Vascular targeting of doxorubicin using cationic liposomes

Tumor vessel has been recognized as an important target for anticancer therapy. Cationic liposomes have been shown to selectively target tumor endothelial cells, thus can potentially be used as a carrier for chemotherapy agents. In this study, cationic liposomes containing 20 mol% cationic lipid dimethyl dioctadecyl ammonium bromide (DDAB) and loaded with doxorubicin (DOX) were prepared and characterized. The cationic liposomal DOX showed 10.8 and 9.1 times greater cytotoxicity than control PEGylated liposomal DOX in KB oral carcinoma and L1210 murine lymphocytic leukemia cells, and 7.7- and 6.8-fold greater cytotoxicity compared to control neutral non-PEGylated liposomal DOX, repectively, in these two cell lines. Although cationic liposomal DOX had higher tumor accumulation at 30 min after intravenous administration compared to control liposomes (p<0.05), DOX uptake of these liposomes at 24h post-injection was similar to that of PEGylated liposomal DOX (p>0.05) and approximately twice the levels of the free drug and non-PEGylated liposomes. In a murine tumor model generated using L1210 cells, increased survival rate was obtained with cationic liposomal DOX treatment compared to free DOX (p<0.01), neutral liposome control (p<0.01), as well as PEGylated liposomes (p<0.05). In conclusion, the cationic liposomal DOX formulation produced superior in vitro cytotoxicity and in vivo antitumor activity, and warrants further investigation.     

Novel tetrapeptide, RGDF, mediated tumor specific liposomal doxorubicin (DOX) preparations

Arginine-glycine-aspartate (RGD) has been shown to possess a strong affinity for the integrins overexpressed in tumor cells, especially during tumor invasion, angiogenesis and metasis. Based on work from others, a novel tetrapeptide, arginine-glycine-aspartate-phenylanaline (RGDF), has been designed and studied as a homing device to direct liposomal doxorubicin (DOX) to tumor cells in this work. In order to incorporate RGDF into liposomal DOX preparations, RGDF was conjugated with three different fatty alcohols to achieve RGDF-fatty alcohol conjugates. Glycine-glycine-aspartate-phenylanaline (GGDF)-lauryl alcohol conjugate was synthesized as a negative control. RGDF-fatty alcohol conjugates (RGDFO(CH(2))(n)CH(3)) and GGDF-lauryl alcohol conjugate (L-GGDFC12-DOX) incorporated liposomal preparations were obtained by first preparing liposomes using the film dispersion method followed by loading DOX using a transmembrane pH gradient method. Because of their amphipathic nature, RGDF- or GGDF-fatty alcohol conjugates are expected to be readily incorporated into liposomes with their fatty alkanyl chains being intercalated between fatty acyl chains of liposomal bilayers and the hydrophilic peptide moiety (RGDF or GGDF) being anchored on the surface of liposomes. The particle size and zeta potential of liposomal DOX preparations containing RGDF-fatty alcohol conjugate (L-RGDF-DOXs) or L-GGDFC12-DOX were measured, and their morphology was studied using transmission electron microscopy. In vitro DOX release profile from RGDF incorporated liposomal DOX was measured. The antitumor activities of RGDF incorporated liposomal DOX preparations were evaluated in ICR mice inoculated with sarcoma S(180), which is known to express α(v)β(3) integrin. Both conventional liposomal DOX preparation (L-DOX) without RGDFO(CH(2))(n)CH(3) and L-GGDFC12-DOX were used as negative controls. Our results showed improved tumor growth inhibition with L-RGDF-DOXs over doxorubicin hydrochloride solution, L-DOX and L-GGDFC12-DOX. Pathological examination of tumor biopsy demonstrated that L-RGDF-DOXs induced enhanced tumor cell death in comparison to negative controls. Pharmacokinetic studies showed that the concentrations of DOX found in tumor sites were increased by 1.7-4.5-fold when liposomal DOX preparation containing RGDF-lauryl alcohol conjugate (L-RGDFC12-DOX) was administered in comparison to when L-GGDFC12-DOX or doxorubicin hydrochloride solution was administered. The concentrations of DOX found in the heart, which is the main site of toxic effects of DOX, were significantly reduced when L-RGDFC12-DOX was administered in comparison to when L-GGDFC12-DOX or doxorubicin hydrochloride solution was administered.     

