Are third-generation active-targeting nanoformulations definitely the best? In vitro and in vivo comparisons of pixantrone-loaded liposomes modified with different sialic acid derivatives.

Treatment with sialic acid-octadecylamine (SA-ODA)-modified pixantrone (Pix) liposomes results in favorable antitumor effects by targeting tumor-associated macrophages (TAMs). To explore the influence of different types of SA decorations on antitumor efficiency, we synthesized a PEGylated SA derivative, SA-PEG2000-DSPE, and combined it with SA-ODA to construct three representative types of SA-modified liposomes (SA-ODA-modified Pix liposomes, SA-ODA-modified Pix liposomes with different PEG densities, and SA-PEG2000-DSPE-modified Pix liposomes, named Pix-SACL, Pix-SPL-0.2/0.5/2.0/5.0, and Pix-SAPL, respectively). All the Pix liposomes were nanoscale formulations, having diameters between 100 and 150 nm, high encapsulation efficiencies (> 90%), and slow drug release properties. The in vivo blood circulation time of the PEGylated formulations (Pix-SPL-0.2/0.5/2.0/5.0 and Pix-SAPL) showed an upward trend with increasing PEG density, but there was no significant difference between adjacent groups. All PEGylated formulations displayed increased tumor accumulation when compared with Pix-SACL, but there was no significant difference among them. However, the antitumor activity of SA-modified liposomes was not positively correlated with circulation time or tumor accumulation in S180-bearing mice. Pix-SPL-0.2 displayed the strongest antitumor effect and lowest toxicity among the formulations tested in this study. With Pix-SPL-0.2 treatment, 66.7% of the mice demonstrated tumor shedding and wound healing.

Innovative development path of ethnomedicines: the interpretation of the path.

One of the primary purposes of the innovative development of ethnomedicines is to use their excellent safety and significant efficacy to serve a broader population. To achieve this purpose, modern scientific and technological means should be referenced, and relevant national laws and regulations as well as technical guides should be strictly followed to develop standards and to perform systemic research in producing ethnomedicines. Finally, ethnomedicines, which are applied to a limited extent in ethnic areas, can be transformed into safe, effective, and quality-controllable medical products to relieve the pain of more patients. The innovative development path of ethnomedicines includes the following three primary stages: resource study, standardized development research, and industrialization of the achievements and efforts for internationalization. The implementation of this path is always guaranteed by the research and development platform and the talent team. This article is based on the accumulation of long-term practice and is combined with the relevant disciplines, laws and regulations, and technical guidance from the research and development of ethnomedicines. The intention is to perform an in-depth analysis and explanation of the major research thinking, methods, contents, and technical paths involved in all stages of the innovative development path of ethnomedicines to provide useful references for the development of proper ethnomedicine use.

The application of EDTA in drug delivery systems: doxorubicin liposomes loaded via NH4EDTA gradient.

The applications of ethylenediaminetetraacetic acid (EDTA) have been expanded from the treatment of heavy metal poisoning to chelation therapies for atherosclerosis, heart disease, and cancers, in which EDTA reduces morbidity and mortality by chelating toxic metal ions. In this study, EDTA was used in a drug delivery system by adopting an NH4EDTA gradient method to load doxorubicin into liposomes with the goal of increasing therapeutic effects and decreasing drug-related cytotoxicity. The particle size of the optimum NH4EDTA gradient liposomes was 79.4±1.87 nm, and the entrapment efficiency was 95.54%±0.59%. In vitro studies revealed that liposomes prepared using an NH4EDTA gradient possessed long-term stability and delayed drug release. The in vivo studies also showed the superiority of the new doxorubicin formulation. Compared with an equivalent drug dose (5 mg/kg) prepared by (NH4)2SO4 gradient, NH4EDTA gradient liposomes showed no significant differences in tumor inhibition ratio, but cardiotoxicity and liposome-related immune organ damage were lower, and no drug-related deaths were observed. These results show that use of the NH4EDTA gradient method to load doxorubicin into liposomes could significantly reduce drug toxicity without influencing antitumor activity.

Self-assembled micelles of novel amphiphilic copolymer cholesterol-coupled F68 containing cabazitaxel as a drug delivery system.

Despite being one of the most promising amphiphilic block copolymers, use of Pluronic F68 in drug delivery is limited due to its high critical micelle concentration (CMC). In this study, we developed a novel F68 derivative, cholesterol-coupled F68 (F68-CHMC). This new derivative has a CMC of 10 µg/mL, which is 400-fold lower than that of F68. The drug-loading capacity of F68-CHMC was investigated by encapsulating cabazitaxel, a novel antitumor drug. Drug-loaded micelles were fabricated by a self-assembly method with simple dilution. The optimum particle size of the micelles was 17.5±2.1 nm, with an entrapment efficiency of 98.1% and a drug loading efficiency of 3.16%. In vitro release studies demonstrated that cabazitaxel-loaded F68-CHMC micelles had delayed and sustained-release properties. A cytotoxicity assay of S180 cells showed that blank F68-CHMC was noncytotoxic with a cell viability of nearly 100%, even at a concentration of 1,000 µg/mL. The IC50 revealed that cabazitaxel-loaded F68-CHMC micelles were more cytotoxic than Tween 80-based cabazitaxel solution and free cabazitaxel. In vivo antitumor activity against S180 cells also indicated better tumor inhibition by the micelles (79.2%) than by Tween 80 solution (56.2%, P<0.05). Based on these results, we conclude that the F68-CHMC copolymer may be a potential nanocarrier to improve the solubility and biological activity of cabazitaxel and other hydrophobic drugs.

