Liposomes for drug delivery to the lungs by nebulization

Preparation of drug-loaded freeze-dried (FD) liposomes, designed for delivery to lungs after rehydration/nebulization was investigated. Rifampicin (RIF) incorporating multilamelar (MLV) and dried rehydrated vesicles (DRV); composed of phosphatidylcholine (PC), dipalmitoyloglycero-PC (DPPC) or distearoyloglycero-PC (DSPC), containing or not Cholesterol (Chol), were prepared. Vesicles were characterized for encapsulation efficiency (EE%), size distribution, zeta-potential, stability during freeze drying (FD) and nebulization (nebulization efficiency (NE%) and retention of RIF after nebulization (NER%)). Mucoadhesion and toxicity in A549 cells was measured. RIF EE% was not affected by liposome type but lipid composition was important; Synthetic lipid vesicles (DPPC and DSPC) had higher EE% compared to PC. As Chol increased EE% decreased. Freeze drying (FD) had no effect on EE%, however trehalose decreased EE% possibly due to RIF displacement. NER% was highly affected by lipid composition. Results of NE% and NER% for RIF-loaded liposomes show that DSPC/Chol (2:1) is the best composition for RIF delivery in vesicular form to lungs, by nebulization. Mucoadhesion and A549 cell toxicity studies were in line with this conclusion, however if mucoadhesion is required, improvement may be needed.     

Liposomes for drug delivery to the lungs by nebulization.

Preparation of drug-loaded freeze-dried (FD) liposomes, designed for delivery to lungs after rehydration/nebulization was investigated. Rifampicin (RIF) incorporating multilamelar (MLV) and dried rehydrated vesicles (DRV); composed of phosphatidylcholine (PC), dipalmitoyloglycero-PC (DPPC) or distearoyloglycero-PC (DSPC), containing or not Cholesterol (Chol), were prepared. Vesicles were characterized for encapsulation efficiency (EE%), size distribution, zeta-potential, stability during freeze drying (FD) and nebulization (nebulization efficiency (NE%) and retention of RIF after nebulization (NER%)). Mucoadhesion and toxicity in A549 cells was measured. RIF EE% was not affected by liposome type but lipid composition was important; Synthetic lipid vesicles (DPPC and DSPC) had higher EE% compared to PC. As Chol increased EE% decreased. Freeze drying (FD) had no effect on EE%, however trehalose decreased EE% possibly due to RIF displacement. NER% was highly affected by lipid composition. Results of NE% and NER% for RIF-loaded liposomes show that DSPC/Chol (2:1) is the best composition for RIF delivery in vesicular form to lungs, by nebulization. Mucoadhesion and A549 cell toxicity studies were in line with this conclusion, however if mucoadhesion is required, improvement may be needed.     

Antiproliferative effect of extracts from Aristolochia baetica and Origanum compactum on human breast cancer cell line MCF-7

Aristolochia baetica L. (Aristolochiaceae) and Origanum compactum Benth. (Lamiaceae) are native plants of Morocco used in traditional medicine. In order to systematically evaluate their potential activity on human breast cancer, four different polarity extracts from each plant were assessed in vitro for their antiproliferative effect on MCF-7 cells. As a result, several extracts of those plants showed potent cell proliferation inhibition on MCF-7 cells. Chloroform extract of A. baetica (IC50: 216.06?±?15 μg/mL) and ethyl acetate of O. compactum (IC50: 279.51?±?16 μg/mL) were the most active. Thin layer chromatography examination of the bioactive extracts of A. baetica and O. compactum showed the presence of aristolochic acid and betulinic acid, respectively. These results call for further studies of these extracts.     

Effects of ginsenoside compound K combined with cisplatin on the proliferation,apoptosis and epithelial mesenchymal transition in MCF-7 cells of human breast cancer

Context: Breast cancer seriously harms the health of women and there are currently few therapeutic options for patients with breast cancer.

Objective: Effects of ginsenoside compound K (CK) in combination with cisplatin (DDP) on the proliferation, apoptosis, and epithelial mesenchymal transition (EMT) of MCF-7 cells were studied.