Effects of intraperitoneal administration of liposomes and methods of preparing liposomes for local therapy.

Many different methods of preparing liposomes exist, but the biological and physical properties of these preparations are not known. We therefore investigated the physical properties of liposomal doxorubicin (DOX) and found it to be most effective when administered intraperitoneally for carcinoma in the abdominal cavity. Liposomal DOX was prepared in three ways, by the Bangham method (BLDOX), pH gradient method, and gelation method. We investigated the physical properties of each preparation. And then we investigated the effects of the liposomes and liposomal lipids on the uptake of DOX by carcinoma cells in vitro and on the survival of Ehrlich ascites carcinoma-bearing mice in vivo. The uptake of DOX by the cells differed significantly with each liposome in vitro. The physical properties of the liposomes, including liposomal membrane lipids, size, zeta potentials and fluidity of liposomal membrane, were not so different, but the leak level of entrapped DOX from the liposomes was. Furthermore, the survival of ascites tumor-bearing mice also differed with each liposome preparation, DMPC containing BLDOX being the most effective when administered intraperitoneally. The method of preparation is an important factor affecting the properties of liposomes, and for local therapy, DMPC containing BLDOX is most effective because of its leaky property.     

Increase of therapeutic activity of doxorubicin by long circulating liposomes in combination with curcumin

In this study, doxorubicin (DOX)-loaded long circulating liposomes combined with curcumin (CUR) (DOX-CUR-LCLs) were successfully prepared as a novel formulation for cancer treatment. The particle size and distribution, zeta potential, drug loading capacity, and entrapment efficiency (EE) of the preparation were characterized. The in vitro anti-tumor activities of DOX-CUR-LCLs and DOX-LCLs against A549 cells were then evaluated and compared with that of free DOX. Cytotoxicity evaluation showed that DOX-CUR-LCLs had a significantly higher antitumor activity than other DOX preparations. These results suggest that novel DOX-CUR-LCLs, combination of DOX and CUR administered in long-circulating liposomes, could improve antitumor activity.     

In vivo distribution and antitumor activity of heparin-stabilized doxorubicin-loaded liposomes

The purpose of this study was to investigate the effect of heparin conjugation to the surface of doxorubicin (DOX)-loaded liposomes on the circulation time, biodistribution and antitumor activity after intravenous injection in murine B16F10 melanoma tumor-bearing mice. The heparin-conjugated liposomes (heparin-liposomes) were prepared by fixation of the negatively charged heparin to the positively charged liposomes. The existence of heparin on the liposomal surface was confirmed by measuring the changes in the particle size, zeta potential and heparin amount of the liposomes. The stability of the heparin-liposomes in serum was higher than that of the control liposomes, due to the heparin-liposomes being better protected from the adsorption of serum proteins. The DOX-loaded heparin-liposomes showed high drug levels for up to 64 h after the intravenous injection and the half-life of DOX was approximately 8.4- or 1.5-fold higher than that of the control liposomes or polyethyleneglycol-fixed liposomes (PEG-liposomes), respectively. The heparin-liposomes accumulated to a greater extent in the tumor than the control or PEG-liposomes as a result of their lower uptake by the reticuloendothelial system cells in the liver and spleen. In addition, the DOX-loaded heparin-liposomes retarded the growth of the tumor effectively compared with the control or PEG-liposomes. These results indicate the promising potential of heparin-liposomes as a new sterically stabilized liposomal delivery system for the enhancement of the therapeutic efficacy of chemotherapeutic agents.     

Enhanced Intracellular Uptake of Sterically Stabilized Liposomal Doxorubicin <Emphasis Type="Italic">in Vitro</Emphasis> Resulting in Improved Antitumor Activity <Emphasis Type="Italic">in Vivo</Emphasis>

