Folate Receptor-positive Cells Research Articles

In vitro and in vivo evaluation of a novel folate-based amphiphilic multifunctional stabilizer for targeting tumors with paclitaxel nanosuspensions

In order to improve the targeted delivery of the drug, a novel folic acid-modified paclitaxel nanosuspensions (FA-PEG-PTX NCs) delivery system was designed in the present study, and its target delivery ability was verified in vitro and in vivo models. Paclitaxel nanosuspensions were prepared by antisolvent precipitation using folic acid-modified polyethylene glycol monostearate (FA-PEG-MA) and D - α-tocopherol polyethylene glycol succinate (TPGS) as functional stabilizers. The FA-PEG-PTX NCs had an average particle size of 195.33 ± 13.31 nm, a PDI of 0.182 ± 0.022 and a zeta potential of - 12.4 ± 0.62 mV. The drug loading (DL) and encapsulation efficiency (EE) of FA-PEG-PTX NCs were 48.45 ± 3.34 % and 96.90 ± 1.81 %. Particle morphology showed that FA-PEG-PTX NCs had a rod-shaped structure with a smooth surface. XRD and DSC indicated that PTX existed mainly in an amorphous structure in the nanocrystals. FTIR showed no chemical reaction between PTX and stabilizer but rather physical interactions. In vitro dissolution experiments showed that nanocrystals significantly improved the PTX dissolution rate. Cytotoxicity assay and apoptosis assay showed that FA-PEG-PTX NCs had a stronger inhibitory effect on the proliferation and apoptosis of folate receptor-positive cells, Bel-7402 cells, compared with PEG-PTX NCs. In vivo studies showed that FA-PEG-PTX NCs and PEG-PTX NCs had long-lasting and targeted effects. In conclusion, FA-PEG-MA seems to be a promising stabilizer with targeting properties.

Nanodroplets Engineered with Folate Carbon Dots for Enhanced Cancer Cell Uptake toward Theranostic Application.

The research in nanotherapeutics is rapidly advancing, particularly in the realm of nanoconstructs for drug delivery. This study introduces folate-based carbon dot-decorated nanodroplets (f-Dnm), synthesized from a binary mixture of negatively charged folic acid carbon dots (f-CDs) and cationic-branched polyethylenimine (PEI). The uniformly spherical nanodroplets with an average diameter of 115 ± 15 nm exhibit notable photoluminescence. Surface potential analysis reveals a significant change upon coacervation, attributed to strong electrostatic interactions between f-CD and PEI. The engineered nanodroplets show excellent colloidal and photostability even after 6 months of storage at room temperature. The pH-dependent self-assembly and disassembly properties of f-Dnm are explored for drug loading and release studies using doxorubicin (DOX) as a model anticancer drug. Moreover, the f-Dnm nanocarrier demonstrates significantly higher drug loading capabilities (∼90%). In vitro release studies of doxorubicin-loaded f-Dnm [f-Dnm(DOX)] reveal 5 times higher drug release at lysosomal pH 5.4 compared to that at physiological blood pH 7.4. Cytocompatibility assessments using the MTT assay on HeLa, A549, and NIH-3T3 cells confirm the nontoxic nature of f-Dnm, even at high concentrations. Additionally, f-Dnm(DOX) exhibits higher cytotoxicity in HeLa cells compared to f-CD(DOX) at similar DOX concentrations. Cellular uptake studies show an increased uptake of f-Dnm in folate receptor-positive HeLa and MDA-MB 231 cells. Hemolysis assay validated the biocompatibility of the developed formulation. Overall, these engineered nanodroplets represent a class of nontoxic nanocarriers that offer promising potential as nanotherapeutics for folate receptor-positive cells.

Luciferase-Decorated Gold Nanorods as Dual-Modal Contrast Agents for Tumor-Targeted High-Performance Bioluminescence/Photoacoustic Imaging.

