Nanoparticle/Engineered Bacteria Based Triple-Strategy Delivery System for Enhanced Hepatocellular Carcinoma Cancer Therapy.

Background: New treatment modalities for hepatocellular carcinoma (HCC) are desperately critically needed, given the lack of specificity, severe side effects, and drug resistance with single chemotherapy. Engineered bacteria can target and accumulate in tumor tissues, induce an immune response, and act as drug delivery vehicles. However, conventional bacterial therapy has limitations, such as drug loading capacity and difficult cargo release, resulting in inadequate therapeutic outcomes. Synthetic biotechnology can enhance the precision and efficacy of bacteria-based delivery systems. This enables the selective release of therapeutic payloads in vivo. Methods: In this study, we constructed a non-pathogenic Escherichia coli (E. coli) with a synchronized lysis circuit as both a drug/gene delivery vehicle and an in-situ (hepatitis B surface antigen) Ag (ASEc) producer. Polyethylene glycol (CHO-PEG2000-CHO)-poly(ethyleneimine) (PEI25k)-citraconic anhydride (CA)-doxorubicin (DOX) nanoparticles loaded with plasmid encoded human sulfatase 1 (hsulf-1) enzyme (PNPs) were anchored on the surface of ASEc (ASEc@PNPs). The composites were synthesized and characterized. The in vitro and in vivo anti-tumor effect of ASEc@PNPs was tested in HepG2 cell lines and a mouse subcutaneous tumor model. Results: The results demonstrated that upon intravenous injection into tumor-bearing mice, ASEc can actively target and colonise tumor sites. The lytic genes to achieve blast and concentrated release of Ag significantly increased cytokine secretion and the intratumoral infiltration of CD4/CD8+T cells, initiated a specific immune response. Simultaneously, the PNPs system releases hsulf-1 and DOX into the tumor cell resulting in rapid tumor regression and metastasis prevention. Conclusion: The novel drug delivery system significantly suppressed HCC in vivo with reduced side effects, indicating a potential strategy for clinical HCC therapy.

A Novel CaCu-Metal-Organic-Framework Based Multimodal Treatment Platform for Enhanced Synergistic Therapy of Hepatocellular Carcinoma.

Metal ions have attracted a lot of interest in antitumor therapy due to their unique mechanism of action. However, multiple death mechanisms associate with metal ions to synergistic antitumors have few studies mainly due to the serious challenges in designing and building metal-associated multimodal treatment platforms. Hence, a series of glutathione-activatable CaCu-based metal-organic-frameworks loaded with doxorubicin and ovalbumin are successfully designed and synthesized with an "all in one" strategy, which is modified by galactosamine-linked hyaluronic acid to prepare multimodal treatment platform (SCC/DOX@OVA-HG) for targeted delivery and synergistic antitumor therapy. SCC/DOX@OVA-HG can be rapidly degraded by the overexpressed glutathione and then releases the "cargoes" in the tumor microenvironment. The released Cu+ efficiently catalyzes H2O2 to produce highly toxic ROS for CDT, and the up-regulation of calcium ion concentration in tumor cells induced by the released Ca2+ enables calcium overload therapy, which synergically enhances the metal-related death pattern. Meanwhile, OVA combined with Ca2+/Cu2+ further activates macrophages into an M1-like phenotype to accelerate tumor cell death through immunotherapy. Besides, the released DOX can also insert into the DNA double helix for chemotherapy. Consequently, the developed SCC/DOX@OVA-HG reveals significantly improved antitumor efficacy through a multimodal synergistic therapy of chemotherapy, chemodynamic therapy, calcium overload, and immunotherapy.

Hollow MIL-125 Nanoparticles Loading Doxorubicin Prodrug and 3-Methyladenine for Reversal of Tumor Multidrug Resistance.