Purpose To investigate the correlation between the in vitro intracellular uptake and the in vivo antitumor activity of anticancer drugs delivered by sterically stabilized liposomes (SSL).Methods Arginine-glycine-aspartic acid (RGD) peptide or RGD mimetic (RGDm) was coupled onto the surface of SSL to obtain the cell-binding carrier to facilitate the intracellular delivery of the encapsulated drugs. DOX-loaded SSL (SSL-DOX), DOX-loaded RGD-modified SSL (RGD-SSL-DOX) and DOX-loaded RGDm-modified SSL (RGDm-SSL-DOX) were prepared by lipid film dispersion followed by remote loading of DOX. The intracellular uptake of DOX from the various liposomal formulations was evaluated in vitro with melanoma B16 cells, and the pharmacokinetics, biodistribution, and antitumor activity were compared in C57BL/6 mice carrying melanoma B16 tumors.Results In vitro intracellular uptake of DOX by B16 cells and in vivo antitumor activity in terms of tumor growth inhibition and mice survival time prolongation for various liposomal DOX were in the following order: RGD-SSL-DOX > RGDm-SSL-DOX > SSL-DOX. The mean survival time of the mice treated with RGD-SSL-DOX, RGDm-SSL-DOX, and SSL-DOX was 55, 49, and 44 days, respectively. The three liposomal DOX formulations produced very close DOX accumulation in tumor, which is significantly higher than that of free DOX. RGD- or RGDm-SSL-DOX demonstrated prolonged circulation time similar to that of SSL-DOX, whereas they showed significantly lower DOX level in blood and remarkably higher uptake by spleen than SSL-DOX.Conclusions Enhanced intracellular uptake of DOX encapsulated in SSL could produce an improved therapeutic effect for the melanoma B16 tumors. Enhancing intracellular delivery of the anticancer drugs encapsulated in SSL may be a promising strategy to improve their therapeutic efficacy for solid tumors.     

Estrogen-functionalized liposomes grafted with glutathione-responsive sheddable chotooligosaccharides for the therapy of osteosarcoma

An estrogen (ES)-functionalized cationic liposomal system was developed and exploited for targeted delivery to osteosarcoma. Natural biocompatible chotooligosaccharides (COS, MW2-5?KDa) were covalently tethered to the liposomal surface through a disulfate bond (-SS-) to confer reduction-responsive COS detachment, whereas estrogen was grafted via polyethylene glycol (PEG 2?K) chain to achieve estrogen receptor-targeting. The liposomal carriers were prepared by the ethanol injection method and fluorescent anticancer drug doxorubicin (DOX) was loaded with ammonium sulfate gradient. The physicochemical properties, reduction-sensitivity, and the roles of estrogen on cellular uptake and tumor-targeting were studied. The Chol-SS-COS/ES/DOX liposomes were spherical with an average size about 110?nm, and high encapsulation efficiency. The liposomes were stable in physiological condition but rapidly release the payload in response to tumoral intracellular glutathione (20?mM). MTT cytotoxicity assay confirmed that Chol-SS-COS/ES/DOX liposomes exhibited higher cytotoxicity to MG63 osteosarcoma cells than to liver cells (LO2). Flow cytometry (FCM) and confocal laser scanning microscopy revealed that cellular uptake of Chol-SS-COS/ES/DOX liposomes by MG63, than the free DOX or Chol-SS-COS/DOX. Ex vivo fluorescence distribution study showed that the multifunctional liposomes selectively accumulated in the MG63 xenografts versus the organs. Chol-SS-COS/ES/DOX liposomes strongly inhibited the tumor growth and enhanced the animal survival rate. Overall, the COS grafted estrogen-functionalized cationic liposomes, fortified with glutathione-responsiveness, showed great potential for specific intracellular drug delivery to estrogen receptor-expressing tumors such as osteosarcoma.     

The effect of PEG coating on in vitro cytotoxicity and in vivo disposition of topotecan loaded liposomes in rats

