Enhanced Tumor Targeting of Doxorubicin by Ganglioside GMl-bearing Long-circulating Liposomes

Abstract

Doxorubicin (DXR) was encapsulated in long-circulating liposomes, composed of ganglioside GM1 (GM1)/distearoylphosphatidylcholine (DSPC)/cholesterol (CH) (0.13:1:1 in molar ratio) and sized to approximately 100 nm in mean diameter, with 98% entrapping efficiency by the transmembrane pH gradient method. Free DXR, DXR-DSPC/CH and DXR-GM1/DSPC/CH liposomes were injected intravenously into Colon 26 tumor-bearing Balb/c mice via the tail vein at a dose of 5.0 mg DXR/kg. DXR-GM1/DSPC/CH liposomes gave a higher blood level of the drug than did DXR-DSPC/CH liposomes or free DXR up to 24 hours after injection, and the area under the blood concentration-time curve (AUC) for DXR-GM1/DSPC/CH liposomes was 1.5 or 526 times higher than that for DXR-DSPC/CH liposomes or free DXR, respectively. DXR-GM1/DSPC/CH liposomes gave a decreased DXR concentration in the reticuloendothelial system (RES) of the liver and the spleen. Both liposomal formulations effectively reduced the DXR concentration in the heart as compared with that in the case of free DXR. At 6 hours after i.v. injection, DXR-GM1/DSPC/CH liposomes provided an approximately 3.3- or 9-fold higher peak DXR level in the tumor as compared with DXR-DSPC/CH liposomes or the free drug, respectively. These high tumor levels of DXR appear to reflect the prolonged residence time of the liposomes. The results suggest that encapsulation of DXR in GM1-bearing long-circulating liposomes will be useful for cancer chemotherapy.     

Targeted thermosensitive liposomes: an attractive novel approach for increased drug delivery to solid tumors

Introduction: Currently available chemotherapy is hampered by a lack in tumor specificity and resulting toxicity. Small and long-circulating liposomes can preferentially deliver chemotherapeutic drugs to tumors upon extravasation from tumor vasculature. Although clinically used liposomal formulations demonstrated significant reduction in toxicity, enhancement of therapeutic activity has not fully met expectations.

Areas covered: Low drug bioavailability from liposomal formulations and limited tumor accumulation remain major challenges to further improve therapeutic activity of liposomal chemotherapy. The aim of this review is to highlight strategies addressing these challenges. A first strategy uses hyperthermia and thermosensitive liposomes to improve tumor accumulation and trigger liposomal drug bioavailability. Image-guidance can aid online monitoring of heat and drug delivery and further personalize the treatment. A second strategy involves tumor-specific targeting to enhance drug delivery specificity and drug internalization. In addition, we review the potential of combinations of the two in one targeted thermosensitive-triggered drug delivery system.

Expert opinion: Heat-triggered drug delivery using thermosensitive liposomes as well as the use of tumor vasculature or tumor cell-targeted liposomes are both promising strategies to improve liposomal chemotherapy. Preclinical evidence has been encouraging and both strategies are currently undergoing clinical evaluation. A combination of both strategies rendering targeted thermosensitive liposomes (TTSL) may appear as a new and attractive approach promoting tumor drug delivery.     

Enhanced Delivery and Antitumor Activity of Doxorubicin Using Long-Circulating Thermosensitive Liposomes Containing Amphipathic Polyethylene Glycol in Combination with Local Hyperthermia