Multidrug resistance (MDR) is a key factor in chemotherapy failure and tumor recurrence. The inhibition of drug efflux and autophagy play important roles in MDR therapy. Herein, a multifunctional delivery system (HA-MIL-125@DVMA) was prepared for synergistically reverse tumor MDR. Tumor-targeted hollow MIL-125-Ti nanoparticles were used to load the doxorubicin-vitamin E succinate (DV) prodrug and 3-methyladenine (3-MA) to enhance reverse MDR effects. The pH-sensitive DV can kill tumor cells and inhibit P-gp-mediated drug efflux, and 3-MA can inhibit autophagy. HA-MIL-125@DVMA had uniformly distributed particle size and high drug-load content. The nanoparticles could effectively release the drugs into tumor microenvironment due to the rapid hydrazone bond-breaking under low pH conditions, resulting in a high cumulative release rate. In in vitro cellular experiments, the accumulation of HA-MIL-125@DVMA and HA-MIL-125@DV in MCF-7/ADR cells was significantly higher than that in the control groups. Moreover, the nanoparticles significantly inhibited drug efflux in the cells, ensuring the accumulation of the drugs in cell cytoplasm and causing drug-resistant cells' death. Importantly, HA-MIL-125@DVMA effectively inhibited tumor growth without changes in body weight in tumor-bearing mice. In summary, the combination of the acid-sensitive prodrug DV and autophagy inhibitor 3-MA in a HA-MIL-125 nanocarrier can enhance the antitumor effect and reverse tumor MDR.

GSH-Activatable Metal-Phenolic Networks for Photothermal-Enhanced Chemotherapy and Chemodynamic Therapy.

Chemotherapy (CT) plays an important role in the antitumor process, but the unsatisfactory therapeutic efficacy and the obvious toxic side effects of CT seriously restrict its application. To overcome the limitations of CT, the strategy of chemotherapy enhanced by chemodynamic therapy (CDT) and photothermal therapy (PTT) has been considered a promising approach to improve the anticancer effect. Herein, a novel GSH-activatable Cu2+-Quercetin network (QC) was synthesized via a convenient strategy to load Au nanoparticles (NPs) and DOX, named QCDA, for the synergistic therapy of CT/CDT/PTT. The results showed that QCDA exhibited GSH-sensitive degradation and "cargos" release in cancer cells, and then PTT and CDT caused by Au NPs and Cu+ significantly enhanced the CT effect of DOX and Quercetin on anticancer. More importantly, the PTT and depleted GSH accelerated the Fenton-like ionization process resulting in facilitating the CDT efficiency. Collectively, the multi-mode synergistic strategy of CT/CDT/PTT, which showed an excellent therapeutic effect, maybe a potential therapeutic pathway for anticancer.

A Purposefully Designed pH/GSH-Responsive MnFe-Based Metal-Organic Frameworks as Cascade Nanoreactor for Enhanced Chemo-Chemodynamic-Starvation Synergistic Therapy.

Metal-organic frameworks (MOFs) have emerged as promising novel therapeutics for treating malignancies due to their tunable porosity, biocompatibility, and modularity to functionalize with various chemotherapeutics drugs. However, the design and synthesis of dual-stimuli responsive MOFs for controlled drug release in tumor microenvironments are vitally essential but still challenging. Meanwhile, the catalytic effect of metal ions selection and ratio optimization in MOFs for enhanced chemodynamic therapy (CDT) is relatively unexplored. Herein, a series of MnFe-based MOFs with pH/glutathione (GSH)-sensitivity are synthesized and then combined with gold nanoparticles (Au NPs) and cisplatin prodrugs (DSCP) as a cascade nanoreactor (SMnFeCGH) for chemo-chemodynamic-starvation synergistic therapy. H+ and GSH can specifically activate the optimal SMnFeCGH nanoparticles in cancer cells to release Mn2+/4+ /Fe2+/3+ , Au NPs, and DSCP rapidly. The optimal ratio of Mn/Fe shows excellent H2 O2 decomposition efficiency for accelerating CDT. Au NPs can cut off the energy supply to cancer cells for starvation therapy and strengthen CDT by providing large amounts of H2 O2 . Then H2 O2 is catalyzed by Mn2+ /Fe2+ to generate highly toxic •OH with the depletion of GSH. Meanwhile, the reduced DSCP accelerates cancer cell regression for chemotherapy. The ultrasensitivity cascade nanoreactor can enhance the anticancer therapeutic effect by combining chemotherapy, CDT, and starvation therapy.

Tumor-targeted glycogen nanoparticles loaded with hemin and glucose oxidase to promote tumor synergistic therapy.

