•Novel hemoglobin loaded nanoliposome (HLL) with high stability was presented here.•The HLL could efficiently accumulate into the ectopic and orthotopic tumor.•The HLL could efficiently alleviate of tumor hypoxia.•HIF-1α and VEGF in tumor was obviously down-regulated after the application of HLL.Hemoglobin-based oxygen carriers were developed as an alternative for blood transfusion. However, the research progress for their further clinic applications was slow in recent several years. Hypoxia is found in most solid tumors, which is responsible for the tumor formation, increased metastasis, drug resistance during therapeutic process as well as poor patient survival. In this work, novel hemoglobin (Hb) loaded nanoliposomes, as artificial red cells for oxygen delivery, were optimized by screening various types of phospholipids and analyzing different mole ratio of phospholipid to cholesterol. The nanoliposomes presented a high encapsulating efficiency to hemoglobin and also significantly enhanced its stability. The obtained hemoglobin loaded nanoliposome (HLL) could be lyophilized for long term storage. HLL did not cause significant cell death in the concentration range of 0–100 μg equivalent Hb/mL under normoxia and hypoxia incubation conditions, suggesting the low cytotoxicity and high biocompatibility of HLL. Importantly, HLL could efficiently accumulate into subcutaneous and deep orthotopic tumors, inducing a significant decrease of hypoxia-inducible factors 1α subunits (HIF-1α) in the tumors and remarkably reduced expression of vascular endothelial growth factor (VEGF). The study of acute and chronic toxicity indicated that HLL did not induce obvious damage to main organs of mice after intravenous injections with total Hb dose of 120 mg/kg. We presented a promising method for relieving the hypoxia degree in solid tumors and down-regulating HIF-1α protein by directly delivering oxygen into tumors, which will be very helpful for subsequent cancer therapy.Download high-res image (183KB)Download full-size image

In the present report, a degradable gene delivery system (PAMS/DNA/10NLS) containing nucleus location signal peptide (NLS) was prepared. The agarose gel electrophoresis, particle size and zeta potential of PAMS/DNA/10NLS were similar to those of PAMS/DNA, which proved that NLS did not affect the interaction between PAMS and DNA. PAMS/DNA/10NLS exhibited marked extracellular and intracellular degradation under acidic conditions. The degradation was believed to allow NLS to come into contact with importins easily, which was able to mediate the nucleus import. With the help of NLS, PAMS/DNA/10NLS exhibited a higher transfection capability than PAMS/DNA. Moreover, the transfection of PAMS/DNA/10NLS was less dependent on the breakdown of the nucleus envelope than PAMS/DNA. Considering that GTPase-activating protein 1 (RanGAP1) was able to activate the endogenous GTPase, which was necessary for NLS-mediated nucleus import, RanGAP1 overexpressed cells (RanGAP1 cells) were produced. This result showed that RanGAP1 cells had higher GTPase activities than normal cells. Both the nucleus import and transfection efficiency of PAMS/DNA/10NLS were markedly higher in RanGAP1 cells than that in normal cells. The in vivo transfection results also showed that the transfection efficiency of PAMS/DNA/10NLS was higher in RanGAP1 pre-treated mice than that in normal mice. These findings showed that PAMS/DNA/10NLS is a promising gene delivery system with the assistance of RanGAP1.Statement of SignificanceThe present report describes the increased transfection efficiency of a degradable gene delivery system (PAMS/DNA/10NLS) containing nuclear location signal (NLS) with the assistance of GTPase-activating protein 1 (RanGAP1). The physicochemical properties of PAMS/DNA/10NLS were similar to those of PAMS/DNA. PAMS/DNA/10NLS exhibited great extracellular and intracellular degradations, which might allow NLS to contact with importins easily. With the help of NLS, PAMS/DNA/10NLS exhibited a higher transfection capability than PAMS/DNA. The transfection of PAMS/DNA/10NLS had less dependence on the breakdown of nuclear envelope. Both the nuclear import and transfection efficiency of PAMS/DNA/10NLS were higher in RanGAP1 overexpressed cells than that in normal cells. Moreover, the transfection efficiency of PAMS/DNA/10NLS was higher in RanGAP1 pre-treated mice than that in normal mice.Download high-res image (131KB)Download full-size image