Enhanced delivery of doxorubicin (DXR) to a solid tumor subjected to local hyperthermia was achieved by using long-circulating, thermosensitive liposomes (TSL) composed of dipalmitoyl phosphatidylcholine (DPPC)/distearoyl phosphatidylcholine (DSPC) (9:1, m/m) and 3 mol% amphipathic polyethylene glycol (PEG) in colon 26-bearing mice. Inclusion of 3 mol% of distearoyl phosphatidylethanolamine derivatives of PEG (DSPE-PEG, amphipathic PEG) with a mean molecular weight of 1000 or 5000 in DPPC/DSPC liposomes resulted in decreased reticuloendothelial system (RES) uptake and a concomitant prolongation of circulation time, affording sustained increased blood levels of the liposomes. Concomitantly, DXR levels in blood were also kept high over a long period. The presence of amphipathic PEG did not interfere with the encapsulation of DXR by the pH gradient method (>90% trapping efficiency) or with the temperature-dependent drug release from the liposomes. The optimal size of these liposomes was 180 – 200 nm in mean diameter for thermosensitive drug release and prolonged circulation time. The DXR levels in the tumor after injection of long-circulating TSL (DXR-PEG1000TSL or DXR-PEG5000TSL, at a dose of 5 mg DXR/ kg) with local hyperthermia were much higher than after treatment with DXR-TSL lacking PEG or with free DXR, reaching 7.0 – 8.5 DXR µg/g tumor (approximately 2 times or 6 times higher than that of DXR-TSL or free DXR, respectively). Furthermore, the combination of DXR-PEGTSL and hyperthermia effectively retarded tumor growth and increased survival time. Our results indicate that the combination of drug-loaded, long-circulating, thermosensitive liposomes with local hyperthermia at the tumor site could be clinically useful for delivering a wide range of chemotherapeutic agents in the treatment of solid tumors.     

Cell Culture and Animal Studies for Intracerebral Delivery of Borocaptate in Liposomal Formulation

The efficacy of a liposomal formulation for intracerebral delivery of borocaptate (BSH) to brain tumor cells has been investigated using cell culture to study BSH uptake and persistence and using tumor-bearing rats to determine BSH distribution in the brain. During a 16-hr incubation, cellular uptake of BSH solution or BSH liposomal formulation was similar. However, the cellular persistence of BSH greatly increased when BSH was present in liposome. The differences in cellular persistence for BSH solution and BSH-loaded liposomes were significant both in 12-hr and 24-hr incubation experiments (p < 0.05 and p < 0.01, respectively). For the studies involving tumor-bearing rats, BSH level in tumor tissue was significantly higher than that in normal brain tissue at 2 hr and 6 hr after intracerebral injection of BSH-loaded liposomes (p < 0.01). Our study indicated that the liposomal formulation enhanced cellular persistence of BSH in tumor cells and therefore favored the boron accumulation in the cells. With the prolonged physical retention of liposomes at the local injection site and the cellular retention of BSH enhanced by the liposomes, the intracerebral delivery of BSH using liposomal formulation may provide an effective boron delivery approach for boron neutron capture therapy of brain tumors.     

Enhanced Therapeutic Effect of Cytidine-5′ -Diphosphate Choline when Associated with GM1 Containing Small Liposomes as Demonstrated in a Rat Ischemia Model

Purpose. Cytidine-5-diphosphate choline (CDPc) was encapsulated in long-circulating unilamellar vesicles (SUVs) to improve the drug's biological effectiveness. Methods. SUVs made up of diaplmitoylphosphatidylcholine/diaplmitoylphosphatidylserine/ cholesterol (7:4:7 molar ratio) and 8 mol % of ganglioside G_Ml were prepared by extrusion through polycarbonate filters (mean diameter 50 nm). The formulation effectiveness was evaluated by an in vivo model of cerebral ischemia on Wistar rats. Results. The enhanced delivery of CDPc into the brain improved the therapeutic effectiveness of the drug. CDPc-loaded SUVs improved the survival rate of ischemized and reperfused Wistar rats (320-350 g) by -66% compared with the free drug. Liposome formulation was also able to effectively protect the brain against peroxidative damage caused by post-ischemic reperfusion. SUVs lowered the conjugated diene levels of the cerebral cortex. The liposomal delivery system did not alter the distribution patterns in the various cerebral lipid fractions of the drug, radiolabeled with ¹⁴C-CDPc. Conclusions. CDPc-loaded SUVs were able to protect the brain against damage induced by ischemia. A possible clinical application is envisaged.     