Amphoteric drugs encapsulated in PEGylated liposomes may not show superior therapeutic antitumor activity due to increased leakage rate of these drugs in presence of PEG-lipids. In order to investigate the effect of PEG coating on in vitro and in vivo characteristics of topotecan loaded liposomes, an amphoteric anticancer drug, PEGylated and conventional liposomes were prepared by lipid film hydration method. Various properties of the prepared nanoliposomes such as encapsulation efficiency, size, zeta potential, physical stability as well as the chemical stability of lactone form of topotecan, cytotoxicity and topotecan pharmacokinetics were evaluated. In vitro cytotoxic activity was evaluated on murine Lewis lung carcinoma (LLC) and human mammary adenocarcinoma (BT20) cells. Pharmacokinetic was evaluated in Wistar rats after i.v. injection of topotecan, formulated in PBS pH 7.4 or in conventional or in PEGylated liposomes. The conventional liposome (CL) formulation was composed of DSPC/cholesterol/DSPG (molar ratio; 7:7:3), while for PEGylated liposome the composition was DSPC/cholesterol/DSPG/DSPE-PEG(2000) (molar ratio; 7:7:3:1.28). The size of both liposomes was around 100 nm with polydispersity index of about 0.1. In comparison with free drug, liposomal topotecan showed more stability for topotecan lactone form in vitro. Compared to free topotecan, PEGylated and conventional liposomes improved cytotoxic effect of topotecan against the two cancer cell line studied. The results of pharmacokinetic studies in rats showed that both CL and PEGylated liposomal formulations increased the concentration of total topotecan in plasma, however, initial concentration and the values of AUC, MRT and t(1/2 beta) were much higher (P<0.001) for PEGylated liposomal drug than for conventional one or free drug. PEGylated liposome resulted in a 52-fold and 2-fold increases in AUC(0-infinity) compared with that of free topotecan and CL, respectively. These results indicated that PEG modified liposome might be an effective carrier for topotecan.     

Reversal of multidrug resistance by transferrin-conjugated liposomes co-encapsulating doxorubicin and verapamil.

PURPOSE: Liposomes co-encapsulating doxorubicin (DOX) and verapamil (VER), and conjugated to transferrin (Tf-L-DOX/VER) were synthesized and evaluated in K562 leukemia cells. The design of this formulation was aimed at selective targeting of tumor cells, reducing cardiotoxicity of DOX and VER, as well as overcoming P-glycoprotein (Pgp)-mediated multidrug resistance (MDR) phenotype. METHODS: The liposomes were prepared by polycarbonate membrane extrusion, followed by pH-gradient driven remote loading and Tf conjugation. Kinetics of in vitro release of DOX and VER from liposomes was determined by measuring changes in the concentration of encapsulated drugs. Uptake of Tf-conjugated liposomes by K562 cells was evaluated by fluorescence microscopy and by fluorometry. Cytotoxicities of various formulations of DOX were determined by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolum bromide (MTT) assay. RESULTS: Efficiencies for liposomal loading of DOX and VER were 95% and 70%, respectively. The mean particle diameter for the liposomes was approximately 110nm. Rates of release for DOX and VER were similar in singly-loaded and co-loaded liposomes. Tf-L-DOX/VER showed efficient uptake by the TfR+ K562 cells. In DOX-resistant K562 cells (K562/DOX), Tf-L-DOX/VER showed 5.2 and 2.8 times greater cytotoxicity (IC50 = 4.18 muM) than non-targeted liposomes (L-DOX/VER) (IC50 = 21.7 muM) and Tf-targeted liposomes loaded with DOX alone (Tf-L-DOX) (IC50 = 11.5 muM), respectively. CONCLUSIONS: The combination of TfR targeting and co-encapsulation of DOX and VER was highly effective in overcoming drug resistance in K562 leukemia cells.     

Preparation and evaluation of doxorubicin-luminespib co-loaded multivesicular liposomes

A combinational therapeutic system that simultaneously administrates various pathways is preferred for anti-cancer treatment. In the present study, we successfully constructed a co-delivery system, multivesicular liposomes (MVLs) co-encapsulating doxorubicin (DOX) and luminespib (AUY922). A simple and accurate dual-wavelength spectrophotometric method was established for the determination of DOX and AUY922 in liposomal formulation. MVL-loading drugs were prepared by a multi-emulsion solvent evaporation method, which exhibited excellent physicochemical properties, such as particle size of 3–8 μm and high entrapment efficiency above 95% for DOX and 73% for AUY922. The synergetic cytotoxic effect for these two drugs was evaluated in MDA-MB-231 cells. The in vitro antitumor studies demonstrated the superior anti-proliferation activity of DOX and AUY922 with a combination index of 0.43, indicating a great synergistic effect. The experimental data suggested that combinational use of DOX and AUY922 within liposomes could be an effective way to develop efficient treatments of cancers.     

