Photobiological and ultrastructural studies of nanoparticles of poly(lactic-co-glycolic acid)-containing bacteriochlorophyll-a as a photosensitizer useful for PDT treatment

The interaction of polymeric nanoparticles formulated from the biodegradable polymer poly(DL-lactide-co-glycolide) loaded with bacteriochlorophyll-a was studied in homogeneous solution and in vitro in the presence of a macrophage cell line (P388-D1-ATCC). Photodynamic therapy (PDT) activity after different laser doses also was investigated. Scanning electron microscopy analysis of cell phagocyte nanoparticles showed that after 30 min of incubation most of the nanoparticles are in a clear adhesion process to the cell surface. The majority of nanoparticles became phagocytic after 2 hr of incubation time. After laser irradiation of the dye-containing system a total photodamage by nanoparticle phagocyte cells was observed and the cell survival was quantified by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide test. Our results indicate that polymeric nanoparticles work as an efficient drug delivery system for PDT drugs. This approach can be widely used for many other hydrophobic photosensitizers with higher aggregation tendency in neoplastic cell treatment.     

Preparation and optimization of N-trimethyl-O-carboxymethyl chitosan nanoparticles for delivery of low-molecular-weight heparin

The aim of this study was preparation, optimization and in vitro characterization of nanoparticles composed of 6-[O-carboxymethyl]-[N,N,N-trimethyl] (TMCMC) for oral delivery of low-molecular-weight heparin. The chitosan derivative was synthesized. Nanoparticles were prepared using the polyelectrolyte complexation method. Box–Behnken response surface experimental design methodology was used for optimization of nanoparticles. The morphology of nanoparticles was studied using transmission electron microscopy. In vitro release of enoxaparin from nanoparticles was determined under simulated intestinal fluid. The cytotoxicity of nanoparticles on a Caco-2 cell line was determined, and finally the transport of prepared nanoparticles across Caco-2 cell monolayer was defined. Optimized nanoparticles with proper physico-chemical properties were obtained. The size, zeta potential, poly-dispersity index, entrapment efficiency and loading efficiency of nanoparticles were reported as 235?±?24.3?nm, +18.6?±?2.57?mV, 0.230?±?0.03, 76.4?±?5.43% and 12.6?±?1.37%, respectively. Morphological studies revealed spherical nanoparticles with no sign of aggregation. In vitro release studies demonstrated that 93.6?±?1.17% of enoxaparin released from nanoparticles after 600?min of incubation. MTT cell cytotoxicity studies showed no cytotoxicity at 3?h post-incubation, while the study demonstrated concentration-dependent cytotoxicity after 24?h of exposure. The obtained data had shown that the nanoparticles prepared from trimethylcarboxymethyl chitosan may be considered as a good candidate for oral delivery of enoxaparin.     

Long-pulsed dye laser-mediated photodynamic therapy combined with topical therapy for mild to severe comedonal, inflammatory, or cystic acne

