME, translated by the DFNA5 gene) during the caspase-3-mediated pyroptosis process is absent in most tumor cells because of the hypermethylation of DFNA5 (deafness autosomal dominant 5) gene. Here, we develop a strategy of combining decitabine (DAC) with chemotherapy nanodrugs to trigger pyroptosis of tumor cells by epigenetics, further enhancing the immunological effect of chemotherapy. DAC is pre-performed with specific tumor-bearing mice for demethylation of the DFNA5 gene in tumor cells. Subsequently, a commonly used tumor-targeting nanoliposome loaded with cisplatin (LipoDDP) is used to administrate drugs for activating the caspase-3 pathway in tumor cells and trigger pyroptosis. Experiments demonstrate that the reversal of GSDME silencing in tumor cells is achieved and facilitates the occurrence of pyroptosis. According to the anti-tumor activities, anti-metastasis results, and inhibition of recurrence, this pyroptosis-based chemotherapy strategy enhances immunological effects of chemotherapy and also provides an important insight into tumor immunotherapy.

PD-1 Blockade for Improving the Antitumor Efficiency of Polymer-Doxorubicin Nanoprodrug.

Chemotherapy is well recognized to induce immune responses during some chemotherapeutic drugs-mediated tumor eradication. Here, a strategy involving blocking programmed cell death protein 1 (PD-1) to enhance the chemotherapeutic effect of a doxorubicin nanoprodrug HA-Psi-DOX is proposed and the synergetic mechanism between them is further studied. The nanoprodrugs are fabricated by conjugating doxorubicin (DOX) to an anionic polymer hyaluronic acid (HA) via a tumor overexpressed matrix metalloproteinase sensitive peptide (CPLGLAGG) for tumor targeting and enzyme-activated drug release. Once accumulated at the tumor site, the nanoprodrug can be activated to release antitumor drug by tumor overexpressed MMP-2. It is found that HA-Psi-DOX nanoparticles can kill tumor cells effectively and initiate an antitumor immune response, leading to the upregulation of interferon-γ. This cytokine promotes the expression of programmed cell death protein-ligand 1 (PD-L1) on tumor cells, which will cause immunosuppression after interacting with PD-1 on the surface of lymphocytes. The results suggest that the therapeutic efficiency of HA-Psi-DOX nanoparticles is significantly improved when combined with checkpoint inhibitors anti-PD-1 antibody (α-PD1) due to the neutralization of immunosuppression by blocking the interaction between PD-L1 and PD-1. This therapeutic system by combining chemotherapy and immunotherapy further increases the link between conventional tumor therapies and immunotherapy.

Dual-Stage Light Amplified Photodynamic Therapy against Hypoxic Tumor Based on an O2 Self-Sufficient Nanoplatform.

Tumor hypoxia severely limits the efficacy of traditional photodynamic therapy (PDT). Here, a liposome-based nanoparticle (designated as LipoMB/CaO2 ) with O2 self-sufficient property for dual-stage light-driven PDT is demonstrated to address this problem. Through a short time irradiation, 1 O2 activated by the photosensitizer methylene blue (MB) can induce lipid peroxidation to break the liposome, and enlarge the contact area of CaO2 with H2 O, resulting in accelerated O2 production. Accelerated O2 level further regulates hypoxic tumor microenvironment and in turn improves 1 O2 generation by MB under another long time irradiation. In vitro and in vivo experiments also demonstrate the superior competence of LipoMB/CaO2 to alleviate tumor hypoxia, suppress tumor growth and antitumor metastasis with low side-effect. The O2 self-sufficient LipoMB/CaO2 nanoplatform with dual-stage light manipulation is a successful attempt for PDT against hypoxic tumor.

Advances in Peptide Functionalization on Mesoporous Silica Nanoparticles for Controlled Drug Release.

During the last decade, using versatile, promising, and fascinating mesoporous silica nanoparticles (MSNs) as site-specific and stimuli-responsive drug delivery systems (DDSs) has received concentrated research interest. As one of the most attractive surface modification units, peptides have inherent bioactivity, biodegradability and biocompatibility. Recent progresses in the utilization of versatile peptides for surface functionalization of MSNs to achieve cell-specific targeting, fluorescence imaging, and intracellular diagnosis and treatment of tumors are summarized in this review. The various functional peptides decorated on the MSNs are introduced and classified into three types, including targeting peptides, stimuli-responsive peptides and multifunctional chimeric peptides. The limitations and challenges of peptide modified MSNs and their potential applications are further discussed.

