Research on the loading and release kinetics of the vincristine sulfate liposomes and its anti-breast cancer activity.

Vincristine (VCR), as a cytotoxic drug, is used clinically to treat acute lymphatic leukemia and breast cancer, and commonly used clinically as vincristine sulfate (VCRS). However, its clinical use is limited by unpredictable pharmacologic characteristics, a narrow therapeutic index, and neurotoxicity. The pH gradient method was used for active drug loading of VCRS, and the process route mainly includes the preparation of blank liposomes and drug-loaded liposomes. VCRS liposomes had suitable particle size, high encapsulation efficiency and good stability. The loading and release kinetics of VCRS liposomes were explored. By calculating the changes of encapsulation efficiency with time at different temperatures, it was confirmed that the drug-loading process of liposomes exhibited a first-order kinetic feature, and the activation energy required for the reaction was determined as 20.6 kcal/mol. The release behavior at different pH was also investigated, and it was demonstrated that the release behavior conformed to the first-order model, suggesting that the release mechanism of VCRS was simple transmembrane diffusion. VCRS liposomes also enhanced in vitro and in vivo antitumor activity. Thus, VCRS liposomes showed great potential for VCRS delivery, and the loading and release kinetics were well researched to provide a reference for investigating active drug loading liposomes.

Dual-Targeted Self-Adjuvant Heterocyclic Lipidoid@Polyester Hybrid Nanovaccines for Boosting Cancer Immunotherapy.

Tumor vaccines have demonstrated a modest response rate, primarily attributed to their inefficient delivery to dendritic cells (DCs), low cross-presentation, DC-intrinsic immunosuppressive signals, and an immunosuppressive tumor microenvironment (TME). Here, draining lymph node (DLN)-targeted and tumor-targeted nanovaccines were proposed to address these limitations, and heterocyclic lipidoid (A18) and polyester (BR647) were synthesized to achieve dual-targeted cancer immunotherapy. Meanwhile, oligo hyaluronic acid (HA) and DMG-PEG2000-Mannose were incorporated to prepare dual-targeted nanovaccines encapsulated with STAT3 siRNA and model antigens. The nanovaccines were designed to target the DLN and the tumor, facilitating the delivery of cargo into the cytoplasm. These dual-targeted nanovaccines improved antigen presentation and DC maturation, activated the stimulator of interferon genes (STING) pathway, enhanced the pro-apoptotic effect, and stimulated antitumor immune responses. Additionally, these dual-targeted nanovaccines overcame immunosuppressive TME, reduced immunosuppressive cells, and promoted the polarization of tumor-associated neutrophils from N2 to N1. Among the four dual-targeted nanovaccines that induced robust antitumor responses, the heterocyclic lipidoid@polyester hybrid nanovaccines (MALO@HBNS) demonstrated the most promising results. Furthermore, a combination strategy involving MALO@HBNS and an anti-PD-L1 antibody exhibited an immensely powerful anticancer role. This work introduced a dual-targeted nanovaccine platform for antitumor treatment, suggesting its potential combination with an immune checkpoint blockade as a comprehensive anticancer strategy.

Enhanced localized therapeutic precision: A face-to-face folate-targeted Cu2+-mediated nanotherapy with thermosensitive sustained-release system.