Materials and methods: MCF-7 cells were divided into CK (50?μmol/L) group, DDP (10?mg/L) group, CK (50?μmol/L)?+DDP (10?mg/L) group, and control (CON) group. The cells in the CON group were not treated with any drugs. Proliferation, apoptosis, expression of E-cadherin, N-cadherin, vimentin, protein kinase B (Akt), phosphorylated Akt (p-Akt), and level of fibronectin (FN) in MCF-7 cells were detected by methyl thiazolyl tetrazolium (MTT), flow cytometry, western blotting, and enzyme-linked immuno sorbent assay (ELISA), respectively.

Results: The proliferation inhibition rates in CK, DDP, and CK?+?DDP groups at 48?h were 19.18?±?2.25, 21.34?±?2.84, and 43.37?±?5.62, respectively. The apoptosis rates were 2.85?±?0.56, 13.37?±?2.28, 20.04?±?2.92, and 30.78?±?4.64 at 24?h and 3.14?±?0.72, 20.36?±?3.28, 27.58?±?4.09, and 41.62?±?5.83 at 48?h in CON, CK, DDP, and CK?+?DDP groups, respectively. CK or DDP alone and their combination all could reduce the levels of N-cadherin, vimentin, p-Akt/Akt, and FN and elevate level of E-cadherin.

Discussion and conclusion: Both CK and DDP can inhibit the proliferation, EMT, and induce the apoptosis in MCF-7 cells, which may be related to the PI3K/Akt pathway. In addition, the combination of CK with DDP can produce a better effect.     

Preparation of pH-sensitive,long-circulating and EGFR-targeted immunoliposomes

A long-circulating formulation of pH-sensitive liposomes (PSLs) with antibodies against epidermal growth factor receptor (EGFR) attached was designed, expecting an increase in binding and delivery of liposomes to the target cells including non-small cell lung cancer (NSCLC) cells. Physicochemical properties of the PSLs were measured by SEM and DLS. Leakage of a self-quenching fluorescent probe, calcein, from the liposome was studied for the evaluation of pH-sensitivity. Encapsulation efficiency of gemcitabine (an anti-cancer drug) in PSLs was about 67%. Average size of liposomes was 88 nm in diameter. The PSL of DOPE/CHEMS (6:4 molar ratio) formulation showed a dramatic pH-sensitivity at/around pH 5.5, whereas non-PSL of DPPC/Chol or PC/CHEMS formulation did not. Anti-proliferation effect of gemcitabine-encapsulating PSLs & Ab-PSLs in A549 cells was 2-fold higher than the free drug, which was further elucidated by the apoptosis of the cells by gemcitabine (∼10% apoptosis for PSL or Ab-PSL formulation vs. ∼1% for free drug or non-PSL formulation) using FACS analysis. These data demonstrate delivery of gemcitabine to tumor cells can be improved by long-circulating PSLs or Ab-PSLs formulations in vitro.     

A nano-liposome vaccine carrying E75, a HER-2/neu-derived peptide,exhibits significant antitumour activity in mice

E75 (HER-2/neu-369–377), is an immunogenic peptide which is highly expressed in breast cancer patients. The purpose of this study was to develop an effective vaccine delivery/adjuvant system by attachment of this peptide to the surface of liposomes consisting of phospholipids including distearoylphosphocholine (DSPC) and distearoyl phosphoglycerol (DSPG) with high transition temperature (Tm) and dioleoylphosphatidylethanolamine (DOPE) (a pH-sensitive lipid for cytosolic antigen delivery) to improve antitumour immune activity against the E75 peptide. For this purpose, the E75 peptide was incorporated into liposomes consisting of DSPC/DSPG/cholesterol (Chol)/DOPE (15/2/3/5 molar ratio) through conjugation with distearoylphosphoethanolamine-N-[maleimide(polyethylene glycol)-2000] (maleimide-PEG₂₀₀₀-DSPE). Immunization of BALB/c mice was performed three times with different forms of liposomal formulations at 2-week intervals and antitumour immunity responses were evaluated. Results of ELISpot and flow cytometry analysis showed that mice vaccinated with DSPC/DSPG/Chol/DOPE/E75 have significantly enhanced the antigen-specific IFN-γ response of CD8⁺ T cells and generated cytotoxic T lymphocytes (CTL) antitumour responses. CTL responses induced by this formulation resulted in inhibition of tumour progression and longer survival time in the mice TUBO tumour model. The results revealed that the liposomes consist of DSPC/DSPG/Chol/DOPE could be suitable candidates for vaccine delivery of E75 peptide for the prevention and therapy of HER2-positive breast cancer and merit further investigation.     