Theranostic GO-based nanohybrid for tumor induced imaging and potential combinational tumor therapy.

Graphene oxide (GO)-based theranostic nanohybrid is designed for tumor induced imaging and potential combinational tumor therapy. The anti-tumor drug, Doxorubicin (DOX) is chemically conjugated to the poly(ethylenimine)-co-poly(ethylene glycol) (PEI-PEG) grafted GO via a MMP2-cleavable PLGLAG peptide linkage. The therapeutic efficacy of DOX is chemically locked and its intrinsic fluorescence is quenched by GO under normal physiological condition. Once stimulated by the MMP2 enzyme over-expressed in tumor tissues, the resulting peptide cleavage permits the unloading of DOX for tumor therapy and concurrent fluorescence recovery of DOX for in situ tumor cell imaging. Attractively, this PEI-bearing nanohybrid can mediate efficient DNA transfection and shows great potential for combinational drug/gene therapy. This tumor induced imaging and potential combinational therapy will open a window for tumor treatment by offering a unique theranostic approach through merging the diagnostic capability and pathology-responsive therapeutic function.

Erythrocyte-Mimicking Nanovesicle Targeting Porphyromonas gingivalis for Periodontitis.

Porphyromonas gingivalis has been demonstrated to have the strongest association with periodontitis. Within the host, P. gingivalis relies on acquiring iron and heme through the aggregation and lysis of erythrocytes, which are important factors in the growth and virulence of P. gingivalis. Additionally, the excess obtained heme is deposited on the surface of P. gingivalis, protecting the cells from oxidative damage. Based on these biological properties of the interaction between P. gingivalis and erythrocytes, this study developed an erythrocyte membrane nanovesicle loaded with gallium porphyrins to mimic erythrocytes. The nanovesicle can target and adhere with P. gingivalis precisely, being lysed and utilized by P. gingivalis as erythrocytes. Ingested gallium porphyrin replaces iron porphyrin in P. gingivalis, causing intracellular metabolic disruption. Deposited porphyrin generates a large amount of reactive oxygen species (ROS) under blue light, causing oxidative damage, and its lethality is enhanced by bacterial metabolic disruption, synergistically killing P. gingivalis. Our results demonstrate that this strategy can target and inhibit P. gingivalis, reduce its invasion of epithelial cells, and alleviate the progression of periodontitis.

In Situ Sprayed Biotherapeutic Gel Containing Stable Microbial Communities for Efficient Anti-Infection Treatment.

Systematic administration of antibiotics to treat infections often leads to the rapid evolution and spread of multidrug-resistant bacteria. Here, an in situ-formed biotherapeutic gel that controls multidrug-resistant bacterial infections and accelerates wound healing is reported. This biotherapeutic gel is constructed by incorporating stable microbial communities (kombucha) capable of producing antimicrobial substances and organic acids into thermosensitive Pluronic F127 (polyethylene-polypropylene glycol) solutions. Furthermore, it is found that the stable microbial communities-based biotherapeutic gel possesses a broad antimicrobial spectrum and strong antibacterial effects in diverse pathogenic bacteria-derived xenograft infection models, as well as in patient-derived multidrug-resistant bacterial xenograft infection models. The biotherapeutic gel system considerably outperforms the commercial broad-spectrum antibacterial gel (0.1% polyaminopropyl biguanide) in pathogen removal and infected wound healing. Collectively, this biotherapeutic strategy of exploiting stable symbiotic consortiums to repel pathogens provides a paradigm for developing efficient antibacterial biomaterials and overcomes the failure of antibiotics to treat multidrug-resistant bacterial infections.

Polymeric nano-system for macrophage reprogramming and intracellular MRSA eradication.

