Theranostic Liposomes Research Articles

Development and In vitro and In vivo efficacy investigation of multifunctional, targeted, theranostic liposomes for imaging and photodynamic therapy of glioblastoma

The challenge of effectively diagnosing and treating glioblastoma, which has a high incidence and low survival rate, remains unresolved. Consequently, research efforts have increased to develop specific and effective diagnostic and therapeutic agents. In this study, N-acetyl glucosamine-modified, IR780-encapsulated, PEG-coated, nanosized, 68Ga-labeled, neutral, and positively charged theranostic liposomes were developed for GLUT1 targeting for PET/CT and NIR imaging and PDT/PTT of GBM. Characterization, in vitro release profile, stability, radiolabeling efficiency, and labeling stability were evaluated. In vitro cell uptake studies were conducted in RG2 and U87 GBM cell lines, NIR images were obtained in the RG2 and U87 cell lines, and fluorescence microscope images were obtained in the RG2 cell line. PDT/PTT activities were assessed after exposure to NIR light (808 nm, 0.8–1 W/cm2). Cytotoxicity was studied in RG2, U87, and L929 cell lines. In vivo studies were performed in GBM model nude mice. The liposomes had an optimum mean particle size (181–193 nm), high encapsulation efficiency (79–84 %), and zeta potential (−5 to −7 mV). The maximum 68Ga labeling efficiency was achieved with an incubation at 80 °C, pH: 3.5, and 5 min. Liposomes remained stable for 30 days at 4 °C. While the intracellular uptake of targeted formulations was higher than non-targeted ones, it was similar for positively charged and neutral formulations. In U87 and RG2 cell lines, cell viability remained above 50 % for liposomal formulations at a concentration of 1.5 μg/mL exposed to NIR light (808 nm, 0.8–1 W/cm2). The formulations showed significant PDT/PTT activity under NIR light and lower cytotoxicity in the L929 than in RG2 and U87 cell lines. In vivo bioluminescence images and biodistribution studies in U87 tumor-bearing nude mice revealed higher tumor uptake of targeted, IR780-encapsulated liposomal formulations in tumor tissue compared to non-targeted ones. Liposomal formulations showed higher tumor uptake than the free IR780 solution. The data suggest that targeted liposomes have potential as theranostic agents for PDT/PTT and multifunctional glioblastoma imaging.

Citrus aurantifolia-derived carbon quantum dots with red fluorescence emission for codelivery with curcumin as theranostic liposomes for lung cancer

BackgroundCarbon dots (CDs) derived from Citrus aurantifolia represent a promising platform for advanced cancer therapy and diagnostics (theranostics). These CDs are synthesized through a sustainable and cost-effective hydrothermal method, utilizing fruit juice as a green carbon source. Despite the potential, research on the synthesis of citrus-based CDs, especially regarding their red fluorescence emission, which is crucial for enhanced tissue penetration and biomedical efficacy, remains limited.ResultsIn this study, CDs were successfully synthesized from C. aurantifolia fruit, yielding nanoparticles below 5 nm in size (PDI 0.231 ± 0.04). Characterization revealed favorable optical properties, including excitation-dependent fluorescent behavior with prominent red emission under higher excitation wavelengths, a quantum yield of 8.17%, and stable photoluminescence. Chemical composition analysis using XPS, FTIR, and XRD confirmed the purity and structure of the CDs.To explore their biomedical application, CDs were co-loaded with curcumin into liposomes. The formulations had a mean size of 177.2 ± 3.6 nm (PDI 0.270 ± 0.012), demonstrated efficient drug entrapment (60.32 ± 2.24%), and exhibited rapid release kinetics, with 90.21 ± 2.16% of the drug release within 8 h. In vitro studies using A549 lung cancer cells demonstrated superior cellular uptake and cytotoxicity of Cur-CD-loaded liposomes compared to curcumin alone (Cur-Suspension), achieving IC50 values of 0.093 ± 0.011 µg/ml and 0.016 ± 0.006 µg/ml, respectively.ConclusionThis research underscores C. aurantifolia as a viable natural source for green CD synthesis. The obtained CDs with red fluorescence emission, optimized through reaction conditions and excitation wavelengths, hold promise for enhanced biological applications, particularly in the realm of lung cancer therapy. The findings advocate for further exploration and refinement of citrus-based CDs as versatile theranostic agents, capitalizing on their sustainable origins and potent biomedical properties. The combination of citrus-derived CDs with curcumin loaded into liposomal formulations represents a potent theranostic strategy for lung cancer treatment, leveraging the unique properties of CDs and their potential for targeted and effective therapy.Graphical abstract