Comparison of antiproliferative effect of epigallocatechin gallate when loaded into cationic solid lipid nanoparticles against different cell lines

Several therapeutic properties have been attributed to epigallocatechin gallate (EGCG), a phytopharmaceutical polyphenol with antioxidant and antiproliferative activity. EGCG is, however, very prone to oxidation in aqueous solutions which changes its bioactive properties. Its loading in nanoparticles has been proposed to reduce its degradation while increasing its in vivo efficacy. The aim of this study was to compare the antiproliferative effect of EGCG before and after its loading in solid lipid nanoparticles (SLNs), against five different cell lines (Caco-2, HepG2, MCF-7, SV-80 and Y-79). EGCG produced concentration- and time-dependent antiproliferative effect, with efficacy dependent on the cell line. The order of potency was: MCF-7>SV-80>HepG2>Y-79>Caco-2, for 24?h exposure (MCF-7 IC₅₀=58.60?±?3.29?µg/mL; Caco-2 IC₅₀>500.00?µg/mL). To the best of our knowledge this is the first study reporting EGCG antiproliferative effect in SV-80 and Y-79 cells. DDAB-SLN physicochemical properties (size ～134?nm; PI～0.179; ZP ～+28mV) were only slightly modified with EGCG loading (EGCG-DDAB-SLN: ～144?nm; PI～0.160; ZP ～+26mV). EGCG loading in SLN, only slightly increases the EGCG antiproliferative effect in MCF-7 and SV-80 cells. SLN exhibited intrinsic toxicity, attributed to the surfactant used in its production. From the obtained results, the biocompatibility of blank SLN must be also considered when testing the efficacy of loaded phytopharmaceutics.     

Bilayer Composition,Temperature, Speciation Effects and the Role of Bilayer Chain Ordering on Partitioning of Dexamethasone and its 21-Phosphate

Purpose

Models to predict membrane-water partition coefficients (K_p) as a function of drug structure, membrane composition, and solution properties would be useful. This study explores the partitioning of dexamethasone (Dex) and its ionizable 21-phosphate (Dex-P) in liposomes varying in acyl chain length, physical state, and pH.

Methods

DMPC:mPEG DMPE, DPPC:mPEG DPPE, and DSPC:mPEG DSPE (95:5 mol%) liposomes were prepared by thin film hydration. K_p values for Dex and Dex-P were determined from pH 1.5–8 by equilibrium dialysis and equilibrium solubility (Dex).

Results

Dex K_p values at 25°C were 705?±?24, 106?±?11, and 58?±?9 in DMPC, DPPC, and DSPC, increasing to 478?±?20 in DPPC liposomes at 45°C. Both neutral and anionic species contributed to the K_p of Dex-P versus solution pH (1.5–8). A linear correlation was found between the natural logarithm of K_p and the inverse of bilayer free surface area (1/a_free) where a_free is a parameter reflecting chain ordering that depends on bilayer composition and temperature.

Conclusions

Models of the pH dependence of partitioning of ionizable compounds must include contributions of both neutral and ionized species. Bilayer free surface area may be an important variable to predict K_p of drug molecules versus lipid composition and temperature.