Gold nanorods (AuNRs) have been considered highly compelling materials for early cancer diagnosis and have aroused a burgeoning fascination among the biomedical sectors. By leveraging the versatile tunable optical properties of AuNRs, herein, we have developed a novel tumor-targeted dual-modal nanoprobe (FFA) that exhibits excellent bioluminescence and photoacoustic imaging performance for early tumor diagnosis. FFA has been synthesized by anchoring the recombinant bioluminescent firefly luciferase protein (Fluc) on the folate-conjugated AuNRs via the PEG linker. TEM images and UV-vis studies confirm the nanorod morphology and successful conjugation of the biomolecules to AuNRs. The nanoprobe FFA relies on the ability of the folate module to target the folate receptor-positive tumor cells actively, and simultaneously, the Fluc module facilitates excellent bioluminescent properties in physiological conditions. The success of chemical engineering in the present study enables stronger bioluminescent signals in the folate receptor-positive cells (Skov3, Hela, and MCF-7) than in folate receptor-negative cells (A549, 293T, MCF-10A, and HepG2). Additionally, the AuNRs induced strong photoacoustic conversion performance, enhancing the resolution of tumor imaging. No apparent toxicity was detected at the cellular and mouse tissue levels, manifesting the biocompatibility nature of the nanoprobe. Prompted by the positive merits of FFA, the in vivo animal studies were performed, and a notable enhancement was observed in the bioluminescent/photoacoustic intensity of the nanoprobe in the tumor region compared to that in the folate-blocking region. Therefore, this synergistic dual-modal bioluminescent and photoacoustic imaging platform holds great potential as a tumor-targeted contrast agent for early tumor diagnosis with high-performance imaging information.

Selective Protein of Interest Degradation through the Split-and-Mix Liposome Proteolysis Targeting Chimera Approach.

Proteolysis Targeting Chimera (PROTAC) technology represents a promising new approach for target protein degradation using a cellular ubiquitin-proteasome system. Recently, we developed a split-and-mix nanoplatform based on peptide self-assembly, which could serve as a self-adjustable platform for multifunctional applications. However, the lower drug efficacy limits further biomedical applications of peptide-based SM-PROTAC. In this study, we develop a novel split-and-mix PROTAC system based on liposome self-assembly (LipoSM-PROTAC), concurrent with modification of FA (folate) to enhance its tumor-targeting capabilities. Estrogen receptors (ERα) were chosen as the protein of interest (POI) to validate the efficacy of Lipo degraders. Results demonstrate that this PROTAC can be efficiently and selectively taken up into the cells by FA receptor-positive cells (FR+) and degrade the POI with significantly reduced concentration. Compared to the peptide-based SM-PROTACs, our designed LipoSM-PROTAC system could achieve therapeutic efficacy with a lower concentration and provide opportunities for clinical translational potential. Overall, the LipoSM-based platform shows a higher drug efficacy, which offers promising potential applications for PROTAC and other biomolecule regulations.

Stimulus-responsive drug/gene delivery system based on polyethylenimine cyclodextrin nanoparticles for potential cancer therapy

Combination therapy through simultaneous delivery of anti-cancer drugs and genes with nano-assembled structure has been proved to be a simple and effective approach for treating breast cancer. In this study, redox-sensitive folate-appended-polyethylenimine-β-cyclodextrin (roFPC) host-guest supramolecular nanoparticles (HGSNPs) were developed as a targeted co-delivery system of doxorubicin (Dox) and Human telomerase reverse transcriptase-small interfering RNA) hTERT siRNA) for potential cancer therapy. The nanotherapeutic system was prepared by loading adamantane-conjugated doxorubicin (Ad-Dox) into roFPC through the supramolecular assembly, followed by electrostatically-driven self-assembly between hTERT siRNA and roFPC/Ad-Dox. The roFPC’ host-guest structures allow pH-dependent intracellular drug release in a sustained manner, as well as simultaneous and effective gene transfection. This co-delivery vector displayed combined anti-tumor properties of the Dox-enhanced gene transfection, good water-solubility, and biocompatibility, possesses considerably enhanced hemocompatibility, and especially targets folate receptor-positive cells only at low N/P levels to prompt effective cell apoptosis for cancer treatment.