The anticancer efficacy of pixantrone-loaded liposomes decorated with sialic acid-octadecylamine conjugate.

Based on the knowledge that sialic acid is a critical element for tumor development and its receptors are highly expressed on the tumor-associated macrophages (TAMs) which play important roles in the growth and metastasis of tumors, we synthesized a sialic acid-octadecylamine conjugate (SA-ODA) and anchored it on the surface of pixantrone (Pix)-loaded liposomes, to achieve an improved anticancer effect. Four Pix formulations (Pix-S, Pix-CL, Pix-PL and Pix-SAL represent solution, conventional liposome, stealth liposome, and SA-ODA modified liposome, respectively) were developed, and various parameters, including drug loading, stability, in vitro release, cytotoxicity and pharmacokinetics, were evaluated. The tumor growth inhibition and toxicity studies were performed in S180-bearing Kunming mice. Pix-S exhibited a strong toxicity to the immune system, accelerated the growth of tumors and reduced the lifespan of mice. In contrast, Pix-SAL displayed the strongest anticancer and life-prolonging effects among all of the formulations in this study. More importantly, injection of Pix-SAL induced a phenomenon whereby the cancerous tissues were "shed" from mice, after which the wound healed. We speculate that this special efficacy may be partly due to the killing of TAMs by Pix-SAL. This study suggests that SA-ODA modified liposomes may serve as an effective intravenous delivery vehicle for Pix.

A noticeable phenomenon: thiol terminal PEG enhances the immunogenicity of PEGylated emulsions injected intravenously or subcutaneously into rats.

Repeated intravenous injection of long-circulating methoxy-polyethylene glycol (PEG)-liposomes alters the pharmacokinetics and biodistribution of the second administration, regarded as the "accelerated blood clearance (ABC) phenomenon." Nevertheless, the effect of terminal groups of distearoylphosphatidylethanolamine-polyethylene glycol (DSPE-PEG) on the induction of the ABC phenomenon had not been reported previously. In this study, rats were injected intravenously or subcutaneously with PEG coated emulsions (DE) which were prepared using PEG terminated with either the methoxyl (OCH3), hydroxyl (OH), amino (NH2), carboxyl (COOH), or thiol (SH) group. DE-OCH3 demonstrated the longest prolonged half-life in vivo after a single intravenous injection, followed by DE-SH and DE-COOH. In contrast, DE-OH was rapidly removed from the blood circulation, as was DE-NH2. Moreover, we observed a strong positive relationship between the circulation time of initially injected PEGylated emulsions and the extent to which the ABC phenomenon was induced, but a exception of DE-SH increasing the ABC effect. Furthermore, the present study suggested that thiols might stimulate the proliferation and differentiation of B cells to induce the fastest clearance of the second intravenous administration by inducing the synthesis of the cell membrane and cytosolic proteins or reacting with follicular dendritic cells. The results strongly suggested that thiol groups played a stimulatory role in the immune response and provided a considerable implication for multiple drug therapy of thiol groups.

Accelerated drug release and clearance of PEGylated epirubicin liposomes following repeated injections: a new challenge for sequential low-dose chemotherapy.

BACKGROUND: Sequential low-dose chemotherapy has received great attention for its unique advantages in attenuating multidrug resistance of tumor cells. Nevertheless, it runs the risk of producing new problems associated with the accelerated blood clearance phenomenon, especially with multiple injections of PEGylated liposomes. METHODS: Liposomes were labeled with fluorescent phospholipids of 1,2-dipalmitoyl-snglycero-3-phosphoethanolamine-N-(7-nitro-2-1,3-benzoxadiazol-4-yl) and epirubicin (EPI). The pharmacokinetics profile and biodistribution of the drug and liposome carrier following multiple injections were determined. Meanwhile, the antitumor effect of sequential low-dose chemotherapy was tested. To clarify this unexpected phenomenon, the production of polyethylene glycol (PEG)-specific immunoglobulin M (IgM), drug release, and residual complement activity experiments were conducted in serum. RESULTS: The first or sequential injections of PEGylated liposomes within a certain dose range induced the rapid clearance of subsequently injected PEGylated liposomal EPI. Of note, the clearance of EPI was two- to three-fold faster than the liposome itself, and a large amount of EPI was released from liposomes in the first 30 minutes in a complement-activation, direct-dependent manner. The therapeutic efficacy of liposomal EPI following 10 days of sequential injections in S180 tumor-bearing mice of 0.75 mg EPI/kg body weight was almost completely abolished between the sixth and tenth day of the sequential injections, even although the subsequently injected doses were doubled. The level of PEG-specific IgM in the blood increased rapidly, with a larger amount of complement being activated while the concentration of EPI in blood and tumor tissue was significantly reduced. CONCLUSION: Our investigation implied that the accelerated blood clearance phenomenon and its accompanying rapid leakage and clearance of drug following sequential low-dose injections may reverse the unique pharmacokinetic-toxicity profile of liposomes which deserved our attention. Therefore, a more reasonable treatment regime should be selected to lessen or even eliminate this phenomenon.