Hypericin-loaded in modified theranostic liposome nanoplatform: a preliminary in vivo study of targeting and diagnosis.

Modified theranostic liposomes were created by combining phospholipid 1,2-dipalmitoyl-sn-3-glycerol-phosphatidylcholine with two previously modified Pluronic® copolymers covalently linked with spermine and folic acid to carry and stabilize the photosensitizer compound hypericin. After physicochemical characterization, the photocytotoxicity was evaluated against different cancer and healthy cells presenting a strong photodynamic effect. The formulation exhibited no photoactivity without illumination and without hypericin. In vivo, pharmacokinetics biodistribution examined the uptake and theranostic potential of this nanoformulation after its intravenous administration in animal models. Fluorescence images revealed the maximum fluorescence between 0.5-4 h post-tail vein injection, making it an appropriate period for photodynamic treatment. The fluorescence of the entire body was monitored for at least 3 days, indicating that the theranostic procedures can be performed within the 0.5-4 h range after administration, after which the intensity decreases, indicating a potent metabolic ability with no significant side effects. The fluorescence images of the main organs consistently showed a signal during the 1st day of its application. After 48 h, only residues of the modified theranostic formulation were detected in the lungs and thyroid. The promising pharmacokinetics observed in our preliminary studies highlight the potential of this system, making it a worthy candidate for further investigation with tumor models.

Tracking Drugs and Lipids: Quantitative Mass Spectrometry Imaging of Liposomal Doxorubicin Delivery and Bilayer Fate in Three-Dimensional Tumor Models.

Targeted therapy to the tumor would greatly advance precision medicine. Many drug delivery vehicles have emerged, but liposomes are cited as the most successful to date. Recent efforts to develop liposomal drug delivery systems focus on drug distribution in tissues and ignore liposomal fate. In this study, we developed a novel method to elucidate both drug and liposomal bilayer distribution in a three-dimensional cell culture model using quantitative matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI qMSI) alongside fluorescence microscopy. Imaging liposomal distribution in a cell culture model is challenging, as lipids forming the bilayer are endogenous to the model system. To resolve this issue, we functionalized the bilayer by chemically cross-linking a fluorescent tag to the alkyne-containing lipid hexynoyl phosphoethanolamine (HPE). We synthesized liposomes incorporating the tagged HPE lipid and encapsulated within them doxorubicin, yielding a theranostic liposome capable of both drug delivery and monitoring liposomal uptake. We employed an "in-tissue" MALDI qMSI approach to generate a calibration curve with R2 = 0.9687, allowing for quantification of doxorubicin within spheroid sections at multiple time points. After 72 h of treatment with the theranostic liposomes, full doxorubicin penetration was observed. The metabolites doxorubicinone and 7-deoxydoxorubicinone were also detected after 48 h. Modification of the bilayer allowed for fluorescence microscopy tracking of liposomes, while MALDI MSI simultaneously permitted the imaging of drugs and metabolites. While we demonstrated the utility of our method with doxorubicin, this system could be applied to examine the uptake, release, and metabolism of many other liposome-encapsulated drugs.

68 Ga-labeled, imatinib encapsulated, theranostic liposomes: Formulation, characterization, and in vitro evaluation of anticancer activity.