BACKGROUND: Acne patients who fail to respond to conventional treatments have been treated with isotretinoin, an effective treatment coming under strict regulation due to the risk of significant side effects. Photodynamic therapy (PDT) may be a viable alternative treatment for recalcitrant acne of various types and levels of severity. OBJECTIVE: To determine the safety and efficacy of combination PDT with topical 5-aminolevulinic acid (ALA) and activation by long-pulsed, pulsed dye laser (LP PDL, 595 nm) energy with topical therapy in patients with mild to severe acne. METHODS: A prospective, controlled pilot, proof-of-principle study of 19 consecutive patients (aged 16-47 years, Fitzpatrick skin types I-VI) with mild to severe cystic, inflammatory, or comedonal acne of the face was conducted. All patients had failed conventional therapy, including oral antibiotics, topical treatments, hormonal therapy, laser procedures (without ALA), and/or oral isotretinoin. Fifteen patients were treated with ALA PDT and 4 patients served as controls; all were continued on topical medications. Patients undergoing PDT were initially randomized to receive either blue light or laser energy. Because recrudescence occurred in 1 patient while undergoing multiple treatments with ALA and blue light, all subsequent patients were treated with ALA and laser energy. The total number of patients treated with LP PDL-mediated ALA PDT was 14. ALA was applied for a short 45-minute incubation followed by 1 minimally overlapping pass with the LP PDL (595 nm, 7.0-7.5 J/cm2 fluence, 10-ms pulse duration, 10-mm spot size, and dynamic cooling spray of 30 ms with a 30-ms delay). Patients treated with conventional therapy (oral antibiotics, oral contraceptives, and topical medications) or laser energy without ALA PDT served as control groups. Patients were followed monthly for up to 13 months. RESULTS: Complete clearance was achieved in 100% (14 out of 14) patients in the LP PDL PDT-treated group. A mean of 2.9 treatments (range 1-6; 2.0-3.7, 95% CI; n=14) was required to achieve complete clearance for a mean follow-up time of 6.4 months (range 1-13; 3.8-8.9 95% CI; n=14). The patient mean percent lesional clearance rate per treatment was 77% (64%-90%, 95% CI; n=14). Improvement in acne lesions became apparent within 1 to 2 weeks after the first treatment. Clearance in the LP PDL PDT group was superior to control groups. In the LP PDL-only control group (n=2), the patient mean percent lesional clearance rate per treatment was 32% without complete clearance after 3 to 4 treatments. In the oral antibiotics, oral contraceptives, and topicals control group (n=2), the clearance rate per treatment was 20%, the mean clearance rate per month was 4%, and complete clearance was not achieved after 6 to 10 months. In the LP PDL-mediated PDT group, treatments were well-tolerated with minimal erythema lasting 1 to 2 days. No cases of crusting, blistering, purpura, scarring, or dyspigmentation occurred. A reduction in the erythema in erythematous acne scars was observed. CONCLUSION: For teenage to adult patients with recalcitrant comedonal, inflammatory, or cystic acne of various degrees of severity, ALA PDT with activation by LP PDL appears to be a safe and effective treatment with minimal side effects. LP PDL-mediated PDT may serve as an important alternative to isotretinoin. Cosmetically well-accepted, LP PDL PDT combined with topical therapy is the first PDT modality to achieve complete clearance with long-term follow-up as compared to controls.     

Silicalites and Mesoporous Silica Nanoparticles for photodynamic therapy

The synthesis of silicalites and Mesoporous Silica Nanoparticles (MSN), which covalently incorporate original water-soluble photosensitizers for PDT applications is described. PDT was performed on MDA-MB-231 breast cancer cells. All the nanoparticles showed significant cell death after irradiation, which was not correlated with (1)O(2) quantum yield of the nanoparticles. Other parameters are involved and in particular the surface and shape of the nanoparticles which influence the pathway of endocytosis. Functionalization with mannose was necessary to obtain the best results with PDT due to an active endocytosis of mannose-functionalized nanoparticles. The quantity of mannose on the surface should be carefully adjusted as a too high amount of mannose impairs the phototoxicity of the nanoparticles. Fluorescein was also encapsulated in MCM-41 type MSN in order to localize the nanoparticles in the organelles of the cells by confocal microscopy. The MSN were localized in lysosomes after active endocytosis by mannose receptors.     

Preparation,characterization, photocytotoxicity assay of PLGA nanoparticles containing zinc (II) phthalocyanine for photodynamic therapy use

Nanoparticles containing Zinc (II) Phthalocyanine (ZnPc) were prepared by a spontaneous emulsification diffusion method utilizing poly-(D,L lactic-co-glycolic acid) (PLGA), characterized and available in cellular culture. The process yield and encapsulation efficiency were 60% and 80%, respectively. The nanoparticles have a mean diameter of 200?nm, a narrow size distribution with polydispersive index of 0.15, smooth surface and spherical shape. ZnPc loaded nanoparticles maintain their photophysical behaviour after the encapsulation process. Photosensitizer released from nanoparticles was sustained with a burst effect of 10% for 3 days. The photocytotoxicity was evaluated on P388-D1 cells. They were incubated with ZnPc loaded Np by 6?h and exposed to light (675?nm) for 120?s, and light dose of 30?J?cm^?2. After 24?h of incubation, the cellular viability was determined, obtaining 60% of cellular death. All the physical-chemical and photobiological measurements performed allowed one conclude that ZnPc loaded PLGA nanoparticles are a promising drug delivery system for PDT.     