     

Preparation and optimisation of anionic liposomes for delivery of small peptides and cDNA to human corneal epithelial cells

Drug delivery to corneal epithelial cells is challenging due to the intrinsic mechanisms that protect the eye. Here, we report a novel liposomal formulation to encapsulate and deliver a short sequence peptide into human corneal epithelial cells (hTCEpi). Using a mixture of Phosphatidylcholine/Caproylamine/Dioleoylphosphatidylethanolamine (PC/CAP/DOPE), we encapsulated a fluorescent peptide, resulting in anionic liposomes with an average size of 138.8?±?34?nm and a charge of ?18.2?±?1.3?mV. After 2?h incubation with the peptide-encapsulated liposomes, 66% of corneal epithelial (hTCEpi) cells internalised the FITC-labelled peptide, demonstrating the ability of this formulation to effectively deliver peptide to hTCEpi cells. Additionally, lipoplexes (liposomes complexed with plasmid DNA) were also able to transfect hTCEpi cells, albeit at a modest level (8% of the cells). Here, we describe this novel anionic liposomal formulation intended to enhance the delivery of small cargo molecules in situ.     

pH-sensitive liposomes--principle and application in cancer therapy

The purpose of this review is to provide an insight into the different aspects of pH-sensitive liposomes. The review consists of 6 parts: the first introduces different types of medications made in liposomal drug delivery to overcome several drawbacks; the second elaborates the development of pH-sensitive liposomes; the third explains diverse mechanisms associated with the endocytosis and the cytosolic delivery of the drugs through pH-sensitive liposomes; the fourth describes the role and importance of pH-sensitive lipid dioleoylphosphatidylethanolamine (DOPE) and research carried on it; the fifth explains successful strategies used so far using the mechanism of pH sensitivity for fusogenic activity; the final part is a compilation of research that has played a significant role in emphasizing the success of pH-sensitive liposomes as an efficient drug delivery system in the treatment of malignant tumours. pH-Sensitive liposomes have been extensively studied in recent years as an amicable alternative to conventional liposomes in effectively targeting and accumulating anti-cancer drugs in tumours. This research suggests that pH-sensitive liposomes are more efficient in delivering anti-cancer drugs than conventional and long-circulating liposomes due to their fusogenic property. Research focused on the clinical and therapeutic side of pH-sensitive liposomes would enable their commercial utility in cancer treatment.     

Development and evaluation of co-formulated docetaxel and curcumin biodegradable nanoparticles for parenteral administration

Context: Technology for development of biodegradable nanoparticles encapsulating combinations for enhanced efficacy.

Objective: To develop docetaxel (DTX) and curcumin (CRM) co-encapsulated biodegradable nanoparticles for parenteral administration with potential for prolonged release and decreased toxicity.

Materials and methods: Modified emulsion solvent-evaporation technique was employed in the preparation of the nanoparticles optimized by the face centered-central composite design (FC-CCD). The uptake potential was studied in MCF-7 cells, while the toxicity was evaluated by in vitro hemolysis test. In vivo pharmacokinetic was evaluated in male Wistar rats.

Results and discussion: Co-encapsulated nanoparticles were developed of 219?nm size, 0.154 PDI, ?13.74?mV zeta potential and 67.02% entrapment efficiency. Efficient uptake was observed by the nanoparticles in MCF-7 cells with decreased toxicity in comparison with the commercial DTX intravenous injection, Taxotere®. The nanoparticles exhibited biphasic release with initial burst release followed by sustained release for 5 days. The nanoparticles displayed a 4.3-fold increase in AUC (391.10?±?32.94 versus 89.77?±?10.58?μg/ml min) in comparison to Taxotere® with a 6.2-fold increase in MRT (24.78?±?2.36 versus 3.58?±?0.21?h).

Conclusion: The nanoparticles exhibited increased uptake, prolonged in vitro and in vivo release, with decreased toxicity thus exhibiting potential for enhanced efficacy.     

Preparation of novel solid lipid microparticles loaded with gentamicin and its evaluation in vitro and in vivo

Objective: To formulate and evaluate solid-reversed-micellar-solution (SRMS)-based solid lipid microparticles (SLMs) for intramuscular administration of gentamicin.

Methods: SRMS formulated with Phospholipon® 90G and Softisan® 154 were used to prepare gentamicin-loaded SLMs. Characterizations based on size and morphology, stability and encapsulation efficiency (EE%) were carried out on the SLMs. In vitro release of gentamicin from the SLMs was performed in phosphate buffer while in vivo release studies were conducted in rats.