Folic acid-appended galactoxyloglucan-capped iron oxide nanoparticles as a biocompatible nanotheranostic agent for tumor-targeted delivery of doxorubicin

Iron oxide nanoparticles (IONPs) are employed as MRI contrast agents and as effective drug delivery vehicles. However, the limited solubility and biodegradability of these nanoparticles need to be improved for safer biomedical applications. In an attempt to improve the bottlenecks associated with IONPs, the current study focuses on the synthesis of folic acid conjugated, galactoxyloglucan-iron oxide nanoparticles (FAPIONPs), for the loading and controlled release of the encapsulated chemotherapeutic agent doxorubicin (DOX). The as-designed DOX@FAPIONPs induced a dose-dependent increase in cytotoxicity in folate receptor-positive cells through a caspase-mediated programmed cell death pathway while bare DOX demonstrated a non-targeted toxicity profile. Using LC-MS/MS analysis, several major biological processes altered in treated cells, from which, cell cycle, cellular function and maintenance were the most affected. Detailed toxicity studies in healthy mice indicated the absence of any major side effects while bare drugs created substantial organ pathology. Gadolinium-based contrast agents have a risk of adverse effects, including nephrogenic systemic fibrosis overcome by the administration of DOX@FAPIONPs in xenograft mice model. Tumor-targeted biodistribution pattern with a favorable DOX pharmacokinetics will be the driving factor behind the appealing tumor reduction capacity and increased survival benefits demonstrated on solid tumor-bearing mice.

Synthesis of folic acid functionalized gold nanoclusters for targeting folate receptor-positive cells

We report on the synthesis of water-soluble gold nanoclusters capped with polyethylene glycol (PEG)-based ligands and further functionalized with folic acid for specific cellular uptake. The dihydrolipoic acid-PEG-based ligands terminated with –OMe, –NH2 and –COOH functional groups are produced and used for surface passivation of Au nanoclusters (NCs) with diameters <2 nm. The produced sub 2 nm Au NCs possess long-shelf life and are stable in physiologically relevant environments (temperature and pH), are paramagnetic and biocompatible. The paramagnetism of Au NCs in solution is also reported. The functional groups on the capping ligands are used for direct conjugation of targeting molecules onto Au NCs without the need for post synthesis modification. Folic acid (FA) is attached via an amide group and effectively target cells expressing the folate receptor. The combination of targeting ability, biocompatibility and paramagnetism in FA-functionalized Au NCs is of relevance for their exploitation in nanomedicine for targeted imaging.

Folate-Conjugated and Dual Stimuli-Responsive Mixed Micelles Loading Indocyanine Green for Photothermal and Photodynamic Therapy.

A folic acid targeted mixed micelle system based on co-assembly of poly(ε-caprolactone)-b-poly(methoxytri(ethylene glycol) methacrylate-co-N-(2-methacrylamido)ethyl folatic amide) and poly(ε-caprolactone)-b-poly(diethylene glycol monomethyl ether methacrylate) is developed to encapsulate indocyanine green (ICG) for photothermal therapy and photodynamic therapy. In this study, the use of folic acid is not only for specific cancer cell recognition, but also in virtue of the carboxylic acid on folic acid to regulate the pH-dependent thermal phase transition of polymeric micelles for controlled drug release. The prepared ICG-loaded mixed micelles possess several superior properties such as a preferable thermoresponsive behavior, excellent storage stability, and good local hyperthermia and reactive oxygen species generation under near-infrared (NIR) irradiation. The photototoxicity induced by the ICG-loaded micelles has efficiently suppressed the growth of HeLa cells (folate receptor positive cells) under NIR irradiation compared to that of HT-29, which has low folate receptor expression. Hence, this new type of mixed micelles with excellent features could be a promising delivery system for controlled drug release, effective cancer cell targeting, and photoactivated therapy.