Preparation, characterization, photocytotoxicity assay of PLGA nanoparticles containing zinc (II) phthalocyanine for photodynamic therapy use

Nanoparticles containing Zinc (II) Phthalocyanine (ZnPc) were prepared by a spontaneous emulsification diffusion method utilizing poly-(D,L lactic-co-glycolic acid) (PLGA), characterized and available in cellular culture. The process yield and encapsulation efficiency were 60% and 80%, respectively. The nanoparticles have a mean diameter of 200 nm, a narrow size distribution with polydispersive index of 0.15, smooth surface and spherical shape. ZnPc loaded nanoparticles maintain their photophysical behaviour after the encapsulation process. Photosensitizer released from nanoparticles was sustained with a burst effect of 10% for 3 days. The photocytotoxicity was evaluated on P388-D1 cells. They were incubated with ZnPc loaded Np by 6 h and exposed to light (675 nm) for 120 s, and light dose of 30 J cm-2. After 24 h of incubation, the cellular viability was determined, obtaining 60% of cellular death. All the physical-chemical and photobiological measurements performed allowed one conclude that ZnPc loaded PLGA nanoparticles are a promising drug delivery system for PDT.     

Photosensitiser delivery for photodynamic therapy. Part 2: systemic carrier platforms

Background: The treatment of solid tumours and angiogenic ocular diseases by photodynamic therapy (PDT) requires the injection of a photosensitiser (PS) to destroy target cells through a combination of visible light irradiation and molecular oxygen. There is currently great interest in the development of efficient and specific carrier delivery platforms for systemic PDT. Objective: This article aims to review recent developments in systemic carrier delivery platforms for PDT, with an emphasis on target specificity. Methods: Recent publications, spanning the last five years, concerning delivery carrier platforms for systemic PDT were reviewed, including PS conjugates, dendrimers, micelles, liposomes and nanoparticles. Results/conclusion: PS conjugates and supramolecular delivery platforms can improve PDT selectivity by exploiting cellular and physiological specificities of the targeted tissue. Overexpression of receptors in cancer and angiogenic endothelial cells allows their targeting by affinity-based moieties for the selective uptake of PS conjugates and encapsulating delivery carriers, while the abnormal tumour neovascularisation induces a specific accumulation of heavy weighted PS carriers by enhanced permeability and retention (EPR) effect. In addition, polymeric prodrug delivery platforms triggered by the acidic nature of the tumour environment or the expression of proteases can be designed. Promising results obtained with recent systemic carrier platforms will, in due course, be translated into the clinic for highly efficient and selective PDT protocols.     

5-氨基酮戊酸对人舌麟癌Tea8113细胞株体外光动力杀伤作用的基础研究

目的初步研究体外5-氨基酮戊酸介导的光动力疗法对人舌鳞癌Tea8113细胞的杀伤效应。方法体外培养Tca8113细胞,以5-氨基酮戊酸为光敏剂,半导体激光治疗仪给予光动力疗法,采用MTr法检测光敏剂不同孵育时间、不同光敏剂浓度对Tea8113细胞抑制率的影响。结果5-AIA—PDT作用后,Tea8113细胞生长受到抑制,孵育时间和浓度效应关系显著（P〈0．05）。药物最佳作用浓度为1mmoL／l,最佳孵育时间为6h。结论5-氨基酮戊酸一光动力疗法能有效杀伤Tea8113细胞,光敏剂孵育时间、光敏剂浓度是影响疗效的重要因素。     

Self-Assembled Biodegradable Nanoparticles Developed by Direct Dialysis for the Delivery of Paclitaxel

Purpose The main objective of this study was to obtain self-assembled biodegradable nanoparticles by a direct dialysis method for the delivery of anticancer drug. The in vitro cellular particle uptake and cytotoxicity to C6 glioma cell line were investigated. Methods Self-assembled anticancer drugs—paclitaxel-loaded poly(d,l-lactic-co-glycolic acid) (PLGA) and poly(l-lactic acid) (PLA) nanoparticles—were achieved by direct dialysis. The physical and chemical properties of nanoparticles were characterized by various state-of-the-art techniques. The encapsulation efficiency and in vitro release profile were measured by high-performance liquid chromatography. Particle cellular uptake was studied using confocal microscopy, microplate reader, and flow cytometry. In addition, the cytotoxicity of this drug delivery system was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay on C6 glioma cell line to predict the possible dose response of paclitaxel-loaded PLGA and PLA nanoparticles. Results PLGA and PLA nanoparticles with or without vitamin E tocopherol polyethylene glycol succinate (TPGS) as an additive were obtained, in which the sustained release of paclitaxel of more than 20 days was achieved. The coumarin6-loaded PLGA and PLA nanoparticles could penetrate the C6 glioma cell membrane and be internalized. The cytotoxicity of paclitaxel-loaded nanoparticles seemed to be higher than that of commercial Taxol? after 3 days incubation when paclitaxel concentrations were 10 and 20 μg/ml. Conclusions Direct dialysis could be employed to achieve paclitaxel-loaded PLGA and PLA nanoparticles, which could be internalized by C6 glioma cells and enhance the cytotoxicity of paclitaxel because of its penetration to the cytoplasm and sustained release property.     