Multidrug resistance (MDR) has been a major obstacle to tumor chemotherapy. Pluronic unimers have been reported to be promising copolymers to reverse MDR, and the intracellular delivery of Pluronic unimers is a problem worth thinking. To exert the excellent reversal effect of Pluronic unimers, DOX-loaded G4.0 PAMAM was modified with PluronicL64 via cis-aconitic acid as a pH-sensitive linkage (PCPAMAM/DOX), which could release DOX and Pluronic unimers into cytoplasm. The Pluronic-modified PAMAM (PCPAMAM) exhibited favorable biocompatibility and pH-sensitivity. PCPAMAM/DOX showed a nano-scale size and a sustained in vitro release profile. Compared with a control formulation, PCPAMAM/DOX showed a higher reversal effect on MCF-7/ADR cells and enhanced intracellular drug accumulation. The results of P-gp activity, subcellular distribution of PluronicL64, the ATP level and mitochondrial transmembrane potential all illustrated that free Pluronic unimers could be released by PCPAMAM functioning as reversal agents. In conclusion, PCPAMAM could be a promising vehicle to enhance DOX accumulation by overcoming MDR in MCF-7/ADR cells. This work also provided an effective method to deliver Pluronic unimers into MDR cells.

To explain thermal stability enhancement of an organic compound, sucralose, with cyclodextrin based metal organic frameworks.Micron and nanometer sized basic CD-MOFs were successfully synthesized by a modified vapor diffusion method and further neutralized with glacial acetic acid. Sucralose was loaded into CD-MOFs by incubating CD-MOFs with sucralose ethanol solutions. Thermal stabilities of sucralose-loaded basic CD-MOFs and neutralized CD-MOFs were investigated using thermogravimetric analysis (TGA), differential scanning calorimetry (DSC) and high performance liquid chromatography with evaporative light-scattering detection (HPLC-ELSD).Scanning electron microscopy (SEM) and powder X-ray diffraction (PXRD) results showed that basic CD-MOFs were cubic crystals with smooth surface and uniform sizes. The basic CD-MOFs maintained their crystalline structure after neutralization. HPLC-ELSD analysis indicated that the CD-MOF crystal size had significant influence on sucralose loading (SL). The maximal SL of micron CD-MOFs (CD-MOF-Micro) was 17.5 ± 0.9% (w/w). In contrast, 27.9 ± 1.4% of sucralose could be loaded in nanometer-sized basic CD-MOFs (CD-MOF-Nano). Molecular docking modeling showed that sucralose molecules preferentially located inside the cavities of γ-CDs pairs in CD-MOFs. Raw sucralose decomposed fast at 90°C, with 86.2 ± 0.2% of the compound degraded within only 1 h. Remarkably, sucralose stability was dramatically improved after loading in neutralized CD-MOFs, with only 13.7 ± 0.7% degradation at 90°C within 24 h.CD-MOFs efficiently incorporated sucralose and maintained its integrity upon heating at elevated temperatures.

•Redox and pH sensitive PAMAM-SS-PEG conjugates were synthesized by a simple process.•Conjugates exhibited excellent stability and effectively diminished uptake by RES.•Drug release from the conjugates was followed a reduction and pH-triggered pattern.•DOX-loaded conjugates showed significant cytotoxicity against B16 and A549 cells.Nanocarriers with low toxicity, high stability, long circulation in blood and triggered drug release at target sites are extremely needed for cancer therapy. In this study, different molar ratios of poly (ethylene glycol) (PEG) were conjugated to poly (amidoamine) dendrimer (PAMAM) by cleavable disulfide bonds, yielding redox and pH dual-responsive nanocarriers (PSSP). The redox sensitivity of the PSSP conjugates was confirmed by measuring the change in particle size and zeta potential under reductive environment by dynamic light scattering (DLS). Meanwhile, all conjugates showed slight hemolytic ratio and high stability in 10% FBS for 24 h. Phagocytosis experiments indicated that the PSSP conjugates could effectively escape the engulfment of macrophage. Doxorubicin (DOX) was loaded into the hydrophobic core of the conjugates with loading content of 10%. In vitro release studies suggested that release rate as well as cumulative release amount of DOX from the PSSP conjugates were pronouncedly enhanced in reductive environment as compared with the non-cleavable counterparts. Besides, all conjugates exhibited excellent acid-triggered release performance. Significantly, PEGylation degree of the conjugates had critical impact on drug release. Cytotoxicity tests displayed that DOX-loaded PSSP conjugates were more potent in killing B16 and A549 tumor cells than the non-cleavable counterparts. Curiously, PSSP conjugate with the highest PEGylation degree showed the best antitumor effect despite the somewhat less cell uptake. Therefore, the dual-responsive PSSP conjugates seem promising candidates for efficient intracellular release of antitumor drugs.