Selective liposome targeting of folate receptor positive immune cells in inflammatory diseases

Activated macrophages play a key role in the development and maintenance of inflammatory diseases such as atherosclerosis, lupus, psoriasis, rheumatoid arthritis, ulcerative colitis, and many others. These activated macrophages, but not resting or quiescent macrophages highly up-regulate folate receptor beta (FR-β). This differential expression of FR-β provides a mechanism to selectively deliver imaging and therapeutic agents utilizing folate as a targeting molecule. In an effort to determine whether inflammatory diseases can be targeted utilizing a folate-linked nanosize carrier, a PEG-coated liposome was prepared that incorporated a folate conjugated PEG that also could transport imaging or therapeutic cargo. We demonstrate that these folate-liposomes specifically bind to folate receptor positive cells and accumulate at sites of inflammation in mouse models of colitis and atherosclerosis. These two animal models show that folate-targeted liposomes could be successfully utilized to deliver fluorescent molecules and an anti-inflammatory drug (betamethasone) for diagnostic and therapeutic applications.

Cellular imaging and folate receptor targeting delivery of gum kondagogu capped gold nanoparticles in cancer cells

In this study, the green synthesis of gum kondagogu capped gold nanoparticles (GK-GNPs) was prepared using a naturally available polysaccharide. The anionic gum capped GK-GNPs enabled the successful coupling of folic acid (FA) and fluorescein isothiocyanate (FITC) to produce a fluorescently labelled GNP (F2-GNP). F2-GNPs were further characterized using different physicochemical methods Cellular viability, cellular imaging, and targeted delivery of F2-GNPs were further evaluated in both folate receptor positive (MCF-7) and folate receptor negative (A549) cancer cells. Physicochemical characterization revealed a nanoparticle with a small size (37 nm), smooth surface (surface charge of −23.7 mV), crystallinity of gold nanoparticles and existence of gum kondagogu in the F2-GNPs. Cellular uptake of F2-GNPs indicated a greater affinity towards folate receptor positive cells. This study shows that the F2-GNPs is as an effective nanocarrier for targeted drug delivery and cellular imaging via folate receptors.

Assessment of folate receptor alpha and beta expression in selection of lung and pancreatic cancer patients for receptor targeted therapies.

A number of folate receptor (FR) targeted small molecular drugs and monoclonal antibodies have been introduced into clinical trials to treat FR positive cancers. Because the therapeutic efficacy of these drugs depends prominently on the level of FR-α expression on the cancer cells, patients have been commonly selected for FR-targeted therapies based on the intensity of a folate-targeted radioimaging agent. Unfortunately, uptake of such imaging agents can be mediated by both major isoforms of the folate receptor, FR-α and FR-β. Logically, if the FR positive cell population in a tumor mass is dominated by FR-β positive macrophages, patients could be selected for therapy that have few FR-expressing cancer cells. Although several IHC studies have examined expression of either FR-α or FR-β, no study to date has investigated expression of both FR-α and FR-β in the same tumor mass. Herein, we utilize monoclonal antibodies specific for FR-α (mAb343) and FR-β (m909) to query each isoform's expression in a range of cancers. We show that lung and pancreatic adenocarcinomas express the full spectrum of FR-α and FR-β combinations with ~76% of lung adenocarcinomas expressing both FR-α and FR-β while pancreatic cancers express primarily FR-β. Thus, while folate-targeted imaging of lung cancer patients might accurately reflect the expression of FR-α on lung cancer cells, imaging of pancreatic cancer patients could mislead a physician into treating a nonresponding patient. Overall, these data suggest that an independent analysis of both FR-α and FR-β should be obtained to predict the potential efficacy of a folate-targeted drug.

Self-Assembled Nanospheres of Folate-Decorated Zein for the Targeted Delivery of 10-Hydroxycamptothecin

Site-specific drug delivery is an effective approach to decreasing drug toxicity and enhancing therapeutic effects. In this study, zein was decorated with folic acid (FA) for targeted delivery of 10-hydroxycamptothecin (HCPT). The reaction process was monitored using an online ATR-UV spectrophotometer, and the conjugation degree was quantified using a UV–vis spectrophotometer. Successful conjugation was evidenced using FT-IR and 1H NMR. The influence of FA conjugation on the self-assembly of zein molecules and cellular uptake was investigated in detail. FA conjugation facilitates the formation of small nanoparticles with good dispersity and stability and improves the cellular uptake in folate receptor positive cells. HCPT nanocrystals (NC) were prepared and incorporated into FA-zein. The drug release behavior of HCPT NC/FA-zein nanoparticles is more consistent with a diffusion mechanism than a matrix swelling/erosion mechanism of HCPT NC/zein particles; and HCPT NC/FA-zein nanoparticles induce higher anti...