Photosensitizer-assembled PEGylated graphene-copper sulfide nanohybrids as a synergistic near-infrared phototherapeutic agent

Objectives: Stimulative nanostructures play a crucial role in developing the smart nanomedicine for high therapeutic efficacy with minimum adverse effects. Herein, a near-infrared (NIR) light-responsive nanohybrids p-nanographene oxide (GO)-copper sulfide (CuS)/indocyanine green (ICG) comprised of GO, CuS nanoparticles and photosensitizer ICG was fabricated to couple the photothermal property of CuS and photodynamic effect of ICG in one system in order to achieve the synergistic phototherapy.

Methods: pGO-CuS/ICG was constructed by self-assembling ICG on pGO-CuS nanostructure. Its physicochemical, photothermal and photodynamic properties were studied by spectroscopic methods. The in vitro cellular uptake, cytotoxicity, the single/combined photothermal therapeutic (PTT) and photodynamic therapeutic (PDT) effects were investigated with biological techniques.

Results: pGO-CuS/ICG exhibited high efficacy of photothermal conversation and singlet oxygen generation under NIR laser excitation. It entered into the target cancer cells probably via passive transmembrane pathway and exerted obvious PTT and PDT effect against the tumor cells upon irradiation with the respective 940 and 808 nm lasers. In particular, the tremendous synergistic efficacy of PDT and PTT had been demonstrated by tuning the NIR laser combined irradiation.

Conclusions: This study promises the future applications of pGO-CuS/ICG as a NIR light activable theranostic nanodrug for deep-seated cancer noninvasive phototherapy.     

Pharmaceutics: Characterization and Trypanocidal Activity of Nifurtimox-containing and Empty Nanoparticles of Polyethylcyanoacrylates

The aim of this study was to evaluate the utility of nanoparticles of polyalkylcyanoacrylate as a targeted delivery system for nifurtimox against Trypanosoma cruzi, responsible for Chagas' disease. Ethylcyanoacrylate nanoparticles were prepared by an emulsion polymerization process and formulations containing different concentrations of nifurtimox, polyethylcyanoacrylates and surfactants were investigated and analysed for size and drug content. The nanoparticles obtained were less than 200 nm in size, as measured by electron microscopy and cytometry. The peak percentage of nifurtimox uptake into the nanoparticles was 33.4% for use of 500 μL polyethylcyanoacrylate, 200 μL surfactant (Tween 20) and 10 mg nifurtimox in 50 mL polymerization medium. The highest release of nifurtimox from the nanoparticles was 65.4% after 6-h incubation at pH 7.4. In-vitro studies using cultures of T. cruzi epimastigotes revealed considerably increased trypanocidal activity compared with a standard solution of nifurtimox. Studies of cell cultures previously infected with metacyclic forms of the parasite showed that only 2-h treatment with solutions of 0.001% of the nanoparticle suspension reduced parasitism by 87–94% both when the nanoparticles were loaded with nifurtimox and when unloaded. Electron-microscopic examination revealed processes of degeneration and lysis, suggesting apoptotic processes, in intracellular amastigotes and free amastigotes treated with the nanoparticles. It was demonstrated that unloaded nanoparticles, by mechanisms not completely elucidated, have trypanocide activity similar to that of a standard solution of nifurtimox. It is concluded that the nanoparticles loaded with nifurtimox constitutes a good carrier of the drug against T. cruzi. The loaded-nanoparticles significantly increase trypanocidal activity.     