Strategies which are used to address the low levels of intracellular hydrogen peroxide and the development of biocompatible catalysts still need to be fulfilled in tumor chemodynamic therapy. Therefore, a novel tumor-targeted glycogen-based nanoparticle system (GN/He/GOx/HA) was developed to co-deliver hemin (He) and GOx, which can self-supply glucose formed upon degradation of glycogen by α-glycosidase in the lysosome environment, in order to achieve synergistic antitumor therapy. Hyaluronic acid (HA) was selected as the outer shell to protect the activity of GOx, and to increase the uptake by tumor cells via CD44 receptor-mediated endocytosis. GN/He/GOx/HA NPs had a good stability in the blood circulation, but fast release of the therapeutic cargos upon intracellular uptake. Hemin had a cascade catalytic reaction with GOx. Furthermore, GN/He/GOx/HA NPs had the strongest cytotoxicity in Hela cells in a glucose concentration dependent manner. The NPs could efficiently produce reactive oxygen species in tumor cells, resulting in a decrease in the mitochondrial membrane potential and apoptosis of tumor cells. The in vivo results showed that the drug-loaded nanoparticles had good safety, biocompatibility, and efficacious antitumor effect. Therefore, the glycogen-based nanoparticle delivery system provides potential application for self-enhancing CDT, which can be used for effective antitumor therapy.

Engineered bacteria combined with doxorubicin nanoparticles suppress angiogenesis and metastasis in murine melanoma models.

Methods capable of distributing antitumour therapeutics uniformly throughout an entire tumour and that can suppress metastasis at the same time, would be of great significance in improving cancer treatment. Bacteria-mediated synergistic therapies have been explored for better specificity, temporal and spatial controllability, as well for providing regulation of the immune microenvironment, in order to provide improved cancer treatment. To achieve this goal, here we developed an engineered bacteria delivery system (GDOX@HSEc) using synthetic biology and interfacial chemistry technologies. The engineered bacteria were concurrently modified to express heparin sulfatase 1 (HSulf-1) inside (HSEc), to attach doxorubicin-loaded glycogen nanoparticles (GDOX NPs) on their surface. Here we demonstrate that HSEc can actively target and colonise tumour sites resulting in HSulf-1 overexpression, thereby suppressing angiogenesis and metastasis. Simultaneously, the GDOX NPs were able to penetrate into tumour cells, leading to intracellular DNA damage. Our results confirmed that a combination of biotherapy and chemotherapy using GDOX@HSEc resulted in significant melanoma suppression in murine models, with reduced side effects. This study provides a powerful platform for the simultaneous delivery of biomacromolecules and chemotherapeutic drugs to tumours, representing an innovative strategy potentially more effective in treating solid tumours. STATEMENT OF SIGNIFICANCE: An original engineered bacteria-based system (GDOX@HSEc) was developed using synthetic biology and interfacial chemistry technologies to concurrently produce naturally occurring heparin sulfatase 1 (HSulf-1) inside and anchor doxorubicin-loaded glycogen nanoparticles on the surface. GDOX@HSEc allowed for combined local delivery of chemotherapeutic agents along with the enzymes and immunostimulatory bacterial adjuvants, which resulted in a synergistic action in the inhibition of tumour growth and metastasis. The study provides a potential therapeutic approach that allows therapeutic agents to be distributed in a spatiotemporally controllable manner in tumours for combinatorial enhanced therapy.

Enzyme-loaded glycogen nanoparticles with tumor-targeting Activatable host-guest supramolecule for augmented chemodynamic therapy.

Chemodynamic therapy (CDT) has advantages in site-specific killing tumor and avoiding systemically side effect. Although numerous nano-systems have been developed to enhance the intracellular hydrogen peroxide (H2O2) for improving CDT effect, the biocompatibility of the materials limits their further biomedical applications. Herein glycogen, as a natural biological macromolecule, was used to construct a new targeted separable GOx@GF/HC nanoparticle system to deliver glucose oxidase (GOx) for CDT/starvation tumor therapy. Amination glycogen-ferrocene (GF) as a guest core and hyaluronic acid-ß-cyclodextrin (HC) as a host shell were synthesized and self-assembled through host-guest interactions to deliver GOx. After being entered into tumor cells, GOx were released to catalyze glucose to produce gluconic acid and H2O2, which in turn cut off the nutrition of tumor cells for starvation therapy and enhanced the generation of OH with ferrous ion through Fenton reaction. Furthermore, GOx@GF/HC also exhibited remarkable tumor-targeting and tumor-suppression in vivo. Therefore, the GOx@GF/HC system can exert excellent synergistic effect of CDT and starvation therapy on cancer treatment through a cascade reaction, which have some potential application for the development of CDT tumor-treatment.

Dexamethasone microspheres and celecoxib microcrystals loaded into injectable gels for enhanced knee osteoarthritis therapy.