Accumulation of Protein-Loaded Long-Circulating Micelles and Liposomes in Subcutaneous Lewis Lung Carcinoma in Mice

Purpose. The purpose of our work was to compare the biodistribution and tumor accumulation of a liposome- or micelle-incorporated protein in mice bearing subcutaneously-established Lewis lung carcinoma. Methods. A model protein, soybean trypsin inhibitor (STI) was modified with a hydrophobic residue of N-glutaryl-phosphatidyl-ethanolamine (NGPE) and incorporated into both polyethyleneglycol(MW 5000)-distearoyl phosphatidyl ethanolamine (PEG-DSPE) micelles (< 20 nm) and PEG-DSPE-modified long-circulating liposomes (ca. 100 nm). The protein was labeled with ¹¹¹In via protein-attached diethylene triamine pentaacetic acid (DTPA), and samples of STI-containing liposomes or micelles were injected via the tail vein into mice bearing subcutaneously-established Lewis lung carcinoma. At appropriate time points, mice were sacrified and the radioactivity accumulated in the tumor and main organs was determined. Results. STI incorporated into PEG-lipid micelles accumulates in sub-cutaneously established Lewis lung carcinoma in mice better than the same protein anchored in long-circulating PEG-liposomes. Conclusions. Small-sized long-circulating delivery systems, such as PEG-lipid micelles, are more efficient in the delivery of protein to Lewis lung carcinoma than larger long-circulating liposomes.     

Effective delivery of an angiogenesis inhibitor by neovessel-targeted liposomes

Angiogenesis is critical for tumor growth and metastasis, and several angiogenesis inhibitors have been developed for the treatment of cancer. Previously, we identified angiogenic vessel-homing peptide, Ala-Pro-Arg-Pro-Gly (APRPG), by use of a phage-displayed peptide library. APRPG peptide-modified liposomes have been revealed to be useful for the delivery of encapsulated drugs to angiogenic vasculature in tumor-bearing animals. In the present study, to assess the usefulness of APRPG-PEG-modified liposomes as a carrier of angiogenesis inhibitors in vitro and in vivo, we designed and validated APRPG-PEG-modified liposomal angiogenesis inhibitor. SU1498, an inhibitor of vascular endothelial growth factor (VEGF) receptor tyrosine kinase, was successfully encapsulated into the liposomes. APRPG-PEG-modified liposomal SU1498 inhibited VEGF-stimulated endothelial cell proliferation in vitro. Moreover, APRPG-PEG-modified liposomal SU1498 significantly decreased tumor microvessel density in Colon26 NL-17 cell-bearing mice and prolonged the survival time of the mice. These findings suggest that APRPG-PEG-modified liposomes effectively deliver SU1498 to angiogenic endothelial cells in tumors and thus inhibit tumor-induced angiogenesis.     

Drug Delivery and Transport to Solid Tumors

Purpose. The purpose of this review is to provide an overview of the principles of and barriers to drug transport and delivery to solid tumors. Methods. This review consists of four parts. Part I provides an overview of the differences in the vasculature in normal and tumor tissues, and the relationship between tumor vasculature and drug transport. Part II describes the determinants of transport of drugs and particles across tumor vasculature into surrounding tumor tissues. Part III discusses the determinants and barriers of drug transport, accumulation, and retention in tumors. Part IV summarizes the experimental approaches used to enhance drug delivery and transport in solid tumors. Results. Drug delivery to solid tumors consists of multiple processes, including transport via blood vessels, transvascular transport, and transport through interstitial spaces. These processes are dynamic and change with time and tumor properties and are affected by multiple physicochemical factors of a drug, multiple tumor biologic factors, and as a consequence of drug treatments. The biologic factors, in turn, have opposing effects on one or more processes in the delivery of drugs to solid tumors. Conclusion. The effectiveness of cancer therapy depends in part on adequate delivery of the therapeutic agents to tumor cells. A better understanding of the processes and contribution of these factors governing drug delivery may lead to new cancer therapeutic strategies.     

Intravenous Cereport (RMP-7) Enhances Delivery of Hydrophilic Chemotherapeutics and Increases Survival in Rats with Metastatic Tumors in the Brain