Cancer is still a global health problem. Among cancer types, breast cancer is the most frequently diagnosed one, and it causes a high mortality rate if not diagnosed in the early stages. In our study, imatinib encapsulated, nanosized, neutral/cationic liposome formulations were prepared as theranostic agents for breast cancer. After the characterization studies in which all liposomes exhibited proper profile owing to their particle size between 133 and 250 nm, polydispersity index valueslower than 0.4, neutral and cationic zeta potential values, and high drug encapsulation efficiency, controlled drug release behaviors with zero-order kinetic were obtained. The higher than 90% radiolabeling efficiency values were obtained thanks to the determination of optimum radiolabeling condition (80°C temperature, 5 mCi radioactivity, and 10 min incubation period). According to the resazurin assay evaluating the cytotoxic profile of liposomes on MCF7 cells, neutral empty liposome was found as biocompatible, while both cationic liposomes (empty and drug-loaded ones) exhibited high nonspecific cytotoxicity at even low drug concentration due to the existence of stearyl amine in the formulations. However, dose-dependent cytotoxic effect and the highest cellular binding capacity were obtained by imatinib loaded neutral liposomes. In conclusion, 68 Ga-radiolabeled, imatinib-loaded, neutral, nanosized liposome formulation is the most promising one as a theranostic agent among all formulations.

Hybrid ultrasound-activated nanoparticles based on graphene quantum dots for cancer treatment

Theranostic liposomes have recently found a broad range of applications in nanomedicine due to stability, the high solubility of biomacromolecules, bioavailability, efficacy, and low adverse effects. However, the limitations of liposomes concerning the short systemic circulation in the body, limited controllability of the release rate, and the inability of in vivo imaging remain challenging. Herein, the development of novel hybrid ultrasound-activated piezoelectric nanoparticles based on a hybrid liposome nanocarrier composed of poly(vinylidene fluoride‐trifluoroethylene), graphene quantum dots (GQDs), and Silibinin (a hydrophobic drug) is presented. The hybrid nanoparticles are an acoustically sensitive drug delivery platform that releases the biomacromolecules in a specific tissue area (through surface labeling with PD-1 antibody) in a non-invasive and controlled manner. We show that the developed hybrid nanoparticles (with an average outer diameter of ∼ 230 ± 20 nm) enable piezoelectric-stimulated drug delivery combined with simultaneous fluorescent imaging of cancer cells in vivo. Significant enhancement (>80 % up to 240 h) and tunable drug release from the nanocarrier through enhanced diffusion from the liposome membrane are demonstrated. Cytotoxicity assays using MCF-7, 4T1, and NIH3T3 cell lines exhibit no confrontational influence of nanoparticles on cell viability up to 125 µg/ml. The PD-1 antibody on the surface of the hybrid nanocarrier allows for selective delivery to breast cancer tumors and low biodistribution to other tissues. Our results affirm that the developed ultrasound-activated piezoelectric nanoparticles have great potential as multifunctional platforms with sustainable release profiles for the delivery of hydrophobic drugs to breast cancer, especially when the ability for adequate labeling and cell monitoring is valued.

Enhanced delivery of theranostic liposomes through NO-mediated tumor microenvironment remodeling.

Highly efficient delivery of nanoagents to the tumor region remains the primary challenge for cancer nanomedicine. Herein, we propose a NO-mediated tumor microenvironment (TME) remodeling strategy for the high-efficient delivery of nanoagents into tumor. Quantum dots (QDs) with bright fluorescence in the near-infrared IIb (NIR-IIb, 1500-1700 nm) window and high photothermal conversion efficiency were encapsulated into liposomes for the imaging-guided photothermal therapy (PTT) of tumor. The fabrication of PEG and arginine-glycine-aspartate (RGD) peptide on liposomes ensured the prolonged circulation in vivo and active targeting to tumor. Moreover, the loading of a natural NO generator L-arginine in liposomes realized the continuous generation of NO in the acidic TME. By co-localization fluorescence imaging and western blot of tumor tissue, we confirmed that the release of NO activated the expression of metalloproteinases in TME and further degraded Collagen I in the peripheral region of the tumor, thus removing the barrier for the permeation of liposomes. Attributed to the enhanced accumulation of liposomes inside the tumor, NIR IIb imaging-guided PTT was achieved with remarkable therapeutic efficacy.