Triblock near-infrared fluorescent polymer semiconductor nanoparticles for targeted imaging

Self-assembled nanoparticles of triblock copolymers incorporating a NIR-emitting fluorophore and folic acid specifically label folate receptor-positive cells.

Novel amphiphilic folic acid-cholesterol-chitosan micelles for paclitaxel delivery

In order to decrease the toxicity of paclitaxel (PTX) and increase the efficiency, we developed an amphiphilic PTX injection system using a biodegradable and biocompatible polymer synthesized by folic acid, cholesterol, and chitosan (FACC). This FACC-based polymer had a low critical concentration (64.13μg/ml) and could self-assemble in aqueous condition to form nanoscale micelles. The particle sizes of FACC-PTX micelles were 253.2±0.56 nm, the encapsulation efficiency and loading capacity of these FACC-PTX micelles were 65.1±0.23% and 9.1±0.16%, respectively. The cumulative release rate was about 85% at pH 5.0 which was higher than that at pH 7.4 (76%). This pH-dependent release behavior was highly suggesting that PTX release from FACC-PTX micelles might be higher in a weak acidic tumor microenvironment and lower toxic for normal cells. The anti-cancer effectiveness of FACC-PTX micelles was investigated by in vitro cytotoxicity and targeting study. The results revealed that FACC micelles have non-toxic on cells as evidenced by high cell viability found (86% to 100%) in the cells cultured with various concentrations of FACC micelles (1 to 500 μg/ml). Targeting study indicated that the cytotoxic efficacy of FACC-PTX micelles was significantly higher than that with Taxol® in the Hela cells (folate receptor-positive cells). These findings indicated that the anticancer efficiency of PTX can be enhanced by adding some cancer cell positive receptor into drug carrier and the FACC micelle was a potential tumor targeting carrier for PXT delivery.

Abstract 2247: A multicenter clinical trial of lung cancer circulating tumor cell assay with the largest sample size (1210 cases) in China

Abstract Background: As a novel biomarker of primary tumors, circulating tumor cells (CTCs) are well known to play an important role in cancer diagnosis, recurrence and metastasis, and disease monitoring. This study investigated the application of CTCs in lung cancer therapy by quantitative determination of folate receptor-positive CTCs. Methods: This study enrolled 1210 subjects (including 560 lung cancer patients, 350 patients with benign lung diseases, 150 healthy subjects and 150 non-lung cancer malignant tumor patients) and quantified the tiny amounts of CTCs in peripheral blood by negative enrichment using immunomagnetic beads in combination with folate receptor-directed polymerase chain reaction (PCR). Results: Following ROC curve analyses of CTC levels in the patients with benign lung diseases or benign lung diseases and the healthy subjects, the cut-off of CTCs was 8.70 CTC Units/3mL with a specificity of 88.2% (441/500) and a sensitivity of 79.6% (446/560) as determined by the method mentioned above. The ROC area under curve of more than 0.8 (AUC = 0.8797, P&amp;lt;0.0001) suggested the excellence of ligand-directed PCR to diagnose lung cancer. For stage II-IV lung cancer patients, the sensitivities were 70.8%(34/48), 79.6% (125/157), and 90.2% (185/205) respectively, and the sensitivity in stage I patients can reach up to 68.0% (102/150) as well. CTC levels appeared to have greater accuracy to diagnose lung cancer (AUC: 0.8833, P&amp;lt;0.0001) than the combination of the other four lung cancer-specific serum tumor markers (CEA, NSE, CYFRA21-1 and SCC)(AUC: 0.8557, P&amp;lt;0.0001). CTCs were related to tumor stage and tumor size other than pathological type. By quantitative determination of folate receptor-positive cells in 150 different types of cancers, the CTC level in lung cancer was found to be higher than those in gastric, breast, colorectal, liver, and esophageal cancers. Conclusions: LT-PCR achieves quantitative determination of folate receptor-positive CTCs to diagnose lung cancer with high sensitivity and specificity, and significantly greater diagnosis accuracy than the model of combined lung cancer-specific serum tumor markers. Citation Format: Jiatao Lou, Caicun Zhou, Jing Wu, Lihua Qiao, Xiaohui Liang, Xiaoqian Wang, Xiaoxia Chen, Xuefei Li, Chao Zhao. A multicenter clinical trial of lung cancer circulating tumor cell assay with the largest sample size (1210 cases) in China. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2247.