Purpose. The following experiments determined whether intravenous infusions of Cereport enhance delivery of chemotherapeutics and prolong survival in rats with metastatic tumors in the brain. Methods. Autoradiography and scintillation were used to examine uptake of the lipophilic (paclitaxel and carmustine) and the hydrophilic (carboplatin) chemotherapeutic agents, as well as the large hydrophilic marker, 70 kDa dextran. Cereport was also tested in combination with the chemotherapeutic drugs carboplatin, vinorelbine, gemcitabine and carmustine to determine if Cereport could enhance the survival benefit beyond that provided by chemotherapy alone. Results. Cereport enhanced the uptake of carboplatin and dextran, but not paclitaxel or carmustine. The pattern of Cereport's uptake effect with carboplatin revealed that Cereport selectively increased the proportion of highly permeable regions. Survival was significantly enhanced when Cereport was combined with either carboplatin, vinorelbine, or gemcitabine, but not carmustine, compared to each chemotherapeutic agent alone. Conclusions. These data provide the first evidence that Cereport, or any receptor-mediated approach intended to enhance the permeability of the blood-brain tumor barrier, can increase the delivery hydrophilic drugs to metastatic tumors in the brain, increasing survival in tumor-bearing rats.     

Direct measurement of the extravasation of polyethyleneglycol-coated liposomes into solid tumor tissue by in vivo fluorescence microscopy

The extravasation of liposomes of different sizes into solid tumors after i.v. injection was visualized by in vivo fluorescence microscopy in mouse neuroblastoma C-1300-bearing mice. Liposomes composed of distearoylphosphatidylcholine/cholesterol (1/1 molar ratio) and 6 mol% distearoylphosphatidylethanolamine derivative of polyethyleneglycol (PEG) were prepared. The PEG-coated liposomes were fluorescently labeled with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI) as a liposome marker or with doxorubicin (DXR) as an aqueous-phase marker. Liposomes with an average diameter of 100–200 nm showed the greatest tumor accumulation. With time after injection of DiI-labeled liposomes, the tumor interstitial fluorescence intensity increased. Most fluorescent spots were located outside and around the vessel wall, indicating extravasation of intact liposomes. The perivascular distribution was heterogeneous. We also obtained the same fluorescence localization pattern with DXR released from extravasated liposomes after injection of DXR-encapsulated liposomes. No fluorescence from extravasated liposomes was detected in normal s.c. tissue; the fluorescent spots were observed only in the vessel wall. Our results indicate that small-size long-circulating liposomes are able to traverse the endothelium of blood vessels in tumors and extravasate into interstitial spaces. Moreover, encapsulated drug was released from extravasated liposomes in the tumor.     

Pharmacokinetics of Differently Designed Immunoliposome Formulations in Rats with Or Without Hepatic Colon Cancer Metastases

Purpose. Compare pharmacokinetics of tumor-directed immunoliposomes in healthy and tumor-bearing rats (hepatic colon cancer metastases). Methods. A tumor cell-specific monoclonal antibody was attached to polyethyleneglycol-stabilized liposomes, either in a random orientation via a lipid anchor (MPB-PEG-liposomes) or uniformly oriented at the distal end of the PEG chains (Hz-PEG-liposomes). Pharmacokinetics and tissue distribution were determined using [³H]-cholesteryloleylether or bilayer-anchored 5-fluoro[³H]deoxyuridine-dipalmitate ([³H]FUdR-dP) as a marker. Results. In healthy animals clearance of PEG-(immuno)liposomes was almost log-linear and only slightly affected by antibody attachment; in tumor-bearing animals all liposomes displayed biphasic clearance. In normal and tumor animals blood elimination increased with increasing antibody density; particularly for the Hz-PEG-liposomes, and was accompanied by increased hepatic uptake, probably due to increased numbers of macrophages induced by tumor growth. The presence of antibodies on the liposomes enhanced tumor accumulation: uptake per gram tumor tissue (2-4% of dose) was similar to that of liver. Remarkably, this applied to tumor-specific and irrelevant antibody. Increased immunoliposome uptake by trypsin-treated Kupffer cells implicated involvement of high-affinity Fc-receptors on activated macrophages. Conclusions. Tumor growth and immunoliposome characteristics (antibody density and orientation) determine immunoliposome pharmacokinetics. Although with a long-circulating immunoliposome formulation, efficiently retaining the prodrug FUdR-dP, we achieved enhanced uptake by hepatic metastases, this was probably not mediated by specific interaction with the tumor cells, but rather by tumor-associated macrophages.     