Safe and effective Cu2+ supplementation in local lesion is crucial for minimizing toxicity of DSF-based chemotherapy. Targeted delivery of Cu2+ appears more promising. Intraperitoneal chemotherapy for peritoneal carcinoma (PC) establishes "face-to-face" contact between targeted nanocarriers and tumor tissue. Herein, this study developed a biodegradable, injectable thermosensitive hydrogel that coencapsulating DSF submicroemulsion (DSF-SE) and folate-modified liposome loading glycyrrhizic acid-Cu (FCDL). FCDL acted as 'beneficial horse' to target the tumor-localized folate receptor, thus liberating Cu2+ in tumor nidus. The prepared FCDL and DSF-SE were found with uniform sizes (160.2 nm, 175.4 nm), low surface charge (-25.77 mV, -16.40 mV) and high encapsulation efficiency (97.93 %, 90.08 %). In vitro drug release profile of FCDL, DSF-SE and FCDL＆DSF-SE@G followed a sustained release pattern. And the release behavior of Cu2+ from FCDL was pH-related, i.e., Cu2+ was released faster under acidic condition. When FCDL and DSF-SE were loaded into an PLGA-PEG-PLGA-based hydrogel system, FCDL＆DSF-SE@G was formed to ensure separated delivery of Cu2+ and DSF in space but synchronized release over time. The rheology experiment showed a satisfactory gelling temperature of 32.7 °C. In vitro cytotoxicity study demonstrated that FCDL＆DSF-SE@G significantly lowered the IC50 of free Cu2+/DSF, Cu2+/DSF hydrogel and non-targeted analogue by almost 70 %, 65 % and 32 %, respectively. Accordingly, in tumor-bearing mice, FCDL＆DSF-SE@G augmented the tumor inhibition rates for the same formulations by 352 %, 145 % and 44 %, respectively. The main mechanism was attributed to higher uptake of FCDL and DSF-SE, resulting in increased Cu(DDTC)2 formation, ROS production and cell apoptosis. In conclusion, this targeted nanotherapy approach with dual-nanocarriers loaded hydrogel system, with its focus on face-to-face contact between nanocarriers and tumor tissues in the peritoneal cavity, holds significant promise for intraperitoneal chemotherapy in PC.

Two-photon fluorescence-guided precise photothermal therapy located in a single cancer cell utilizing bifunctional N-doped carbon quantum dots.

The utilization of carbon quantum dots (CQDs) for photothermal therapy has emerged as a hot research topic. However, there has been limited research on killing one single cancer cell which is critical in reducing unnecessary damage to the surrounding healthy tissues. In this work, we developed a two-photon fluorescence-guided precise photothermal therapy in a single human malignant melanoma (A375) cancer cell utilizing bifunctional N-doped CQDs. Resulting from the two-photon fluorescence of the CQDs, one single cancer cell can be located and simultaneously destroyed by the photothermal effect of the same CQDs. Specifically, the balanced two-photon absorption cross-section (7000 GM) and photoluminescence quantum yield (8.4%) of the CQDs enable the fluorescence-guided photothermal treatment to be achieved in only 5 s under the irradiation of 800 nm laser of 27.5 mW, much faster than the control experiment without the guidance of fluorescence. The heat generated by the aggregated CQDs is in sufficient amounts while being confined in a small area, as evidenced by the numerical simulations and photothermal experiments, to limit the range of thermal treatment in the cells. This work provides a new approach for realizing photothermal therapy with minimal damage and establishes a new application scenario of CQDs for precise tumor ablation.

Design of carboxymethylcellulose-conjugated polymeric prodrug micelles for enhanced in vivo performance of docetaxel.

Docetaxel (DTX) has become one of the most important cytotoxic drugs to treat cancer; nevertheless, its poor hydrophilicity and non-specific distribution of DTX lead to detrimental side effects. In this article, we devised carboxymethylcellulose (CMC)-conjugated polymeric prodrug micelles (mPEG-CMC-DTX PMs) for DTX delivery. The ester-bonded polymeric prodrug, mPEG-CMC-DTX, was synthesized and exhibited the capacity for self-assembling into polymeric micelles. The CMC is profusely substituted and acetylated to promote the coupling rate of DTX. Covalent binding of DTX and CMC through an ester bond can be hydrolyzed to dissociate the bond under the action of esterase in the tumor. The mPEG-CMC-DTX PMs displayed promoted drug loading (>50 %, wt), commendable stability, and sustained release behavior in vitro. The gradual release of the prodrug amplified the selectivity of cytotoxicity between normal cells and tumor cells, mitigating the systemic toxicity of mPEG-CMC-DTX PMs and enabling dose intensification. Notably, mPEG-CMC-DTX PMs demonstrated a superior antitumor efficacy and low systemic toxicity due to the elevated tolerance dosage (even at 40 mg/kg DTX). In summation, mPEG-CMC-DTX PMs harmonized the antitumor efficacy and toxicity of DTX. In essence, innovative perspectives for the rational design of CMC-conjugated polymeric prodrug micelles for the delivery of potently toxic drugs were proffered.