注射用多西他赛脂质体的制备及其体内外评价

 目的：制备多西他赛脂质体,对其体内外性质进行评价。方法：采用改良薄膜分散法结合冷冻干燥工艺制备多西他赛脂质体冻干粉;用激光粒度仪考察了脂质体的粒径分布和Zeta电位;超滤法测定了包封率;以市售多西他赛注射液为对照,比较体外释放、大鼠骨髓抑制和裸鼠体内肿瘤抑制情况。结果：多西他赛脂质体平均粒径为92 nm,Zeta电位为-52.3 mV,包封率＞95%,24 h累积释放率为84.1%;相同剂量下,多西他赛脂质体比多西他赛注射液的骨髓抑制作用降低,但二者对HT-29移植瘤的抑制作用相同。结论：本实验所制备的多西他赛脂质体包封率较高、稳定性较好;与多西他赛注射液相比,毒性较低,同时具有相同的抗肿瘤效力。      

In vitro cytotoxicity of Stealth liposomes co-encapsulating doxorubicin and verapamil on doxorubicin-resistant tumor cells

Multidrug resistance (MDR) is a major obstacle to successful clinical cancer chemotherapy. A novel doxorubicin anti-resistant Stealth liposomes (DARSLs), prepared by co-encapsulating doxorubicin (DOX) and verapamil (VER) into stealth liposomes, has been developed. The average particle size of DARSLs was 118.1+/-22.3 nm. Encapsulation efficiencies of DOX and VER in DARSLs were greater than 95% and 70%, respectively. The IC(50) of DARSLs as measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT) assay in multidrug resistant rat prostate cancer Mat-LyLu-B2 (MLLB2) cells was 0.079+/-0.017 microM, 13 fold less than that for liposomal DOX with free VER (LDFV 0.96+/-0.46 microM) but only about 2 times less than FDFV. The IC(50) cytotoxicity on MLLB2 cells of the various formulations was as follows: DARSLs approximately LDLV相似文献   

Development and evaluation of inhalational liposomal system of budesonide for better management of asthma

Budesonide is a corticosteroid used by inhalation in the prophylactic management of asthma. However, frequent dosing and adverse effects (local and systemic) remain a major concern in the use of budesonide. Liposomal systems for sustained pulmonary drug delivery have been particularly attractive because of their compatibility with lung surfactant components. In the present investigation, pulmonary liposomal delivery system of budesonide was prepared by film hydration method and evaluated for sustained release. Various parameters were optimized with respect to entrapment efficiency as well as particle size of budesonide liposomes. For better shelf life of budesonide liposomes, they were freeze dried using trehalose as cryoprotectant. The liposomes were characterized for entrapment efficiency, particle size, and surface topography; in vitro drug release was evaluated out in simulated lung fluid at 37° at pH 7.4. The respirable or fine particle fraction was determined by using twin stage impinger. The stability study of freeze dried as well as aqueous liposomal systems was carried out at 2-8° and at ambient temperature (28±40). The freeze dried liposomes showed better fine particle fraction and drug content over the period of six months at ambient as well as at 2-8° storage condition compared to aqueous dispersion of liposomes.     

米托蒽醌隐形阳离子脂质体的制备及体内外抗肿瘤活性

目的:制备米托蒽醌隐形阳离子脂质体(YM),并研究其制剂学性质及体内外抗肿瘤作用。方法:膜材料中引入2-二油酰基羟丙基-3-N,N,N-三甲铵氯(DOTAP)和聚乙二醇2000-二硬脂酰磷脂乙醇胺(DSPE-PEG₂₀₀₀)分别作为阳性和隐形膜材,采用硫酸铵梯度法制备YM,并以泄漏率为指标,考察其稳定性;高内涵扫描仪考察2 h时其在B16F10细胞中的摄取情况;MTT 法测定其对B16F10细胞的增殖影响;B16F10皮下移植瘤模型研究其体内抗肿瘤作用。结果:YM的粒径和Zeta电位分别为(118.2±4.6)nm和(31.6±4.9)mV;载药量和包封率分别为(6.5±0.2)%和(96.2±1.8)%;血清中48 h累积泄漏率<30%,稳定性良好;YM的细胞摄取量是普通长循环脂质体(CM)的10倍,IC₅₀与CM相比显著降低(P<0.05),体内抑瘤率为58.8%,显著高于CM组的42.7%。结论:YM稳定性良好,具有较好的体内外抗肿瘤活性。     