Targeted delivery of antibody-based therapeutic and imaging agents to CNS tumors: crossing the blood–brain barrier divide

Introduction: Brain tumors are inherently difficult to treat in large part due to the cellular blood–brain barriers (BBBs) that limit the delivery of therapeutics to the tumor tissue from the systemic circulation. Virtually no large molecules, including antibody-based proteins, can penetrate the BBB. With antibodies fast becoming attractive ligands for highly specific molecular targeting to tumor antigens, a variety of methods are being investigated to enhance the access of these agents to intracranial tumors for imaging or therapeutic applications.

Areas covered: This review describes the characteristics of the BBB and the vasculature in brain tumors, described as the blood–brain tumor barrier (BBTB). Antibodies targeted to molecular markers of central nervous system (CNS) tumors will be highlighted, and current strategies for enhancing the delivery of antibodies across these cellular barriers into the brain parenchyma to the tumor will be discussed. Noninvasive imaging approaches to assess BBB/BBTB permeability and/or antibody targeting will be presented as a means of guiding the optimal delivery of targeted agents to brain tumors.

Expert opinion: Preclinical and clinical studies highlight the potential of several approaches in increasing brain tumor delivery across the BBB divide. However, each carries its own risks and challenges. There is tremendous potential in using neuroimaging strategies to assist in understanding and defining the challenges to translating and optimizing molecularly targeted antibody delivery to CNS tumors to improve clinical outcomes.     

In vitro evaluation and biodistribution of HER2-targeted liposomes loaded with an 125I-labelled DNA-intercalator

Background: Increasing attention is currently focussed on the issue of finding strategies for the delivery of Auger-electron-emitting radionuclides into tumor cell nuclei.

Purpose: In this study, we investigated tumor-cell uptake and cell-killing ability in vitro as well as in vivo biodistribution of an ¹²⁵I-labelled anthracycline derivative administered by means of HER2-targeted liposomes.

Methods: Anthracycline derivative Comp1 was radiolabelled with Auger-emitting ¹²⁵I and encapsulated in liposomes (DSPC:Chol:DSPE-PEG) using pH-gradient loading. Single-chain fragment F5 was anchored to the liposomes as targeting device for HER2. Uptake and specificity of ¹²⁵I-Comp1 delivered via targeting and non-targeting liposomes were analysed in cultured HER2-overexpressing cells. Cell-killing efficacy was evaluated in SKOV3 cells and biodistribution for up to 48?h was studied after intraperitoneal injection in tumor-bearing female BALB/c nu/nu mice.

Results: ¹²⁵I-Comp1 was specifically taken up by the cultured cells when administered by means of HER2-targeted liposomes and a clear dose-effect correlation in survival of cells was seen with increasing specific activity. The biodistribution studies revealed that ¹²⁵I-Comp1 accumulated in tumors when distributed using HER2-targeted liposomes and that this effect was absent when using non-targeting liposomes.

Conclusion: The HER2-targeted liposomes possess the properties needed to bring about tumor-specific delivery and therapeutic effect of ¹²⁵I-Comp1.     

Cell culture and animal studies for intracerebral delivery of borocaptate in liposomal formulation

The efficacy of a liposomal formulation for intracerebral delivery of borocaptate (BSH) to brain tumor cells has been investigated using cell culture to study BSH uptake and persistence and using tumor-bearing rats to determine BSH distribution in the brain. During a 16-hr incubation, cellular uptake of BSH solution or BSH liposomal formulation was similar. However, the cellular persistence of BSH greatly increased when BSH was present in liposome. The differences in cellular persistence for BSH solution and BSH-loaded liposomes were significant both in 12-hr and 24-hr incubation experiments (p < 0.05 and p < 0.01, respectively). For the studies involving tumor-bearing rats, BSH level in tumor tissue was significantly higher than that in normal brain tissue at 2 hr and 6 hr after intracerebral injection of BSH-loaded liposomes (p < 0.01). Our study indicated that the liposomal formulation enhanced cellular persistence of BSH in tumor cells and therefore favored the boron accumulation in the cells. With the prolonged physical retention of liposomes at the local injection site and the cellular retention of BSH enhanced by the liposomes, the intracerebral delivery of BSH using liposomal formulation may provide an effective boron delivery approach for boron neutron capture therapy of brain tumors.     