Sheddable Coatings for Long-Circulating Nanoparticles

Nanoparticles, such as liposomes, polymeric micelles, lipoplexes and polyplexes are frequently studied as targeted drug carrier systems. The ability of these particles to circulate in the bloodstream for a prolonged period of time is often a prerequisite for successful targeted delivery. To achieve this, hydrophilic ‘stealth’ polymers, such as poly(ethylene glycol) (PEG), are used as coating materials. Such polymers shield the particle surface and thereby reduce opsonization by blood proteins and uptake by macrophages of the mononuclear phagocyte system. Yet, after localizing in the pathological site, nanoparticles should deliver their contents in an efficient manner to achieve a sufficient therapeutic response. The polymer coating, however, may hinder drug release and target cell interaction and can therefore be an obstacle in the realization of the therapeutic response. Attempts have been made to enhance the therapeutic efficacy of sterically stabilized nanoparticles by means of shedding, i.e. a loss of the coating after arrival at the target site. Such an ‘unmasking’ process may facilitate drug release and/or target cell interaction processes. This review presents an overview of the literature regarding different shedding strategies that have been investigated for the preparation of sterically stabilized nanoparticulates. Detach mechanisms and stimuli that have been used are described.     

Sharp pH-sensitive amphiphilic polypeptide macrophotosensitizer for near infrared imaging-guided photodynamic therapy

Tumor environmental sensitive polypeptide integrated photosensitizer is a platform for imaging-guided photodynamic therapy (PDT). However, the photosensitizer leakage during blood circulation, poor accumulation in tumor tissue and inferior quantum yield of singlet oxygen are still challenges. Herein, NHS-active boron-dipyrromethene derivative with bromine substituted NHS-BODIPY-Br₂ was first synthesized, which possessed high singlet oxygen generation efficiency and near infrared (NIR) fluorescence, and then it was conjugated to a sharp pH (6.36) sensitive polypeptide to achieve a macrophotosensitizer for NIR imaging-guided PDT. In vitro study showed that the macrophotosensitizer nanoparticles exhibited good cellular uptake and ability to kill cancer cells. Once accumulating in the tumor tissues, the nanoparticles can be demicellized by tumor acidity to promote cellular uptake, which could enlarge fluorescence signal intensity and enhance in vivo PDT therapeutic effect upon NIR laser irradiation. It provides a strategy to design photosensitizer conjugated tumor acidity sensitive polypeptide for NIR imaging-guided photodynamic therapy.     

Short incubation PDT versus 5-FU in treating actinic keratoses

The efficacy of photodynamic therapy (PDT) using broad area treatment with 5-aminolevulinic acid (ALA) has not been compared to topical 5-fluorouracil (5-FU) in the treatment of actinic keratoses (AK). The purpose of this study was to compare the efficacy and tolerability of PDT using short incubation time, broad area treatment with ALA plus activation with either blue light or laser light to topical 5-FU in the treatment of AK of the face and scalp. Thirty-six subjects with AK of either the face or scalp were randomized to receive either application of ALA for 1 hour followed by activation with blue light or pulsed dye laser or topical 5-FU. Efficacy was evaluated by grading AK lesions and photoaging signs. Tolerability was assessed by scoring crusting/erosions, erythema and stinging/burning. Treatment with PDT using ALA plus blue light was as effective as topical 5-FU in clearing AK. PDT using ALA plus laser light was the least effective treatment. All treatments made improvements in the signs of photoaging. Both PDT treatments were better tolerated than 5-FU. In conclusion, broad area PDT treatment with ALA plus activation with blue light appears to be as effective as 5-FU in the treatment of AK. ALA plus laser light is somewhat less effective than the above therapies. Efficacy could likely be improved with further study of laser parameters and incubation times.     

Photodynamic therapy of experimental B-16 melanoma in mice with tumor-targeted 5,10,15,20-tetraphenylporphin-loaded PEG-PE micelles

Polyethylene glycol (PEG)-diacyl lipid micelles have been prepared by loading with the hydrophobic meso-5,10,15,20-tetraphenyl-21H,23H-porphine (TPP) and used for the photodynamic treatment of B-16 melanoma cells in vitro and in vivo. The use of PEG-PE micelles allowed for a 150-fold increased the solubilization of TPP, compared with the native drug. The average size of the PEG-PE micelles was in the range of 10–12 nm with a narrow size distribution. At 50 μg/ml of TPP in micelles with an irradiation intensity of 4.5–21.5 mW/cm², the viability of B-16 melanoma cells in vitro decreased in a fluence-dependent manner. A highly effective outcome of photodynamic therapy (PDT) with TPP-loaded PEG-PE micelles can be further increased by modifying such micelles with cancer-specific monoclonal antibody 2C5 to TPP-loaded micelles to tumor cells. TPP-containing 2C5-modified micelles provided the strongest phototoxic effect against B-16 cells in vitro compared with TPP-loaded plain micelles at the same TPP concentration. The association of TPP-loaded immuno-targeted micelles with melanoma cells was also studied by flow cytometry. An increase in cell association was found for 2C5-targeted micelles compared with non-targeted micelles. In vivo, the PDT treatment of subcutaneous melanoma-bearing C57BL/6 mice with 100 mW/cm² of 630 nm laser light 9 h after the administration of the micellar TPP (1 mg/kg of TPP) resulted in a significant inhibition of tumor growth. Compared with controls, the weight of postmortem tumors was approx. 3.5- and 7.5-fold smaller with TPP-loaded PEG-PE micelles and TPP-loaded PEG-PE 2C5-immunomicelles, respectively.     