In vitro and in vivo evaluation of N,N,N-trimethylphytosphingosine-iodide (TMP) in liposomes for the treatment of angiogenesis and metastasis

Phytosphingosine and methyl derivatives are important mediators on cellular processes, and are associated with cell growth and death. The antitumor activity of N,N,N-trimethylphytosphingosine-iodide (TMP) as a novel potent inhibitor of angiogenesis and metastasis was evaluated in B16F10 murine melanoma cells. The results indicated that TMP itself effectively inhibited in vitro cell migration, tube formation, and the expression of angiogenic factors as well as in vivo lung metastasis. However, TMP slightly suppressed in vivo experimental tumor metastasis in its free form and induced side effects including hemolysis and local side effects. Therefore, in an attempt to reduce the toxicity and the undesirable side effects of TMP, a liposomal formulation was prepared and tested for its effectiveness. TMP liposomes retained the effectiveness of TMP in vitro while side effects were reduced, and both in vivo experimental and spontaneous tumor metastasis were significantly suppressed. These results support the conclusion that TMP effectively inhibits in vitro angiogenesis as well as in vivo metastasis, and a liposomal formulation is more efficient delivery system for TMP treatment than solution.     

Octreotide-targeted liposomes loaded with CPT-11 enhanced cytotoxicity for the treatment of medullary thyroid carcinoma

Medullary thyroid carcinoma (MTC) is a rare endocrine tumor that frequently metastasizes, but treatment with irinotecan (CPT-11) is limited because of side effects. MTC is known to overexpress the somatostatin receptor subtype 2 (SSTR2). Octreotide (Oct) is a somatostatin analogue that has a high binding affinity for SSTR and can be used as a tumor-targeting ligand. We prepared Oct-targeted liposomes loaded with CPT-11 using Oct-poly (ethylene glycol) (PEG)-lipid and evaluated Oct-mediated association and cytotoxicity of the liposomes with an MTC cell line TT. The association of higher concentrations of modified Oct-targeted liposomes with TT cells was significantly higher than PEGylated liposomes and was significantly inhibited by empty Oct-targeted liposomes but not by free Oct. With exposure for 96 h, the cytotoxicity of Oct-targeted liposomal CPT-11 (IC50: 1.05 ± 0.47 μM) was higher than free CPT-11 (IC50: 3.76 ± 0.61 μM) or PEGylated liposomal CPT-11 (IC50: 3.05 ± 0.28 μM). In addition, empty Oct-targeted liposomes showed significantly higher cytotoxicity than empty nontargeted liposomes at a concentration where free Oct did not show cytotoxicity, suggesting that Oct as a ligand showed cytotoxicity. Moreover, Oct-targeted liposomal CPT-11 led to significantly higher antitumor activity and prolonged the survival time compared with nontargeted liposomal and free CPT-11 at a one-third dose and lower administration times with free CPT-11. These findings indicated that Oct-targeted liposomes loaded with CPT-11 may offer considerable potential for MTC chemotherapy because cytotoxicity of both CPT-11 and Oct was enhanced by effective cellular uptake via SSTR2.     

Liposomal bleomycin: increased therapeutic activity and decreased pulmonary toxicity in mice

Conventional and sterically stabilized liposomes derived from phosphatidylcholine or the antitumor agents, hexadecylphosphocholine and octadecyl-(1,1-dimethyl-4-piperidino-4-yl)-phosphate, as bilayer forming constituents, containing bleomycin, were developed and tested. Liposomal encapsulation of bleomycin enhanced strongly the antitumor activity against P388 leukemia and the Lewis lung carcinoma. This effect was clearly dependent on the size and lipid composition of the bleomycin-containing liposomes. The therapeutic effects were nearly equal for liposomal and free bleomycin in the B16 melanoma. The partial replacement of phosphatidylcholine by alkylphospholipids and the inclusion of polyethylene glycol modified lipids for sterical stabilization did not further improve the therapeutic efficacy but increased, in some cases, the toxicity of liposomes. Bleomycin-induced lung injury was not observed if liposomal bleomycin was administered.     