Application of Asymmetrical Flow Field-Flow Fractionation for Characterizing the Size and Drug Release Kinetics of Theranostic Lipid Nanovesicles.

Liposome size and in vitro release of the active substance belong to critical quality attributes of liposomal carriers. Here, we apply asymmetric flow field-flow fractionation (AF4) to characterize theranostic liposomes prepared by thin lipid film hydration/extrusion or microfluidics. The vesicles’ size was derived from multi-angle laser light scattering following fractionation (AF4) and compared to sizes derived from dynamic light scattering measurements. Additionally, we adapted a previously developed AF4 method to study zinc phthalocyanine (ZnPc) release/transfer from theranostic liposomes. To this end, theranostic liposomes were incubated with large acceptor liposomes serving as a sink (mimicking biological sinks) and were subsequently separated by AF4. During incubation, ZnPc was transferred from donor to acceptor fraction until reaching equilibrium. The process followed first-order kinetics with half-lives between 119.5–277.3 min, depending on the formulation. The release mechanism was postulated to represent a combination of Fickian diffusion and liposome relaxation. The rate constant of the transfer was proportional to the liposome size and inversely proportional to the ZnPc/POPC molar ratio. Our results confirm the usefulness of AF4 based method to study in vitro release/transfer of lipophilic payload, which may be useful to estimate the unwanted loss of drug from the liposomal carrier in vivo.

Insight into theranostic nanovesicles prepared by thin lipid hydration and microfluidic method

Liposomes are phospholipid-based self-assembled nanoparticles. Various components can be solubilized in the lipid bilayer, encapsulated in the aqueous core or attached to the surface, making liposomes attractive platforms for multimodality functionalization. Here we describe theranostic liposomes delivering a magnetic resonance contrast agent (lipid derivative of gadopentetic acid) and a hydrophobic photosensitizer (zinc phthalocyanine, ZnPc) for photodynamic therapy of cancer. For the first time, this theranostic system was prepared by the microfluidic method. Analogous formulations were produced by thin lipid film hydration (TLH) with down-sizing performed by extrusion for comparison purposes. We demonstrated double the loading capacity of ZnPc into liposomes made by microfluidics compared to TLH/extrusion. Microfluidics resulted in the theranostic nanoliposomes characterized by sizes =2.5x smaller than vesicles prepared by TLH/extrusion. Increased relaxivity was observed for liposomes manufactured by microfluidics compared to TLH, despite a slightly lower Gd chelate recovery. We attributed the improved relaxation to the increased surface area/volume ratio of vesicles and decreased phosphatidylcholine/ZnPc molar ratio, which affected water molecules' diffusion through the liposomal membrane. Finally, we showed photodynamic efficacy of ZnPc loaded into theranostic liposomes in head and neck cancer model, resulting in IC50 of 0.22 – 0.61 μM, depending on the formulation and cell line used.We demonstrate microfluidics' feasibility to be used for theranostic liposome manufacturing and co-entrapment of therapeutic and imaging components in a single-step process with a high yield.