Improved photodynamic activity of porphyrin loaded into nanoparticles: an in vivo evaluation using chick embryos

Hydrophobic porphyrins are potentially interesting molecules for the photodynamic therapy (PDT) of solid cancers or ocular vascularization diseases. Their pharmaceutical development is, however, hampered by their lipophilicity, which renders formulation difficult especially when intravenous administration is needed. Encapsulation of a lipophilic derivative of porphyrin, the meso-tetra(p-hydroxyphenyl)porphyrin (p-THPP), into polymeric biodegradable poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles proved to enhance its photodynamic activity against mammary tumour cells when compared to free drug. In order to further investigate these carriers, the efficacy of the encapsulated drug was assessed on the chick embryo chorioallantoic membrane (CAM) model. First, we identified a suitable solvent for the drug in terms of p-THPP solubility and tolerability by chick embryos. This solution was used as a reference. Then, the fluorescence pharmacokinetics and the photodynamic effects of the porphyrin on CAM vessels were evaluated after intravenous administration of either a p-THPP solution (free drug) or the drug loaded into nanoparticles. The results showed that: (i) the drug remained longer in the vascular compartment when incorporated into nanoparticles and (ii) vascular effects of p-THPP after light irradiation were enhanced with nanoparticle carriers. These results are discussed taking into account the extravasation of intravascular circulating photosensitizers and its influence on PDT performance.     

Formulation and Lyoprotection of Poly(Lactic Acid-Co-Ethylene Oxide) Nanoparticles: Influence on Physical Stability and In Vitro Cell Uptake

Purpose. To investigate the feasibility of producing freeze-dried poly-(ethylene oxide) (PEO)-surface modified nanoparticles and to study their ability to avoid the mononuclear phagocytic system (MPS), as a function of the PEO chain length and surface density. Methods. The nanoparticles were produced by the salting-out method using blends of poly(D,L-lactic acid) (PLA) and poly(D,L-lactic acid-co-ethylene oxide) (PLA-PEO) copolymers. The nanoparticles were purified by cross-flow filtration and freeze-dried as such or with variable amounts of trehalose as a lyoprotectant. The redispersibility of the particles was determined immediately after freeze-drying and after 12 months of storage at –25° C. The uptake of the nanoparticles by human monocytes was studied in vitro by flow cytometry. Results. PLA-PEO nanoparticles could be produced from all the polymeric blends used. Particle aggregation after freeze-drying was shown to be directly related to the presence of PEO. Whereas this problem could be circumvented by use of trehalose, subsequent aggregation was shown to occur during storage. These phenomena were possibly related to the specific thermal behaviours of PEO and trehalose. In cell studies, a clear relationship between the PEO content and the decrease of uptake was demonstrated. Conclusions. The rational design of freeze-dried PEO-surface modified nanoparticles with potential MPS avoidance ability is feasible by using the polymer blends approach combined with appropriate lyoprotection and optimal storage conditions.     