Ultrasound enhanced antitumor activity of liposomal doxorubicin in mice

Liposomal encapsulation of doxorubicin (DXR) improves tumor accumulation and reduces adverse effects. One possible strategy for further optimization of this delivery technology would be to design the liposome carrier to release its content within the tumor tissue in response to specific stimuli such as ultrasound (US). In this study, the tumor uptake properties and therapeutic efficacy of 1,2 distearoyl-sn-glycero-3-phosphatidylethanolamine-based liposomes containing DXR were investigated in nude mice bearing tumor xenografts. The liposomal DXR formulation alone showed no inhibitory effect on tumor growth. However, upon exposure to low frequency US in situ inhibition of tumor growth was demonstrated.     

Therapeutic efficacy of the combination of doxorubicin-loaded liposomes with inertial cavitation generated by confocal ultrasound in AT2 Dunning rat tumour model

Abstract

The combination of liposomal doxorubicin (DXR) and confocal ultrasound (US) was investigated for the enhancement of drug delivery in a rat tumour model. The liposomes, based on the unsaturated phospholipid dierucoylphosphocholine, were designed to be stable during blood circulation in order to maximize accumulation in tumour tissue and to release drug content upon US stimulation. A confocal US setup was developed for delivering inertial cavitation to tumours in a well-controlled and reproducible manner. In vitro studies confirm drug release from liposomes as a function of inertial cavitation dose, while in vivo pharmacokinetic studies show long blood circulation times and peak tumour accumulation at 24–48?h post intravenous administration. Animals injected 6?mg kg^?1 liposomal DXR exposed to US treatment 48?h after administration show significant tumour growth delay compared to control groups. A liposomal DXR dose of 3?mg kg^?1, however, did not induce any significant therapeutic response. This study demonstrates that inertial cavitation can be generated in such a fashion as to disrupt drug carrying liposomes which have accumulated in the tumour, and thereby increase therapeutic effect with a minimum direct effect on the tissue. Such an approach is an important step towards a therapeutic application of cavitation-induced drug delivery and reduced chemotherapy toxicity.     

Delivery of contrast agents for positron emission tomography imaging by liposomes

Liposomes encapuslating positron emitters are applicable for diagnostic imaging and are useful to investigate the real-time liposomal trafficking in vivo. Long-circulating liposomes encapsulaing [2-(18)F]-2-fluoro-2-deoxyglucose were administrated to tumor-bearing mice, and a PET scan was performed. Small-sized long-circulating liposomes (100 nm) tended to accumulate in tumor tissues of tumor-bearing mice as compared with conventional liposomes. Then the size effect on trafficking of long-circulating liposomes was investigated. Large-sized liposomes (>300 nm) accumulated in liver and spleen in a time dependent manner. On the contrary, small-sized ones (<200 nm) were transiently accumulated in the liver right after injection, but the accumulation decreased time dependently, suggesting that, although the majority of small long-circulating liposomes remain in bloodstream, some extravasate once into interstitial spaces in liver which re-enter into bloodstream again. Next the trafficking of so-called long-circulating liposomes, i.e., liposomes modified with ganglioside GM1, palmityl glucuronide (PGlcUA), and polyethylene glycol (PEG), in tumor-bearing mice was examined. The accumulation of all three kinds of long-circulating liposomes in liver decreased time-dependently, and PGlcUA-liposomes could avoid liver-trapping the most efficiently. Tumor accumulation of liposomes was obvious for PGlcUA-liposomes and PEG-liposomes from immediately after injection, but not for GM1-liposomes. Finally, the trafficking of differently charged liposomes was investigated in normal mice. The accumulation of positively charged liposomes containing 1,2-dimyristyloxypropyl-3-dimethyl-hydroxyethyl bromide was different from that of neutral and negatively charged DCP-liposomes. The agglutinability of and serum protein ginding to positively charged liposomes were marked, suggesting that these factors affect the high accumulation of DMRIE-liposomes in liver. Non-invasive PET analysis of liposomal trafficking is beneficial for obtaining information about liposomal drug delivery, and long-circulating liposomes might be useful for diagnostic tumor imaging by PET.     