Results: Maximum EE% of 90.0, 91.6 and 83.0% were obtained for SLMs formed with SRMS 1:1, 1:2 and 2:1, respectively. Stable, spherical and smooth SLMs of size range 9.80?±?1.46?µm to 33.30?±?6.42?µm were produced. The release of gentamicin in phosphate buffer varied widely with the lipid contents. Moreover, significant (p?<?0.05) amount of gentamicin was released in vivo from the SLMs.

Conclusion: SRMS-based SLMs would likely offer a reliable means of delivering gentamicin intramuscularly.     

Development of a bone targeted thermosensitive liposomal doxorubicin formulation based on a bisphosphonate modified non-ionic surfactant

Bone is among the most common sites of metastasis in cancer patients, so it is an urgent need to develop drug delivery systems targeting tumor bone metastasis with the feature of controlled release. This study aimed to delivery of thermosensitive liposomal doxorubicin to bone for tumor metastasis treatment. First, Brij78 (polyoxyethylene stearyl ether) was conjugated with Pamidronate (Pa). By incorporating Pa-Brij78 to DPPC/Chol liposomes, we developed Pa surface functionalized liposomes. The Pa-Brij78/DPPC/Chol liposomes (PB-liposomes) exhibited a stronger binding affinity to hydroxyapatite (HA), a major component of bone, than Brij78/DPPC/Chol liposomes (B-liposomes). Doxorubicin (Dox) was then encapsulated in PB-liposomes and the results demonstrated complete release of Dox from PB-liposomes or the complex of HA/PB-liposomes within 10?min at 42?°C. Next, human lung cancer A549 cells were treated with the thermosensitive complex of HA/PB-liposomes/Dox to mimic tumor bone metastasis treatment through bone targeted delivery of therapeutic agents. Pre-incubation of HA/PB-liposomes/Dox with mild heat at 42?°C induced subsequent higher cytotoxicity to A549 cells than incubation of the same complex at 37?°C, suggesting more active drug release triggered by heat. In conclusion, we synthesized a novel surfactant Pa-Brij78 and it has the potential to be used for development of a bone targeted thermosensitive liposome formulation for treatment of tumor bone metastasis.     

Preparation of psoralen polymer–lipid hybrid nanoparticles and their reversal of multidrug resistance in MCF-7/ADR cells

Multidrug resistance (MDR) is the leading cause of failure for breast cancer in the clinic. Thus far, polymer–lipid hybrid nanoparticles (PLN) loaded chemotherapeutic agents has been used to overcome MDR in breast cancer. In this study, we prepared psoralen polymer–lipid hybrid nanoparticles (PSO-PLN) to reverse drug resistant MCF-7/ADR cells in vitro and in vivo. PSO-PLN was prepared by the emulsification evaporation-low temperature solidification method. The formulation, water solubility and bioavailability, particle size, zeta potential and entrapment efficiency, and in vitro release experiments were optimized in order to improve the activity of PSO to reverse MDR. Optimal formulation: soybean phospholipids 50?mg, poly(lactic-co-glycolic) acid (PLGA) 15?mg, PSO 3?mg, and Tween-80 1%. The PSO-PLN possessed a round appearance, uniform size, exhibited no adhesion. The average particle size was 93.59?±?2.87?nm, the dispersion co-efficient was 0.249?±?0.06, the zeta potential was 25.47?±?2.84?mV. In vitro analyses revealed that PSO resistance index was 3.2, and PSO-PLN resistance index was 5.6, indicating that PSO-PLN versus MCF-7/ADR reversal effect was significant. Moreover, PSO-PLN is somewhat targeted to the liver, and has an antitumor effect in the xenograft model of drug-resistant MCF-7/ADR cells. In conclusion, PSO-PLN not only reverses MDR but also improves therapeutic efficiency by enhancing sustained release of PSO.     