DNA as Tunable Adaptor for siRNA Polyplex Stabilization and Functionalization.

siRNA and microRNA are promising therapeutic agents, which are engaged in a natural mechanism called RNA interference that modulates gene expression posttranscriptionally. For intracellular delivery of such nucleic acid triggers, we use sequence-defined cationic polymers manufactured through solid phase chemistry. They consist of an oligoethanamino amide core for siRNA complexation and optional domains for nanoparticle shielding and cell targeting. Due to the small size of siRNA, electrostatic complexes with polycations are less stable, and consequently intracellular delivery is less efficient. Here we use DNA oligomers as adaptors to increase size and charge of cargo siRNA, resulting in increased polyplex stability, which in turn boosts transfection efficiency. Extending a single siRNA with a 181-nucleotide DNA adaptor is sufficient to provide maximum gene silencing aided by cationic polymers. Interestingly, this simple strategy was far more effective than merging defined numbers (4–10) of siRNA units into one DNA scaffolded construct. For DNA attachment, the 3′ end of the siRNA passenger strand was beneficial over the 5′ end. The impact of the attachment site however was resolved by introducing bioreducible disulfides at the connection point. We also show that DNA adaptors provide the opportunity to readily link additional functional domains to siRNA. Exemplified by the covalent conjugation of the endosomolytic influenza peptide INF-7 to siRNA via a DNA backbone strand and complexing this construct with a targeting polymer, we could form a highly functional polyethylene glycol–shielded polyplex to downregulate a luciferase gene in folate receptor–positive cells.

Design, preparation, and in vitro characterization of a trimodally-targeted nanomagnetic onco-theranostic system for cancer diagnosis and therapy

In this study, the aim was to introduce and characterize a new trimodally-targeted nanomagnetic onco-theranostic system for simultaneous early diagnosis and efficient treatment of cancer. The onco-theranostic system was designed as it could target the tumor site through three targeting approach, i.e. magnetic, folic acid receptor, and pH sensitivity, and concurrently, due to the presence of superparamagnetic iron oxide nanoparticles (SPIONs) with super paramagnetic characteristics could be useful as MRI contrast agent for early cancer diagnosis. To achieve this goal, SPIONs were coated with chitosan and folic acid-conjugated chitosan via ionic gelation method in order to obtain non-targeted nanomagnetic onco-diagnostic (NT/NOD) and targeted nanomagnetic onco-diagnostic (T/NOD) systems. Finally, doxorubicin was loaded successfully into NT/NOD and T/NOD in order to obtain nanomagnetic onco-theranostic (NT/NOT) and targeted nanomagnetic onco-theranostic (T/NOT) systems. The entrapment efficiency and drug loading of T/NOT was determined to be 62.33±5.20% and 10.26±1.36%, respectively. MTT assay revealed that all systems were biocompatible within the concentration range investigated. Also, the T/NOT system showed the lowest IC50 comparing with free doxorubicin and NT/NOT system. In addition, uptake studies and competitive inhibition study verified the folate receptor mediated endocytosis of targeted system by MCF-7 as a folate receptor-positive cell line. The finding revealed that the extent of drug release from theranostic systems was pH-sensitive as it was higher at acidic media compared to that of in the neutral condition. Finally, T2-weighted phantom images, with an acceptable and dose-dependent resolution, proved the potential of T/NOT system as promising T2 MR contrast agent for diagnostic purpose. These finding proved that the prepared T/NOT system have great potential as a novel tumor-targeting nanotheranostic agent for simultaneous MRI imaging and treatment of folate receptor-positive cancers. Further studies are needed to test their behavior in vivo.