Research on the preparation process of the cytarabine/daunorubicin dual-encapsulation liposome and its physicochemical properties and performances in vitro/vivo.

As the only Food and Drug Administration (FDA)-approved dual-encapsulation liposome injection for treating Acute myeloid leukemia (AML), CPX-351 outperforms the standard chemotherapy treatment "DA 7 + 3″ in terms of clinical effectiveness. Although research on dual-loaded liposomes has increased in recent years, little attention has been paid to their preparation, which can affect their quality, efficacy, and safety. This study explored various preparation processes to create the cytarabine/daunorubicin co-loaded liposome (the Cyt/Daun liposome) and eventually settled on two methods: the sequential loading approach, thin film hydration-extrusion-copper ion gradient, and the simultaneous encapsulation technique, copper ion gradient-concentration gradient. Different preparation methods resulted in different particle sizes and encapsulation efficiencies; the two aforementioned preparation processes generated dual-loaded liposomes with comparable physicochemical properties. The sequential encapsulation technique was selected for the subsequent research owing to its higher encapsulation efficiency prior to purification; the prepared Cyt/Daun liposomes had small and uniform particle size (108.6 ± 1.02 nm, Polydispersity index (PDI) 0.139 ± 0.01), negative charge (-(60.2 ± 1.15) mV), high drug encapsulation efficiency (Cyt 88.2 ± 0.24 %, Duan 94.2 ± 0.45 %) and good plasma stability. To improve its storage stability, the Cyt/Daun liposome was lyophilized (-40 °C for 4 h, maintained for 130 min, and dried for 1200 min) using sucrose-raffinose (mass ratio 7:3; glycolipid ratio 4:1, w/w) as a lyoprotectant. The lyophilized liposomes were purple cakes, redissolved rapidly with insignificant alterations in particle size and encapsulation efficiency, and possessed well storage stability. The pharmacokinetic and tissue distribution studies demonstrated that the Cyt/Daun liposome could achieve long circulation and maintain synergic proportions of drugs within 24 h, increasing the accumulation of drugs at tumor sites. Furthermore, the in vitro/in vivo pharmacodynamic studies confirmed its good anti-tumor activity and safety.

Intratumoral injection of norcantharidin liposome emulsion hybrid delivery system amplifies the cancer-fighting effects of oral sorafenib against hepatocellular carcinoma.

Interventional therapies are increasingly used in clinical trials for hepatocellular carcinoma (HCC). Sorafenib is the front-line remedy for HCC, however, chemoresistance occurs immutably and affects the effectiveness of treatment. In a previous study, a norcantharidin liposome emulsion hybrid (NLEH) delivery system for HCC was developed. This study aims to examine the therapeutic effects of the combination of intratumoral injection of NLEH and sorafenib in treating HCC. Sorafenib combined with NLEH activated the apoptosis pathway by synergistically upregulating caspase-9, promoting cytotoxicity, apoptosis (64.57%), and G2/M cell cycle arrest (48.96%). Norcantharidin could alleviate sorafenib resistance by counteracting sorafenib-induced phosphorylation of Akt. Additionally, intratumoral injection of NLEH exhibited a sustained accumulation in the tumor within 24 h and didn't distribute to other major organs. Intratumoral injection of NLEH in combination with oral sorafenib displayed the most potent tumor growth inhibitory effect (77.91%) in vivo. H&E staining results and the indicators of the renal and liver function tests demonstrated the safety of this combination therapy. Overall, these results showed that intratumoral injection of NLEH in combination with oral sorafenib treatment represented a rational potential therapeutic option for HCC.

Bone-homing metastatic breast cancer cells impair osteocytes' mechanoresponse in a 3D loading model.