Evaluation of pharmacokinetic, biodistribution, pharmacodynamic, and toxicity profile of free juglone and its sterically stabilized liposomes

The present study was aimed to formulate and compare the pharmacokinetic, biodistribution, pharmacodynamic, and toxicity profiles of free 5-hydroxy-1,4-naphthoquinone (juglone) with sterically stabilized liposomal form. The liposomes were optimized for size, zeta potential, entrapment efficiency (EE), and in vitro release properties. The optimized formulation had a mean size, zeta potential, and EE value of 137.1 nm, -43.1 mV, and 67.2%, respectively. In vitro release studies showed biphasic pattern with initial burst followed by sustained release over the study period, releasing about 61% after 24 h. In vitro cytotoxicity studies against melanoma cells indicated that liposomal juglone was more toxic than free juglone. Free juglone had short plasma half-life of about 2 h, whereas liposomal juglone exhibited significantly improved pharmacokinetics with a 12-fold increase in plasma half-life. Further, biodistribution studies indicated rapid renal elimination of free juglone, evidenced by its significant localization in kidneys. Conversely, the accumulation of liposomal juglone in kidneys reduced significantly with enhanced tumor localization, thereby resulting in enhanced antitumor activity. The histological studies revealed lower levels of nephrotoxicity for liposomal juglone compared with that of free juglone. To conclude, sterically stabilized liposomes could be a promising approach for the intravenous delivery of hydrophobic compounds such as juglone.     

Intracellular delivery of doxorubicin with RGD-modified sterically stabilized liposomes for an improved antitumor efficacy: in vitro and in vivo

Passive targeting by sterically stabilized liposomes (SSL), once combined with efficient intracellular delivery, may be a very useful strategy to improve the antitumor efficacy for the anticancer agents. The arginine-glycine-aspartic acid tripeptide (RGD) is known to serve as a recognition motif for several different integrins located on cell surface. In this study, the RGD tripeptide was coupled to the distal end of the poly (ethylene glycol)-coated liposomes (RGD-SSL) aimed to achieve increased tumor accumulation and enhanced intracellular uptake. DOX-loaded RGD-SSL (RGD-SSL-DOX), DOX-loaded SSL (SSL-DOX), and free DOX were compared with respect to their in vitro uptake and cytotoxicity and their in vivo biodistribution and therapeutic efficacy in tumor-bearing mice. Flow cytometry and confocal microscopy studies revealed that RGD-SSL could facilitate the DOX uptake into melanoma cells by integrin-mediated endocytosis. RGD-SSL-DOX displayed higher cytotoxicity on melanoma cells than SSL-DOX. While RGD-SSL-DOX demonstrated prolonged circulation time and increased tumor accumulation as SSL-DOX did, it showed remarkably higher splenic uptake than SSL-DOX. Mice receiving RGD-SSL-DOX (5 mg DOX/kg) showed effective retardation in tumor growth compared with those receiving same dose of SSL-DOX, free DOX solution, or saline. These results suggest that RGD-modified SSL may be a feasible intracellular targeting carrier for efficient delivery of chemotherapeutic agents into tumor cells.     

Development and characteristics of temperature-sensitive liposomes for vinorelbine bitartrate

A novel liposome with temperature-sensitivity for vinorelbine bitartrate (VB) was designed to enhance VB targeted delivery and antitumor effect. Liposomes without drugs were prepared by thin film hydration, and then VB was entrapped into liposomes by pH gradient loading method. The mean particle size of the liposomes was about 100 nm, and the drug entrapment efficiency was more than 90%. Stability data indicated that the liposome was physically and chemically stable for at least 6 months at 4 °C. In vitro drug release study showed that drugs hardly released at 37 °C; while at 42 °C, drugs released quickly. For in vivo experiments, the lung tumor model was established by subcutaneous inoculation of cell suspension on mice, liposomes and free VB were injected i.v. in mice, followed by exposure the tumors to hyperthermia (HT) for 30 min after administration. The ratio of inhibition tumor of temperature-sensitive liposomes group was significantly higher than the normal injection group. Combining temperature-sensitive liposomes with HT enhanced the delivery of VB and, consequently, its antitumor effects. This liposome could potentially produce viable clinical strategies for improved targeting and delivery of VB for treatment of cancer.