Effect of particle size on solubility, dissolution rate, and oral bioavailability: evaluation using coenzyme Q10 as naked nanocrystals.

In this paper work, four naked nanocrystals (size range 80-700 nm) were prepared without any surfactant or polymer using the solvent/nonsolvent method. The effects of particle size on their solubility, dissolution, and oral bioavailability were investigated. Solubility and dissolution testing were performed in three types of dissolution medium, and the studies demonstrated that the equilibrium solubilities of coenzyme Q10 nanocrystals and bulk drugs were not affected by the dissolution media but the kinetic solubilities were. Kinetic solubility curves and changes in particle size distribution were determined and well explained by the proposed solubilization model for the nanocrystals and bulk drugs. The particle size effect on dissolution was clearly influenced by the diffusion coefficients of the various dissolution media, and the dissolution velocity of coenzyme Q10 increased as particle size decreased. The bioavailability of coenzyme Q10 after oral administration in beagle dogs was improved by reducing the particle size. For 700 nm nanocrystals, the AUC0₋48 was 4.4-fold greater than that for the coarse suspensions, but a further decrease in particle size from 700 nm to 120 nm did not contribute to improvement in bioavailability until the particle size was reduced to 80 nm, when bioavailability was increased by 7.3-fold.

Repeated injection of PEGylated solid lipid nanoparticles induces accelerated blood clearance in mice and beagles.

Surface modification of nanocarriers with amphiphilic polymer polyethylene glycol (PEG), known as PEGylation, is regarded as a major breakthrough in the application of nanocarriers. However, PEGylated nanocarriers (including liposomes and polymeric nanoparticles) induce what is referred to as the "accelerated blood clearance (ABC) phenomenon" upon repeated injection and consequently they lose their sustained circulation characteristics. Despite this, the present authors are not aware of any reports of accelerated clearance due to repeated injection for PEGylated solid lipid nanoparticles (SLNs), another promising nanocarrier. This study investigated the pharmacokinetics of PEGylated SLNs upon repeated administration in mice; moreover, the impact of circulation time on the induction of the ABC phenomenon was studied in beagles for the first time. The ABC index, selected as the ratio of the area under the concentration-time curve from time 0 to the last measured concentration of a second injection to that of the first injection, was used to evaluate the extent of this phenomenon. Results showed that the PEGylated SLNs exhibited accelerated clearance from systemic circulation upon repeated injection, both in mice and in beagles, and the ratio for the different time intervals, which showed that the ABC index exhibited significant difference within 30 minutes following the second injection, was good enough to evaluate the magnitude of ABC. This ABC index indicated that the 10 mol% PEG SLNs with a suitable prolonged circulation time induced the most marked ABC phenomenon in this research. This study demonstrated that, like PEGylated nanocarriers such as liposomes and polymeric nanoparticles, PEGylated SLNs induced the ABC phenomenon upon repeated injection--the beagle was a valuable experimental animal for this research. Furthermore, the authors considered that a relatively extended circulation time of the initial dose may be the underlying major factor determining the induction of the ABC phenomenon.

A frustrating problem: accelerated blood clearance of PEGylated solid lipid nanoparticles following subcutaneous injection in rats.

Colloidal particles have preferential access to the lymphatic system following subcutaneous administration, achieving lymphatic targeting by drug accumulation in the regional lymph nodes. Moreover, the surface PEGylated colloidal particles have shown enhanced drainage into lymphatics and uptake by macrophages of the regional lymph nodes after subcutaneous injection. Nevertheless, it is reported that upon repeated intravenous injection, the PEG-specific IgM produced by the administration of the PEGylated colloidal particles markedly accelerates the clearance of subsequent doses of PEGylated particles. In this article, we report that the first subcutaneous injection of PEGylated solid lipid nanoparticles also induces the intravenously administered PEGylated particles to be cleared very rapidly from the circulation, and the "ABC index," a parameter for the intensity of accelerated blood clearance, for subcutaneous injection was equivalent to or even lower than that following the first intravenous injection. Moreover, the small quantities of distributed particles in the spleen after the first s.c. dose but the significantly higher elimination rate of the second i.v. dose, strongly suggest that, in addition to the spleen, the regional lymph nodes also play a promotive role in this phenomenon, although the exact lymphocytes causing this phenomenon remain unclear. Our observations may thus have important implications for considering combination therapy with PEGylated productions requiring different administration routes such as intravenous and subcutaneous injection, and great care is needed.