Intracellular Methicillin-Resistant Staphylococcus aureus (MRSA) remains a major factor of refractory and recurrent infections, which cannot be well addressed by antibiotic therapy. Here, we design a cellular infectious microenvironment-activatable polymeric nano-system to mediate targeted intracellular drug delivery for macrophage reprogramming and intracellular MRSA eradication. The polymeric nano-system is composed of a ferrocene-decorated polymeric nanovesicle formulated from poly(ferrocenemethyl methacrylate)-block-poly(2-methacryloyloxyethyl phosphorylcholine) (PFMMA-b-PMPC) copolymer with co-encapsulation of clofazimine (CFZ) and interferon-γ (IFN-γ). The cellular-targeting PMPC motifs render specific internalization by macrophages and allow efficient intracellular accumulation. Following the internalization, the ferrocene-derived polymer backbone sequentially undergoes hydrophobic-to-hydrophilic transition, charge reversal and Fe release in response to intracellular hydrogen peroxide over-produced upon infection, eventually triggering endosomal escape and on-site cytosolic drug delivery. The released IFN-γ reverses the immunosuppressive status of infected macrophages by reprogramming anti-inflammatory M2 to pro-inflammatory M1 phenotype. Meanwhile, intracellular Fe2+-mediated Fenton reaction together with antibiotic CFZ contributes to increased intracellular hydroxyl radical (•OH) generation. Ultimately, the nano-system achieves robust potency in ablating intracellular MRSA and antibiotic-tolerant persisters by synchronous immune modulation and efficient •OH killing, providing an innovative train of thought for intracellular MRSA control.

Deep-penetration functionalized cuttlefish ink nanoparticles for combating wound infections with synergetic photothermal-immunologic therapy.

The challenge of wound infections post-surgery and open trauma caused by multidrug-resistant bacteria poses a constant threat to clinical treatment. As a promising antimicrobial treatment, photothermal therapy can effectively resolve the problem of drug resistance in conventional antibiotic antimicrobial therapy. Here, we report a deep-penetration functionalized cuttlefish ink nanoparticle (CINP) for photothermal and immunological therapy of wound infections. CINP is decorated with zwitterionic polymer (ZP, namely sulfobetaine methacrylate-methacrylate copolymer) to form CINP@ZP nanoparticles. Natural CINP is found to not only exhibit photothermal destruction of methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli), but also trigger macrophages-related innate immunity and enhance their antibacterial functions. The ZP coating on the surface of CINP enables nanoparticles to penetrate into deeply infected wound environment. In addition, CINP@ZP is further integrated into the thermosensitive Pluronic F127 gel (CINP@ZP-F127). After in situ spraying gel, CINP@ZP-F127 is also documented notable antibacterial effects in mice wound models infected with MRSA and E. coli. Collectively, this approach combining of photothermal therapy with immunotherapy can promote delivery efficiency of nanoparticles to the deep foci of infective wounds, and effectively eliminate wound infections.

PEG-stabilized micellar system with positively charged polyester core for fast pH-responsive drug release.

PURPOSE: To design functional drug carriers for fast pH-responsive drug release. METHODS: Functional diblock terpolymers of monomethoxy poly(ethylene glycol)-block- copoly(6,14-dimethyl-1,3,9,11-tetraoxa-6,14-diaza-cyclohexadecane-2,10-dione-co-ε-caprolactone) [mPEG-b-poly(ADMC-co-CL)] were fabricated via biosynthetic pathway. The self-assembled nanosphere and drug-loaded micelles of the copolymers were further prepared by dialysis method. The pH-tunable morphology variation and drug release pattern were observed at different pH. RESULTS: A collection of three PEGylated terpolymers with varied compositions in poly(ADMC-co-CL) block was designed with high cell-biocompatibility. The copolymers could readily self-assemble into nanoscale micelles (~ 100 nm) in aqueous medium and exhibit high stability over 80-h incubation in different mediums including deionized water, neutral NaCl solution, and heparin sodium solution. Due to the protonation-deprotonation of tertiary amine groups in ADMC units, acid-induced structural deformation of micelles was disclosed in terms of the variation in CAC value and hydrodynamic size at different pH. Drug loading efficiency was comparable to that of reported PEG-polyester micelles with specifically designed structures purposed for drug-loading improvement. Remarkably accelerated drug release triggered by acidity was distinctly detected for ibuprofen-loaded mPEG-b-poly(ADMC-co-CL) micelle system, suggesting a fast pH-responsive characteristic. CONCLUSION: Functional PEG-stabilized micellar carriers with positively charged polyester core were successfully developed for fast pH-responsive drug release.