Cyclodextrin-based nanosponges: effective nanocarrier for Tamoxifen delivery

The purpose of the present study was to develop Tamoxifen loaded β-cyclodextrin nanosponges for oral drug delivery. The three types of Tamoxifen loaded β-cyclodextrin nanosponges were synthesized by varying the molar ratios of β-cyclodextrin to carbonyldiimidazole as a crosslinker viz. 1:2, 1:4 and 1:8. The Tamoxifen nanosponge complex (TNC) with particle size of 400–600?nm was obtained by freeze drying method. Differential scanning calorimetry, Fourier transformed infra-red spectroscopy and X-ray powder diffraction studies confirmed the complexation of Tamoxifen with cyclodextrin nanosponge. AUC and C_max of TNC formulation (1236.4?±?16.12 µg·mL^?1 h, 421.156?±?0.91 µg/mL) after gastric intubation were 1.44 fold and 1.38 fold higher than plain drug (856.079?±?15.18 µg·mL^?1 h, 298.532?±?1.15 µg/mL). Cytotoxic studies on MCF-7 cells showed that TNC formulation was more cytotoxic than plain Tamoxifen after 24 and 48?h of incubation.     

Effects of sialic acid derivative on long circulation time and tumor concentration of liposomes

The effects of a sialic acid derivative, Neu5Aβ-PA, on the blood circulation and tissue distribution of liposomes composed of dipalmitoylphosphatidylcholine (DPPC), cholesterol (Chol) and Neu5Aβ-PA were investigated compared with liposomes composed of DPPC, Chol and monosialoganglioside GM₁, in mice and rats. When liposomes containing Neu5Aβ-PA were intravenously administered into mice, the plasma concentration of liposomes containing Neu5Ac/3-PA was increased, and the liver and spleen uptakes were decreased; there was no significant difference in tissue distribution between liposomes containing Neu5Aβ-PA (DPPC/Chol/Neu5Aβ-PA= 10: 10:3) and those containing GM₁, (DPPC/Chol/GM₁= 10:10:1). On the other hand, the plasma concentration of liposomes containing Neu5Aβ-PA was significantly greater than that of liposomes containing GM₁, at all times determined in rats, and was about 30.7- and 10.3-fold that of liposomes containing GM₁, at 6 and 24 h, respectively. The liver and spleen uptakes of liposomes containing Neu5Aβ-PA at 6 h were significantly reduced compared with those of liposomes containing GM, in rats. The tumor accumulation of liposomes was also examined. The liver/tumor ratio of liposomes containing Neu5Aβ-PA was similar to that of liposomes containing GM₁ in mice and lower than that of liposomes containing GM₁ in rats.     

Colon-targeted celecoxib-loaded Eudragit® S100-coated poly-ϵ-caprolactone microparticles: Preparation,characterization and in vivo evaluation in rats

Context: Celecoxib suffers from low and variable bioavailability following oral administration of solutions or capsules. Recent studies proved that chemoprevention of colorectal cancer is possible with celecoxib.

Objective: This work aimed to tailor colon-targeted celecoxib-loaded microparticles using time-dependant and pH-dependant coats. Estimation of drug pharmacokinetics following oral administration to fasted rats was another goal.

Methods: A 2³ factorial design was adopted to develop poly-?-caprolactone (PCL) celecoxib-loaded microparticles (F1–F8). To minimize drug-percentages released before colon, another coat of Eudragit^® S100 was applied. In vitro characterization of microparticles involved topography, determination of particle size and entrapment efficiency (EE %). Time for 50% drug release (t_50%) and drug-percentages released after 2 hours (Q_2h) and 4 hours (Q_4h) were statistically compared. Estimation of drug pharmacokinetics following oral administration of double-coat microparticles (F10) was studied in rats.

Results: PCL-single-coat microparticles were spherical, discrete with a size range of 60.66?±?4.21–277.20?±?6.10 μm. Direct correlations were observed between surfactant concentration and EE%, Q_2h and Q_4h. The PCL M.wt. and drug: PCL ratio had positive influences on EE% and negative impacts on Q_2h and Q_4h. When compared to the best achieved PCL-single-coat microparticles (F2), the double-coat microparticles (F10) showed satisfactory drug protection; Q_2h and Q_4h were significantly (P?<?0.01) decreased from 31.84?±?1.98% and 54.72?±?2.10% to 15.92?±?1.78% and 26.93?±?2.76%, respectively. When compared to celecoxib powder, F10 microparticles enhanced the bioavailability and extended the duration of drug-plasma concentration in rats.