Breast cancer predominantly metastasizes to the skeleton. Mechanical loading is reliably anabolic in bone, and also inhibits bone metastatic tumor formation and bone loss in vivo. To study the underlying mechanisms, we developed a 3D culture model for osteocytes, the primary bone mechanosensor. We verified that MLO-Y4s responded to perfusion by reducing their rankl and rankl:opg gene expression. We next cultured MLO-Y4s with tumor-conditioned media (TCM) collected from human breast cancer cells (MDA-MB-231s) and a corresponding bone-homing subclone to test the impacts on osteocytes' mechanosensation. We found that TCM from the bone-homing subclone was more detrimental to MLO-Y4 growth and viability, and it abrogated loading-induced changes to rankl:opg. Our studies demonstrate that MLO-Y4s, including their mechanoresponse to perfusion, were more negatively impacted by soluble factors from bone-homing breast cancer cells compared to those from parental cells.

Double-Heterostructured Nickel Phosphate-Phosphides as High-Activity Electrocatalyst for Ammonia Borane Electrooxidation.

The direct electrooxidation reaction of ammonia borane (ABOR) as the anodic reaction of direct ammonia borane fuel cells (DABFCs) is greatly dependent on the properties of electrocatalysts. Both the active sites and charge/mass transfer characteristics are the key to promoting the processes of kinetics and thermodynamics, which can further improve the electrocatalytic activity. Hence, the catalyst double-heterostructured Ni2 P/Ni2 P2 O7 /Ni12 P5 (d-NPO/NP) with the optimistic redistribution of electrons and active sites is prepared for the first time. The d-NPO/NP-750 catalyst obtained after pyrolysis at 750 °C shows the outstanding electrocatalytic activity toward ABOR with an onset potential of -0.329 V vs RHE which is better than all the published catalysts. The density functional theory (DFT) computations illustrate that the Ni2 P2 O7 /Ni2 P acts as the activity enhancement heterostructure with a high d-band center (-1.60 eV) and the low activation energy barrier, while the Ni2 P2 O7 /Ni12 P5 acts as the conductivity enhancement heterostructure with the highest density of valence electrons.

Graves Disease and Inflammatory Bowel Disease: A Bidirectional Mendelian Randomization.

CONTEXT: Both Graves disease (GD) and inflammatory bowel disease (IBD) are common autoimmune diseases that severely damage a patient's quality of life. Previous epidemiological studies have suggested associations between GD and IBD. However, whether a causal relationship exists between these 2 diseases remains unknown. OBJECTIVE: To infer a causal relationship between GD and IBD using bidirectional 2-sample Mendelian randomization (MR). METHODS: We performed bidirectional 2-sample MR to infer a causal relationship between GD and IBD using genome-wide association study summary data obtained from Biobank Japan and the International Inflammatory Bowel Disease Genetic Consortium. Several methods (random-effect inverse variance weighted, weighted median, MR-Egger regression, and MR-PRESSO) were used to ensure the robustness of the causal effect. Heterogeneity was measured based on Cochran's Q value. Horizontal pleiotropy was evaluated by MR-Egger regression and leave-one-out analysis. RESULTS: Genetically predicted IBD may increase the risk of GD by 24% (odds ratio [OR] 1.24, 95% CI 1.01-1.52, P = .041). Crohn disease (CD) may increase the risk of GD, whereas ulcerative colitis (UC) may prevent patients from developing GD. Conversely, genetically predicted GD may slightly increase the risk of CD, although evidence indicating that the presence of GD increased the risk of UC or IBD was lacking. Outlier-corrected results were consistent with raw causal estimates. CONCLUSION: Our study revealed a potentially higher comorbidity rate for GD and CD. However, UC might represent a protective factor for GD. The underlying mechanism and potential common pathways await discovery.

PD-L1 blockade liberates intrinsic antitumourigenic properties of glycolytic macrophages in hepatocellular carcinoma.