Folic-Acid-Modified Conducting Polymer: Electrochemical Detection of the Cell Attachment.

Here, postfunctionalization and bioapplication of a π-conjugated polymer named 4-[4H-dithieno(3,2-b:2',3'-d)pyrrol-4-yl]aniline (DTP-aryl-NH2 ) are reported, which is successfully synthesized via electropolymerization onto the glassy carbon electrode. Folic acid (FA) is used to modify the amino functional polymer via N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride/N-hydroxysuccinimide chemistry for the further steps. The selective adhesion of folate receptor positive cells on the surface is followed by the electrochemical methods. Cyclic voltammetry and electrochemical impedance spectroscopy have been used to characterize stepwise modification of the electroactive surface. After optimization studies such as scan rate during the polymer deposition, FA amount for the efficient surface targeting, incubation time with the cells etc., analytical characterization is carried out. The surface morphologies at each step are imaged by using fluorescence microscopy.

Size controlled protein nanoemulsions for active targeting of folate receptor positive cells

Bovine serum albumin (BSA) nanoemulsions were produced by high pressure homogenization with a tri-block copolymer (Poloxamer 407), which presents a central hydrophobic chain of polyoxypropylene (PPO) and two identical lateral hydrophilic chains of polyethylene glycol (PEG). We observed a linear correlation between tri-block copolymer concentration and size – the use of 5mg/mL of Poloxamer 407 yields nanoemulsions smaller than 100nm. Molecular dynamics and fluorescent tagging of the tri-block copolymer highlight their mechanistic role on the size of emulsions. This novel method enables the fabrication of highly stable albumin emulsions in the nano-size range, highly desirable for controlled drug delivery. Folic Acid (FA)-tagged protein nanoemulsions were shown to promote specific folate receptor (FR)-mediated targeting in FR positive cells. The novel strategy presented here enables the construction of size controlled, functionalized protein-based nanoemulsions with excellent characteristics for active targeting in cancer therapy.

Apoptosis of HeLa cells induced by a new targeting photosensitizer-based PDT via a mitochondrial pathway and ER stress.

Photodynamic therapy (PDT) is emerging as a viable treatment for many cancers. To decrease the cutaneous photosensitivity induced by PDT, many attempts have been made to search for a targeting photosensitizer; however, few reports describe the molecular mechanism of PDT mediated by this type of targeting photosensitizer. The present study aimed to investigate the molecular mechanism of PDT induced by a new targeting photosensitizer (PS I), reported previously by us, on HeLa cells. Apoptosis is the primary mode of HeLa cell death in our system, and apoptosis occurs in a manner dependent on concentration, irradiation dose, and drug–light intervals. After endocytosis mediated by the folate receptor, PS I was primarily localized to the mitochondria and the endoplasmic reticulum (ER) of HeLa cells. PS I PDT resulted in rapid increases in intracellular reactive oxygen species (ROS) production and Ca2+ concentration, both of which reached a peak nearly simultaneously at 15 minutes, followed by the loss of mitochondrial membrane potential at 30 minutes, release of cytochrome c from mitochondria into the cytoplasm, downregulation of Bcl-2 expression, and upregulation of Bax expression. Meanwhile, activation of caspase-3, -9, and -12, as well as induction of C/EBP homologous protein (CHOP) and glucose-regulated protein (GRP78), in HeLa cells after PS I PDT was also detected. These results suggest that apoptosis of HeLa cells induced by PS I PDT is not only triggered by ROS but is also regulated by Ca2+ overload. Mitochondria and the ER serve as the subcellular targets of PS I PDT, the effective activation of which is responsible for PS I PDT-induced apoptosis in HeLa cells.