Cooperative Effect of 5-Aminolevulinic Acid and Gold Nanoparticles for Photodynamic Therapy of Cancer

An enhanced capacity for protoporphyrin IX (PpIX) synthesis through 5-aminolevulinic acid (ALA) administration has been reported in cancer cells. We compared the effect of ALA and ALA combined with gold nanoparticles (ALA–AuNPs) for photodynamic therapy (PDT) on human cervical cancer cell line. Because PpIX after photoactivation produces reactive oxygen species (ROS), ALA-AuNPs combinations can enhance this production and then induce higher phototoxicity. With this aim, two different-sized AuNPs (14 and 136 nm, AuNPl and AuNP2, respectively) were successfully synthesized and characterized by UV-visible spectrophotometry and transmission electron microscopy. AuNPs were combined with ALA to evaluate their cooperative action in the intracellular ROS production, cell viability, and cell death mechanism. Results showed that ALA-AuNPs combinations induced cell death via ROSmediated apoptosis after PDT. When exposed to light at their resonance wavelength, AuNP2 combined with ALA result in cytotoxicity and cell injury in greater extension than ALA and ALA-AuNPl combination. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:2660-2669, 2013     

Improved drug targeting of cancer cells by utilizing actively targetable folic acid-conjugated albumin nanospheres

Folic acid-conjugated albumin nanospheres (FA-AN) have been developed to provide an actively targetable drug delivery system for improved drug targeting of cancer cells with reduced side effects. The nanospheres were prepared by conjugating folic acid onto the surface of albumin nanospheres using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as a catalyst. To test the efficacy of these nanospheres as a potential delivery platform, doxorubicin-loaded albumin nanospheres (DOX-AN) and doxorubicin-loaded FA-AN (FA-DOX-AN) were prepared by entrapping DOX (an anthracycline, antibiotic drug widely used in cancer chemotherapy that works by intercalating DNA) into AN and FA-AN nanoparticles. Cell uptake of the DOX was then measured. The results show that FA-AN was incorporated into HeLa cells (tumor cells) only after 2.0 h incubation, whereas HeLa cells failed to incorporate albumin nanospheres without conjugated folic acid after 4.0 h incubation. When HeLa cells were treated with the DOX-AN, FA-DOX-AN nanoparticles or free DOX, cell viability decreased with increasing culture time (i.e. cell death increases with time) over a 70 h period. Cell viability was always the lowest for free DOX followed by FA-DOX-AN4 and then DOX-AN. In a second set of experiments, HeLa cells washed to remove excess DOX after an initial incubation for 2 h were incubated for 70 h. The corresponding cell viability was slightly higher when the cells were treated with FA-DOX-AN or free DOX whilst cells treated with DOX-AN nanoparticles remained viable. The above experiments were repeated for non-cancerous, aortic smooth muscle cells (AoSMC). As expected, cell viability of the HeLa cells (with FA receptor alpha, FRα) and AoSMC cells (without FRα) decreased rapidly with time in the presence of free DOX, but treatment with FA-DOX-AN resulted in selective killing of the tumor cells. These results indicated that FA-AN may be used as a promising actively targetable drug delivery system to improve drug targeting to cancer cells.     

Synchronous delivery of oxygen and photosensitizer for alleviation of hypoxia tumor microenvironment and dramatically enhanced photodynamic therapy

Photosensitizer, proper laser irradiation, and oxygen are essential components for effective photodynamic therapy (PDT) in clinical cancer therapy. However, native hypoxic tumoral microenvironment is a major barrier hindering photodynamic reactions in vivo. Thus, we have prepared biocompatible liposomes by loading complexes of oxygen-carrier (hemoglobin, Hb) and photosensitizer (indocyanine green, ICG) for enhanced PDT against hypoxic tumor. Ideal oxygen donor Hb, which is an oxygen-carried protein in red blood cells, makes such liposome which provide stable oxygen supply. ICG, as a photosensitizer, could transfer energy from lasers to oxygen to generate cytotoxic reactive oxygen species (ROS) for treatment. The liposomes loading ICG and Hb (LIH) exhibited efficient tumor homing upon intravenous injection. As revealed by T₂-weighted magnetic resonance imaging and immunohistochemical analysis, the intratumoral hypoxia was greatly alleviated, and the level of hypoxia inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF) in tumor was obviously down-regulated. A weak PDT efficiency was found in cells incubated in simulated hypoxia condition in vitro, while PDT effect was dramatically enhanced in LIH treated hypoxia cells under near-infrared (NIR) laser, which was mainly attributed to massive generation of ROS with sufficient oxygen supply. ROS trigger oxidative damage of tumors and induce complete suppression of tumor growth and 100% survival rate of mice, which were also in good health condition. Our work highlights a liposome-based nanomedicine that could effectively deliver oxygen to tumor and alleviate tumor hypoxia state, inducing greatly improved efficacy compared to conventional cancer PDT and demonstrates the promise of modulating unfavorable tumor microenvironment with nanotechnology to overcome limitations of cancer therapies.