Using different chain lengths of PEG as linkers to develop a novel folate (FA) and TAT peptide co-modified doxorubicin (DOX)-loaded liposome (FA/TAT-LP-DOX) and evaluate its potential for tumor targeted intracellular drug delivery.FA/TAT-LP-DOX was prepared by pH gradient method and post-insertion method and the optimal ligand density was screened by MTT assay. In vitro evaluation was systematically performed through cytotoxicity assay, cellular uptake studies, subcellular localization and cellular uptake mechanism in folate receptor (FR) over-expressing KB tumor cells. In vivo tumor targeted delivery of FA/TAT-LP-DOX was also studied by in vivo fluorescence imaging in a murine KB xenograft model.The particle size and zeta potential determination indicated that FA and TAT were successfully inserted into the liposome and cationic TAT peptide was completely shielded. With the optimal ligand density (5% of FA and 2.5% TAT), the FA/TAT-LP-DOX exhibited improved cytotoxity and cellular uptake efficiency compared with its single-ligand counterparts (FA-LP-DOX and PEG/TAT-LP-DOX). Competitive inhibition and uptake mechanism experiments revealed that FA and TAT peptide played a synergistic effect in facilitating intracellular transport of the liposome, and association between FA and FA receptors activated this transport process. In vivo imaging further demonstrated the superiority of FA/TAT-LP in tumor targeting and accumulation.Folate and TAT peptide co-modified liposome using different chain lengths of PEG as linkers may provide a useful strategy for specific and efficient intracellular drug delivery.

pH-sensitive self-aggregated nanoparticles (SNPs), based on amphiphilic deoxycholic acid (DOCA) modified carboxymethyl chitosan (DCMC), were prepared for delivery of the anticancer drug doxorubicin (DOX). DCMCs with different degrees of substitution (DS) of DOCA were initially synthesized and characterized. Based on self-aggregation, DCMC formed nanoparticles with size ranging from 87 to 174 nm. The critical aggregation concentration (CAC) decreased on increasing the DS of DOCA. Moreover, the DCMC SNPs showed an acidic pH-induced aggregation and deformation behavior. The DOX-loaded SNPs ([D]NP) exhibited a sustained drug release manner, which could be accelerated by an acidic pH, but delayed by a higher DS of DOCA. Antitumor efficacy results showed that [D]NP could suppress both sensitive and resistant MCF-7 cells effectively in a dose- and time-dependent manner. The enhanced cellular uptake and greater retention of [D]NP in drug-resistant cells, as evidenced by confocal microscopy and flow cytometry, contributed to a superior efficacy of [D]NP over free DOX. These results suggest the potential of DCMC SNPs as carriers for the hydrophobic drug DOX for effective cancer therapy against drug-resistant tumors.Graphical abstractHighlights► We have developed a chitosan-derived nanocarrier system for hydrophobic anticancer drugs. ► This system exhibited an acidic pH-induced aggregation, deformation and accelerated drug release profile. ► This system was also much more effective than the free drug against drug-resistant cancer cells.

Although most researches and therapies have been focused on the tumor itself, it is becoming clear that immune cells can not only suppress tumor development but support and maintain their malignant type. Promising recent developments in immunology will provide opportunities for tumor-specific immunotherapy, which can orchestrate the patients immune system to target, fight and eradicate cancer cells without destroying healthy cells. However, antitumor immunity driven by self-immune system alone may be therapeutically insufficient. Developments in nanoparticle based drug delivery system can promote immunotherapy and re-educate immunosuppressive tumor microenvironment (TME), which provide promising strategies for cancer therapy. In this review, we will focus on nanoparticle-based immunotherapeutic approaches against cancer, ranging from nanovaccines, artificial antigen presenting cells (aAPCs) to nanoparticles reversing tumor immunosuppressive microenvironment.Download high-res image (152KB)Download full-size image