Hierarchy-Assembled Dual Probiotics System Ameliorates Cholestatic Drug-Induced Liver Injury via Gut-Liver Axis Modulation.

Cholestatic drug-induced liver injury (DILI) induced by drugs or other xenobiotics is a severe and even fatal clinical syndrome. Here, living materials of hierarchy-assembled dual probiotics system are fabricated by sequentially encapsulating probiotic Lactobacillus delbrueckii subsp. bulgaricus (LDB) and Lactobacillus rhamnosus GG (LGG) into Ca2+ -complexed polymer microspheres for effective prevention of cholestatic DILI. Upon entering intestinal tract of the constructed living materials, LGG is released because of pH-triggered dissolution of outer enteric polymer coating. The released LGG can inhibit hepatic bile acids (BAs) synthesis by activating intestinal farnesoid X receptor-fibroblast growth factor 15（FGF-15) signaling pathway. BAs excretion is also facilitated by LGG through increasing the abundance of bile salt hydrolase (BSH)-active gut commensal bacteria. Furthermore, exposed positively-charged chitosan shell can absorb the excessive BAs via electrostatic interaction, which leads to steady BAs fixation by the imprisoned LDB, decreasing the total BAs amounts in enterohepatic circulation. Together, the fabricated living materials, obtained here, can effectively prevent cholestatic DILI through dredging cholestasis via gut-liver axis modulation. The therapeutic effect is demonstrated in α-naphthylisothiocyanate and clinical antiepileptic drug valproate acid-induced cholestatic DILI mouse models, which reveal the great potential for effective cholestatic DILI management.

Tumor-triggered targeting ammonium bicarbonate liposomes for tumor multimodal therapy.

Tumor-triggered targeting ammonium bicarbonate (TTABC) liposomes were proposed to improve the uptake of ammonium bicarbonate (ABC) liposomes in tumor cells and retain their long circulation in vivo in our previous study. However, it must be solved how to precisely release the loaded drugs of the TTABC liposomes into tumor cells. In addition, synergistic multimodal therapy could result in better tumor treatment outcomes than monomodal chemotherapy. In the research, we prepared indocyanine green (ICG) and doxorubicin (DOX) encapsulated TTABC liposomes (ICG&DOX@TTABC) to achieve near-infrared (NIR) light-controlled chemo/photothermal/photodynamic multimodal therapy guided by fluorescence and photothermal imaging. In vitro and vivo studies show that ICG&DOX@TTABC can specifically accumulate in tumor tissues, effectively transform NIR light into local thermo-therapy, and have excellent anti-tumor ability without obvious side effects. ICG&DOX@TTABC could be promising for fluorescence and photothermal imaging-guided chemo/photothermal/photodynamic tumor treatment.

Biomaterial-mediated modulation of oral microbiota synergizes with PD-1 blockade in mice with oral squamous cell carcinoma.

Because a host's immune system is affected by host-microbiota interactions, means of modulating the microbiota could be leveraged to augment the effectiveness of cancer therapies. Here we report that patients with oral squamous cell carcinoma (OSCC) whose tumours contained higher levels of bacteria of the genus Peptostreptococcus had higher probability of long-term survival. We then show that in mice with murine OSCC tumours injected with oral microbiota from patients with OSCCs, antitumour responses were enhanced by the subcutaneous delivery of an adhesive hydrogel incorporating silver nanoparticles (which inhibited the growth of bacteria competing with Peptostreptococcus) alongside the intratumoural delivery of the bacterium P. anaerobius (which upregulated the levels of Peptostreptococcus). We also show that in mice with subcutaneous or orthotopic murine OSCC tumours, combination therapy with the two components (nanoparticle-incorporating hydrogel and exogenous P. anaerobius) synergized with checkpoint inhibition with programmed death-1. Our findings suggest that biomaterials can be designed to modulate human microbiota to augment antitumour immune responses.

Targeted delivery in breast cancer cells via iodine: nuclear localization sequence conjugate.