OBJECTIVE: Patients with increased PD-L1+ host cells in tumours are more potent to benefit from antiprogrammed death-1/programmed death ligand-1 (PD-L1) treatment, but the underlying mechanism is still unclear. We aim to elucidate the nature, regulation and functional relevance of PD-L1+ host cells in hepatocellular carcinoma (HCC). DESIGN: A total of untreated 184 HCC patients was enrolled randomly. C57BL/6 mice are given injection of Hepa1-6 cells to form autologous hepatoma. ELISpot, flow cytometry and real-time PCR are applied to analyse the phenotypic characteristics of PD-L1+ cells isolated directly from HCC specimens paired with blood samples or generated from ex vivo and in vitro culture systems. Immunofluorescence and immunohistochemistry are performed to detect the presence of immune cells on paraffin-embedded and formalin-fixed samples. The underlying regulatory mechanisms of metabolic switching are assessed by both in vitro and in vivo studies. RESULTS: We demonstrate that PD-L1+ host macrophages, which constructively represent the major cellular source of PD-L1 in HCC tumours, display an HLA-DRhighCD86high glycolytic phenotype, significantly produce antitumourigenic IL-12p70 and are polarised by intrinsic glycolytic metabolism. Mechanistically, a key glycolytic enzyme PKM2 triggered by hepatoma cell derived fibronectin 1, via a HIF-1α-dependent manner, concurrently controls the antitumourigenic properties and inflammation-mediated PD-L1 expression in glycolytic macrophages. Importantly, although increased PKM2+ glycolytic macrophages predict poor prognosis of patients, blocking PD-L1 on these cells eliminates PD-L1-dominant immunosuppression and liberates intrinsic antitumourigenic properties. CONCLUSIONS: Selectively modulating the 'context' of glycolytic macrophages in HCC tumours might restore their antitumourigenic properties and provide a precise strategy for anticancer therapy.

Hepatic stellate cell activation and senescence induced by intrahepatic microbiota disturbances drive progression of liver cirrhosis toward hepatocellular carcinoma.

BACKGROUND: The significance of the relationship between the microbiota and diseases is increasingly being recognized. However, the characterization of tumor microbiome and their precise molecular mechanisms through which microbiota promotes hepatocellular carcinoma (HCC) development are still unclear. METHODS: The intrahepatic microbiota was investigated from tumor, normal adjacent tissues in 46 patients with HCC and normal hepatic tissues in 33 patients with hemangioma by 16S rRNA gene sequencing. Taxonomic composition differences in patients were evaluated using Linear discriminant analysis Effect Size (LefSe) and Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) to predict microbial functional pathways. Associations between the most relevant taxa and clinical characteristics of HCC patients were analyzed by Spearman rank correlations. The effects of microbe on hepatic stellate cells (HSCs) activation and HCC progression were examined. RESULTS: We observed intrahepatic microbiota disturbances by reduced microbial diversity in HCC. The tumor microbiota of the HCC patients with cirrhosis showed higher abundance of Stenotrophomonas maltophilia (S. maltophilia). S. maltophilia provoked senescence-associated secretory phenotype (SASP) in HSCs by activating TLR-4-mediated NF-κB signaling pathway, which in turn induced NLRP3 inflammasome complex formation and secreted various inflammatory factors in the liver, thus facilitating HCC progression in mice. Moreover, signs of SASP were also observed in the HSCs in the area of HCC with higher S. maltophilia enrichment arising in patients with cirrhosis. CONCLUSIONS: Our analysis of the hepatic microbiota revealed for the first time that patients with HCC exhibited a dysbiotic microbial community with higher S. maltophilia abundance, which induced the expression SASP factors of HSCs and cirrhosis in the liver, concurring in the process of hepatocarcinogenesis.

Influence of gut and intratumoral microbiota on the immune microenvironment and anti-cancer therapy.

Microbiota has been implicated in the regulation of tumor progression and therapeutic efficacy. However, the effect of microbiota on disease progression is context dependent, differing according to tumor types, therapeutic regimens, and composition of the microbiota, calling for a deeper understanding of host-microbiome interactions. Previous studies have demonstrated that gut microbiota influences disease progression by regulating local and systemic immunity. Notably, with the advent of next-generation sequencing technology, intratumoral microbiota has also been found and constitutes an important component of the tumor microenvironment. In this review, we summarize recent knowledge about the identification of intra-tumor microbiota and discuss the role of gut and intratumoral microbiota in solid tumors in the angle of immune microenvironment interaction. Furthermore, we discuss how these findings may benefit current anti-cancer approaches. Key problems to be solved in ongoing and future research are highlighted.