Conclusion: The developed double-coat microparticles could be considered as a promising celecoxib extended-release colon-targeting system.     

Emodin induces cytotoxic effect in human breast carcinoma MCF-7 cell through modulating the expression of apoptosis-related genes

Abstract

Context: The poor prognostic outcome of breast cancer is largely due to its resistance to cancer therapies. Development of therapeutic agents that can inhibit growth and induce apoptosis in breast cancer cells can help solve the problem. Emodin is an active anthraquinone that has been reported to have diverse biological effects.

Objective: In this study, the anticancer effects of emodin on growth inhibition, apoptosis induction and the expression of apoptosis-related genes in MCF-7 cells were investigated.

Materials and methods: Growth inhibition induced by emodin was investigated by the MTS assay and the colony formation assay; while emodin-induced apoptosis was determined by the COMET assay and DNA fragmentation detection. Emodin (35?μM)-induced alterations in the expression of apoptotic-related genes were detected by using real-time PCR.

Results: Emodin had significant growth inhibitory effects on MCF-7 cells with IC₅₀?=?7.22?µg/ml (～30?μM). It also exerted a concentration-dependant inhibitory effect on the colony-forming ability of MCF-7 cells with IC₅₀?=?7.60?µg/ml (～30?µM). Hallmarks of apoptosis, such as single-strand DNA breakage and DNA fragmentation, were observed in emodin-treated MCF-7 cells. The gene expression of Fas ligand (FASL) was up-regulated (p?<?0.01) but those of MCL1, CCND1 and C-MYC were down-regulated (p?<?0.05) in emodin-treated MCF-7 cells.

Discussion and conclusion: This study indicated that emodin could induce growth inhibition and apoptosis in MCF-7 cells through the modulation of the expression of apoptosis-related genes. The growth inhibitory effects of emodin might involve both the intrinsic and the extrinsic apoptotic pathways and cell cycle arrest.     

Estrogen-anchored pH-sensitive liposomes as nanomodule designed for site-specific delivery of doxorubicin in breast cancer therapy

The present investigation reports the development of nanoengineered estrogen receptor (ER) targeted pH-sensitive liposome for the site-specific intracellular delivery of doxorubicin (DOX) for breast cancer therapy. Estrone, a bioligand, was anchored on the surface of pH-sensitive liposome for drug targeting to ERs. The estrone-anchored pH-sensitive liposomes (ES-pH-sensitive-SL) showed fusogenic potential at acidic pH (5.5). In vitro cytotoxicity studies carried out on ER-positive MCF-7 breast carcinoma cells revealed that ES-pH-sensitive-SL formulation was more cytotoxic than non-pH-sensitive targeted liposomes (ES-SL). The flow cytometry analysis confirmed significant enhanced uptake (p < 0.05) of ES-pH-sensitive-SL by MCF-7 cells. Intracellular delivery and nuclear localization of the DOX was confirmed by fluorescence microscopy. The mechanism for higher cytotoxicity shown by estrone-anchored pH-sensitive liposomal-DOX was elucidated using reactive oxygen species (ROS) determination. The in vivo biodistribution studies and antitumor activities of formulations were evaluated on tumor bearing female Balb/c mice followed by intravenous administration. The ES-pH-sensitive-SL efficiently suppressed the breast tumor growth in comparison to both ES-SL and free DOX. Serum enzyme activities such as LDH and CPK levels were assayed for the evaluation of DOX induced cardiotoxicity. The ES-pH-sensitive-SL accelerated the intracellular trafficking of encapsulated DOX, thus increasing the therapeutic efficacy. The findings support that estrone-anchored pH-sensitive liposomes could be one of the promising nanocarriers for the targeted intracellular delivery of anticancer agents to breast cancer with reduced systemic side effects.