The combination of corticosteroids and nonsteroidal anti-inflammatory drugs (NSAIDs) has been commonly used for inflammation and chronic articular pain in the clinic. Nonetheless, the long-term administration of both medications might result in osteonecrosis of the knee due to repeated injections of steroids and side effects in the gastrointestinal and cardiovascular systems. To overcome these unmet medical needs, we designed a microsphere-microcrystal-gel delivery system for intra-articular injection. Dexamethasone (DEX)-loaded microspheres (DMs) were optimized by Plackett-Burman and Taguchi orthogonal designs to extend their retention time in the knee joint. Celecoxib (CLX) microcrystals (CMs) were manufactured using an ultrasonic method to improve solubility and bioavailability. Moreover, a green solvent-free method was employed to crosslink and synthesize a novel poloxamer 407/Gantrez® S97-based gel system (GZF), which can undergo the sol-gel transition at lower concentrations. Then, DM and CM were loaded by GZF to form intra-articular injectable gels (DM/CM/Gel). The in vitro release of DEX and CLX showed a fast phase in 24 h followed by a controlled release of ∼8 d. Both blank microspheres and GZF gels displayed great biocompatibility against RAW264.7 macrophages. The most suitable dosages of 5 nM DEX and 125 nM CLX in the formulation were chosen because of their significant effects against macrophage inflammation with a lower administrative amount. An In vivo animal evaluation showed that DM/CM/Gel suppressed the release of inflammatory cytokines (TNF-α and IL-6) after 21 d of treatment. In addition, a histological evaluation revealed that DM/CM/Gel interrupted the progression of cartilage surface denudation and matrix loss. Therefore, DM/CM/Gel provides a prospective strategy for reforming traditional therapy for chronic articular disease.

Bacteria-mediated cancer therapies: opportunities and challenges.

In recent years, cancer therapy strategies utilizing live tumor-targeting bacteria have presented unique advantages. Engineered bacteria have the particular ability to distinguish tumors from normal tissues with less toxicity. Live bacteria are naturally capable of homing to tumors, resulting in high levels of local colonization because of insufficient oxygen and low pH in the tumor microenvironment. Bacteria initiate their antitumor effects by directly killing the tumor or by activating innate and adaptive antitumor immune responses. The bacterial vectors can be reprogrammed following advanced DNA synthesis, sophisticated genetic bioengineering, and biosensors to engineer microorganisms with complex functions, and then produce and deliver anticancer agents based on clinical needs. However, because of the lack of knowledge on the mechanisms and side effects of microbial cancer therapy, developing such smart microorganisms to treat or prevent cancer remains a significant challenge. In this review, we summarized the potential, status, opportunities and challenges of this growing field. We illustrated the mechanism of tumor regression induced by engineered bacteria and discussed the recent advances in the application of bacteria-mediated cancer therapy to improve efficacy, safety and drug delivery. Finally, we shared our insights into the future directions of tumor-targeting bacteria in cancer therapy.

Fabrication and characterization of DDAB/PLA-alginate composite microcapsules as single-shot vaccine.

The most effective method to reduce chronic hepatitis B virus infection is the universal implementation of vaccination. The commercial aluminum-based vaccines need multiple-injection protocols for complete protection resulting in poor compliance in developing countries. It is necessary to develop single-shot vaccine formulations. In this study, novel antigen-loaded DDAB/PLA (didodecyldimethylammonium bromide/poly(lactic acid)) nanoparticles (NPs)-alginate composite microcapsules were developed as a single-shot vaccine. The hepatitis B surface antigen (HBsAg)-loaded DDAB/PLA NPs were successfully encapsulated into alginate microcapsules by a modified spray-solidification technique. The response surface method was applied to optimize the preparation parameters employing encapsulation efficiency of HBsAg and particle size of microcapsules as response variables. The antigen-loaded DDAB/PLA NPs-alginate composite microcapsules were prepared under these optimal conditions: the size of composite microcapsules was 24.25 µm, the Span value was 1.627, and the encapsulation efficiency of HBsAg was 68.4%. The obtained microcapsules were spherical gel microparticles with excellent dispersity and narrow size distributions. In vitro release profile indicated a slow release rate of encapsulated HBsAg especially in phosphate buffered saline solution. The microcapsules showed little toxicity in vivo. This vaccine delivery system could induce stronger immune responses by a single shot, which exhibited much higher cytokine secretion levels closely related to cellular immunity and comparable IgG titers to the traditional aluminum-adjuvanted vaccine with three shots.