The nonviral vector with iodine-nuclear localization sequence (namely, NLS-I) targeting breast cancer cells was fabricated. Ternary complexes were formed via charge interactions among NLS-I peptides, PEI 1800, and DNA, and we investigated their cellular internalization, nuclear accumulation as well as transfection efficiency. All the experiments were assessed by employing MCF-7 cells that express sodium/iodide symporter and HeLa cells that lack the expression of the symporter. In MCF-7 cells, cell internalization and nuclear accumulation of NLS-I was markedly increased compared to that in NLS. In addition, compared to that of the PEI1800/DNA complex, PEI1800/DNA/NLS-I complexes exhibited much enhanced luciferase reporter gene expression by up to 130-fold. By contrast, in HeLa cells, the evident improvements of cellular internalization, nuclear accumulation, and transfection efficiency by NLS-I were not observed. This study demonstrates an alternative method to construct a nonviral delivery system for targeted gene transfer into breast cancer cells.

Hyperbranched polycarbonate-based multimolecular micelle with enhanced stability and loading efficiency.

We herein develop a facile catalyst-free method to prepare hyperbranched hydroxyl-enriched aliphatic polycarbonate according to SCROP strategy. PEG-attached multiarm hyperbranched copolymer HEHDO-star-mPEG was further designed. It was found that HEHDO-star-mPEG can self-assemble into supramolecular multimolecular micelles in water. HEHDO-star-mPEG micelle showed excellent stability with respect to micellar size upon dilution, and displayed good cell-biocompatibility. An anticancer drug of doxorubicin with hydrogen-bonding functionality was incorporated into obtained micelles to establish a drug delivery system model. A high drug-loading content as well as sustained release pattern for HEHDO-star-mPEG based delivery system was achieved.

Porphyrin and galactosyl conjugated micelles for targeting photodynamic therapy.

PURPOSE: To study the targeting and photodynamic therapy efficiency of porphyrin and galactosyl conjugated micelles based on amphiphilic copolymer galactosyl and mono-aminoporphyrin (APP) incoporated poly(2-aminoethyl methacrylate)-polycaprolactone (Gal-APP-PAEMA-PCL). METHODS: Poly(2-aminoethyl methacrylate)-polycaprolactone (PAEMA-PCL) was synthesized by the combination of ring opening polymerization and reversible addition-fragmentation chain transfer (RAFT) polymerization, and then Gal-APP-PAEMA-PCL was obtained after conjugation of lactobionic acid and 5-(4-aminophenyl)-10,15,20-triphenylporphyrin (APP) to PAEMA-PCL. The chemical structures of the copolymers were characterized, and their biological properties were evaluated in human laryngeal carcinoma (HEp2) and human hepatocellular liver carcinoma (HepG2) cells. RESULTS: Both APP-PAEMA-PCL and Gal-APP-PAEMA-PCL did not exhibit dark cytotoxicity to HEp2 cells and HepG2 cells. However, Gal-APP-PAEMA-PCL was taken up selectively by HepG2 cells and had the higher phototoxicity effect. Both polymers preferentially localized within cellular vesicles that correlated to the lysosomes. CONCLUSIONS: The results indicated that porphyrin and galactosyl conjugated polymer micelles exhibited higher targeting and photodynamic therapy efficacy in HepG2 cells than in HEp2 cells.

Injectable hydrogel helps bone marrow-derived mononuclear cells restore infarcted myocardium.

BACKGROUNDS: Experimental and clinical studies have suggested that cell implantation could improve cardiac function after myocardial infarction (MI). However, this technique was limited by decreased engraftment and survival of transplanted cells within the ischemic tissue. The present study was performed to investigate whether implantation of bone marrow-derived mononuclear cells (BMMNCs) encapsulated in hydrogel could increase cell engraftment and help to restore cardiac function of MI rabbits. METHODS: MI was induced in rabbits by coronary artery ligation. One week later, cell culture medium, Dex-PCL-HEMA/PNIPAAm hydrogel, BMMNCs in medium or BMMNCs in hydrogel were injected into the infarcted area of the left ventricle (LV). RESULTS: Increased cell engraftment was observed 48 h after injection when cells were encapsulated in hydrogel; 30 days after treatment, echocardiographic studies showed that injection of BMMNCs in hydrogel preserved LV ejection fraction and attenuated LV dilatation compared with other groups. Histological analysis indicated that injection of BMMNCs in hydrogel enhanced neovascular formation and prevented scar expansion compared with the other groups. CONCLUSION: Injection of hydrogel-encapsulated BMMNCs increased cell engraftment and improved LV function; this technique may serve as an effective approach to restore infarcted myocardium.