The present report describes the synthesis of a hydroxyl terminal PAMAM dendrimer (PAMAM-OH) derivative (PAMSPF). The hydroxyls of PAMAM-OH were attached to S-Methyl-l-cysteine (SMLC) via an acid-labile ester bond, named as β-thiopropionate bond, followed by modification with folic acid (FA) through a polyethylene glycol (PEG) linker. The degrees of attachment of SMLC and FA to the PAMAM-OH backbone were 83.9% and 12.8%, respectively. PAMSPF could condense DNA to form spherical nanoparticles with particle sizes of ∼200 nm and remain stable in the presence of heparin and nuclease. The β-thiopropionate bond in PAMSPF was hydrolyzed completely and the DNA release rate was 95.8 ± 3.3% after incubation under mildly acidic conditions at 37 °C for 3 h. PAMSPF/DNA was less cytotoxic to KB and HepG2 cells and exhibited a higher gene transfection efficiency than native PAMAM/DNA. The uptake assays showed that PAMSPF/DNA entered KB cells within 0.5 h through folate receptor-mediated endocytosis and escaped from endosomes within 2 h. In addition, PAMSPF/DNA displayed long circulation time along with excellent targeting of tumor sites in vivo. These findings demonstrate that PAMSPF is an excellent carrier for safe and effective gene delivery.Download high-res image (185KB)Download full-size image

Treatment of brain tumor remains a great challenge worldwide. Development of a stable, safe, and effective siRNA delivery system which is able to cross the impermeable blood-brain barrier (BBB) and target glioma cells is necessary. This study aims to investigate the therapeutic effects of intravenous administration of T7 peptide modified core-shell nanoparticles (named T7-LPC/siRNA NPs) on brain tumors. Layer-by-layer assembling of protamine/chondroitin sulfate/siRNA/cationic liposomes followed by T7 peptide modification has been carried out in order to obtain a targeted siRNA delivery system. In vitro cellular uptake experiments demonstrated a higher intracellular fluorescence intensity of siRNA in brain microvascular endothelial cells (BMVECs) and U87 glioma cells when treated with T7-LPC/siRNA NPs compared with PEG-LPC/siRNA NPs. In the co-culture model of BMVECs and U87 cells, a significant down-regulation of EGFR protein expression occurred in the U87 glioma cells after treatment with the T7-LPC/siEGFR NPs. Moreover, the T7-LPC/siRNA NPs had an advantage in penetrating into a deep region of the tumor spheroid compared with PEG-LPC/siRNA NPs. In vivo imaging revealed that T7-LPC/siRNA NPs accumulated more specifically in brain tumor tissues than the non-targeted NPs. Also, in vivo tumor therapy experiments demonstrated that the longest survival period along with the greatest downregulation of EGFR expression in tumor tissues was observed in mice with an intracranial U87 glioma treated with T7-LPC/siEGFR NPs compared with mice receiving other formulations. Therefore, we believe that these transferrin receptor-mediated core-shell nanoparticles are an important potential siRNA delivery system for brain tumor-targeted therapy.To develop a systemic stable, safe, and effective siRNA delivery system that is able to cross the impermeable blood-brain barrier (BBB) and target glioma cells is necessary. The targeted siRNA delivery system (named as T7-LPC/siRNA NPs) is obtained from the layer-by-layer assembling of protamine/chondroitin sulfate/siRNA/cationic liposomes followed by T7 peptide modification. The longest survival period along with the strongest downregulation of EGFR expression in tumor tissues were observed from the mice bearing intracranial U87 glioma after intravenous administration of T7-LPC/siEGFR NPs.Download high-res image (171KB)Download full-size image

Multidrug resistance (MDR) has been a major obstacle to tumor chemotherapy. Pluronic unimers have been reported to be promising copolymers to reverse MDR, and the intracellular delivery of Pluronic unimers is a problem worth thinking. To exert the excellent reversal effect of Pluronic unimers, DOX-loaded G4.0 PAMAM was modified with PluronicL64 via cis-aconitic acid as a pH-sensitive linkage (PCPAMAM/DOX), which could release DOX and Pluronic unimers into cytoplasm. The Pluronic-modified PAMAM (PCPAMAM) exhibited favorable biocompatibility and pH-sensitivity. PCPAMAM/DOX showed a nano-scale size and a sustained in vitro release profile. Compared with a control formulation, PCPAMAM/DOX showed a higher reversal effect on MCF-7/ADR cells and enhanced intracellular drug accumulation. The results of P-gp activity, subcellular distribution of PluronicL64, the ATP level and mitochondrial transmembrane potential all illustrated that free Pluronic unimers could be released by PCPAMAM functioning as reversal agents. In conclusion, PCPAMAM could be a promising vehicle to enhance DOX accumulation by overcoming MDR in MCF-7/ADR cells. This work also provided an effective method to deliver Pluronic unimers into MDR cells.