Use of polyglycerol (PG), instead of polyethylene glycol (PEG), prevents induction of the accelerated blood clearance phenomenon against long-circulating liposomes upon repeated administration

The accelerated blood clearance (ABC) phenomenon accounts for the rapid systemic clearance of PEGylated nanocarriers upon repeated administrations. IgM production against the polyethylene glycol (PEG) coating in PEGylated liposomes is now known to be responsible for such unexpected pharmacokinetical alterations. The ABC phenomenon poses a remarkable clinical challenge by reducing the therapeutic efficacy of encapsulated drugs and causing harmful effects due to the altered tissue distribution pattern of the drugs. In this study, we investigated the in vivo performance of liposomes modified with polyglycerol (PG) upon repeated injection, and the in vivo therapeutic efficacy of such liposomes when they encapsulated a cytotoxic agent, doxorubicin (DXR). Repeated injection of PEG-coated liposomes in rats induced the ABC phenomenon, while repeated injection of PG-coated liposomes did not. In addition, DXR-containing PG-coated liposomes showed antitumor activity that was superior to that of free DXR and similar to that of DXR-containing PEG-coated liposomes upon repeated administration. These results indicate that polyglycerol (PG) might represent a promising alternative to PEG via enhancing the in vivo performance of liposomes by not eliciting the ABC phenomenon upon repeated administration.     

地塞米松脂质体的制备及其对乳腺癌作用的体内外研究

目的制备地塞米松脂质体,探讨地塞米松对乳腺癌4T1细胞的生长抑制作用及对荷瘤鼠的抗肿瘤药效。方法采用薄膜分散–超声法,以粒径和多分散指数(PDI)为指标进行单因素实验考察了大豆磷脂(SPC)与甲氧基聚乙二醇磷脂(DSPE-mPEG2000)的质量比、SPC与地塞米松的质量比、超声时间对地塞米松脂质体粒径的影响从而筛选得到最佳处方和最佳工艺条件。采用MTT法比较地塞米松注射液和地塞米松脂质体对4T1细胞的作用。建立4T1 BAL B/c荷瘤小鼠模型,研究地塞米松脂质体对4T1荷瘤小鼠的体内抗肿瘤作用。结果当SPC与DSPE-mPEG2000质量比为5∶1、SPC与地塞米松质量比为50∶3、超声时间为20 min时制备得到的脂质体粒径最小,粒径分布最窄,室温放置15 d稳定,于生理介质中稳定。MTT测定结果显示地塞米松注射液和脂质体对4T1细胞生长抑制作用均较弱,但在4T1荷瘤鼠的体内实验中,在5mg/kg的给药剂量下,地塞米松脂质体的抑瘤率却高达78.9%,显著高于地塞米松注射液(33.4%,P<0.05)和8mg/kg紫杉醇注射液(55%,P<0.05)。结论制备的地塞米松脂质体放置于生理介质中均能稳定存在,能口服能静脉给药。地塞米松脂质体对4T1荷瘤小鼠肿瘤生长有较强的抑制作用,但体外对4T1细胞抑制抑制作用并不强,推测地塞米松脂质体是通过调节肿瘤微环境来抑制肿瘤生长。     

Glutathione PEGylated liposomes: pharmacokinetics and delivery of cargo across the blood–brain barrier in rats

Abstract

Partly due to poor blood–brain barrier drug penetration the treatment options for many brain diseases are limited. To safely enhance drug delivery to the brain, glutathione PEGylated liposomes (G-Technology®) were developed. In this study, in rats, we compared the pharmacokinetics and organ distribution of GSH-PEG liposomes using an autoquenched fluorescent tracer after intraperitoneal administration and intravenous administration. Although the appearance of liposomes in the circulation was much slower after intraperitoneal administration, comparable maximum levels of long circulating liposomes were found between 4 and 24?h after injection. Furthermore, 24?h after injection a similar tissue distribution was found. To investigate the effect of GSH coating on brain delivery in vitro uptake studies in rat brain endothelial cells (RBE4) and an in vivo brain microdialysis study in rats were used. Significantly more fluorescent tracer was found in RBE4 cell homogenates incubated with GSH-PEG liposomes compared to non-targeted PEG liposomes (1.8-fold, p?<?0.001). In the microdialysis study 4-fold higher (p?<?0.001) brain levels of fluorescent tracer were found after intravenous injection of GSH-PEG liposomes compared with PEG control liposomes. The results support further investigation into the versatility of GSH-PEG liposomes for enhanced drug delivery to the brain within a tolerable therapeutic window.