Flow inside a bone scaffold: Visualization using 3D phase contrast MRI and comparison with numerical simulations.

We report on results of experimental flow measurements inside a bone scaffold model, subjected to a uniform incoming flow (applied perfusion). Understanding the flow behavior inside a tissue engineered scaffold is essential for mechanistic studies of mechanobiology, particularly flow-sensitive bone cells. Nearly all existing studies that quantify interstitial flow inside engineered bone scaffolds have been based on numerical results, in part due to the difficulties associated with quantitative measurements and visualization of flow inside large, opaque bone or bone mimics. Thus, an experimental platform to complement and validate in silico studies is needed. Therefore, we developed a flow visualization method using Phase-Contrast Magnetic Resonance Imaging (PC-MRI) to measure flow velocities within a 3D-printed microCT-based rendering of a bone scaffold. We designed and built a non-magnetic recirculating water tunnel to apply uniform perfusion to the 3D-printed model and we measured flow distribution within the scaffold and compared these experimental results with CFD results. Both magnitude and distribution of flow velocities observed at different slices of the scaffold were in quantitative agreement numerically and experimentally. This experimental approach can be used to both validate numerical studies and provide insight into the flow behavior inside tissue-engineered scaffolds for a range of applications, including fundamental mechanobiology of healthy cells, and in the context of diseases, such as cancer.

Multiphysics simulation of a compression-perfusion combined bioreactor to predict the mechanical microenvironment during bone metastatic breast cancer loading experiments.

Incurable breast cancer bone metastasis causes widespread bone loss, resulting in fragility, pain, increased fracture risk, and ultimately increased patient mortality. Increased mechanical signals in the skeleton are anabolic and protect against bone loss, and they may also do so during osteolytic bone metastasis. Skeletal mechanical signals include interdependent tissue deformations and interstitial fluid flow, but how metastatic tumor cells respond to each of these individual signals remains underinvestigated, a barrier to translation to the clinic. To delineate their respective roles, we report computed estimates of the internal mechanical field of a bone mimetic scaffold undergoing combinations of high and low compression and perfusion using multiphysics simulations. Simulations were conducted in advance of multimodal loading bioreactor experiments with bone metastatic breast cancer cells to ensure that mechanical stimuli occurring internally were physiological and anabolic. Our results show that mechanical stimuli throughout the scaffold were within the anabolic range of bone cells in all loading configurations, were homogenously distributed throughout, and that combined high magnitude compression and perfusion synergized to produce the largest wall shear stresses within the scaffold. These simulations, when combined with experiments, will shed light on how increased mechanical loading in the skeleton may confer anti-tumorigenic effects during metastasis.

Perfusion applied to a 3D model of bone metastasis results in uniformly dispersed mechanical stimuli.

Breast cancer most frequently metastasizes to the skeleton. Bone metastatic cancer is incurable and induces wide-spread bone osteolysis, resulting in significant patient morbidity and mortality. Mechanical cues in the skeleton are an important microenvironmental parameter that modulate tumor formation, osteolysis, and tumor cell-bone cell signaling, but which mechanical signals are the most beneficial and the corresponding molecular mechanisms are unknown. We focused on interstitial fluid flow based on its well-known role in bone remodeling and in primary breast cancer. We created a full-scale, microCT-based computational model of a 3D model of bone metastasis undergoing applied perfusion to predict the internal mechanical environment during in vitro experimentation. Applied perfusion resulted in uniformly dispersed, heterogeneous fluid velocities, and wall shear stresses throughout the scaffold's interior. The distributions of fluid velocity and wall shear stress did not change within model sub-domains of varying diameter and location. Additionally, the magnitude of these stimuli is within the range of anabolic mechanical signals in the skeleton, verifying that our 3D model reflects previous in vivo studies using anabolic mechanical loading in the context of bone metastasis. Our results indicate that local populations of cells throughout the scaffold would experience similar mechanical microenvironments.