Ligand-targeted Theranostic Liposomes combining methylene blue attached upconversion nanoparticles for NIR activated bioimaging and photodynamic therapy against HER-2 positive breast cancer

Methylene Blue (MB) based Photodynamic therapy (PDT) has gained much interest recently due to its FDA approval for treating various diseased conditions in human. Herein, we report for the first time the systematic evaluation of NIR-controlled PDT efficiency of free MB and MB attached on NaYF4:Yb, Er nanoparticles (UCNPs) i.e., MB@UCNPs encapsulated within Liposomes (LPs). The UCNPs act as an upconverting energy source for the photosensitizers dye MB for enhanced ROS generation and utilizing the remnant energy for selective tracking or bioimaging. The specificity of our nano LPs system was achieved by conjugating our newly discovered anti-HER2 peptides by phage display technique and demonstrated its specificity by confocal imaging of HER-2 overexpressing in vitro model. A significant decline in the cell viability by 83% and 64% was achieved by the LPs containing MB@UCNPs and free MB with UCNPs, respectively after 975 nm NIR laser exposure (0.60 W/cm−2) for 5 min continuous exposure. Also, the flowcytometry assay confirmed that enhanced ROS generation capability of LPs with MB@UCNPs was higher than the free MB with UCNPs, which states the relevance of the minimal distance between MB and UCNPs for higher anti-cancer efficiency.

Lipid-Iron Nanoparticle with a Cell Stress Release Mechanism Combined with a Local Alternating Magnetic Field Enables Site-Activated Drug Release.

Simple SummaryA novel active release system magnetic sphingomyelin-containing liposome encapsulated with indocyanine green, fluorescent marker, or the anticancer drug cisplatin was evaluated. The liposomal sphingomyelin is a target for the sphingomyelinase enzyme, which is released by stressed cells. Thus, sphingomyelin containing liposomes behave as a sensitizer for biological stress situations. In addition, the liposomes were engineered by adding paramagnetic beads to act as a receiver of outside given magnetic energy. The enzymatic activity towards liposomes and destruction caused by the applied magnetic field caused the release of the content from the liposomes. By using these novel liposomes, we could improve the drug release feature of liposomes. The improved targeting and drug-release were shown in vitro and the orthotopic tongue cancer model in mice optical imaging. The increased delivery of cisplatin prolonged the survival of the targeted delivery group versus free cisplatin.Most available cancer chemotherapies are based on systemically administered small organic molecules, and only a tiny fraction of the drug reaches the disease site. The approach causes significant side effects and limits the outcome of the therapy. Targeted drug delivery provides an alternative to improve the situation. However, due to the poor release characteristics of the delivery systems, limitations remain. This report presents a new approach to address the challenges using two fundamentally different mechanisms to trigger the release from the liposomal carrier. We use an endogenous disease marker, an enzyme, combined with an externally applied magnetic field, to open the delivery system at the correct time only in the disease site. This site-activated release system is a novel two-switch nanomachine that can be regulated by a cell stress-induced enzyme at the cellular level and be remotely controlled using an applied magnetic field. We tested the concept using sphingomyelin-containing liposomes encapsulated with indocyanine green, fluorescent marker, or the anticancer drug cisplatin. We engineered the liposomes by adding paramagnetic beads to act as a receiver of outside magnetic energy. The developed multifunctional liposomes were characterized in vitro in leakage studies and cell internalization studies. The release system was further studied in vivo in imaging and therapy trials using a squamous cell carcinoma tumor in the mouse as a disease model. In vitro studies showed an increased release of loaded material when stress-related enzyme and magnetic field was applied to the carrier liposomes. The theranostic liposomes were found in tumors, and the improved therapeutic effect was shown in the survival studies.

Gold liposomes for brain-targeted drug delivery: Formulation and brain distribution kinetics

This work was aimed to formulate transferrin (Tf) receptor targeted gold based theranostic liposomes which contain both docetaxel (DCX) and glutathione reduced gold nanoparticles (AuGSH) for brain-targeted drug delivery and imaging. AuGSH was prepared by reducing chloroauric acid salt using glutathione. The co-loading of DCX and AuGSH into liposomes was achieved by the solvent injection technique, and Tf was post-conjugated on the surface of the liposomes using carboxylated Vit-E TPGS (TPGS-COOH) as a linker. The liposomes were characterized for various parameters such as size, shape, surface charge, and drug release. The Tf receptor targeted gold liposomes were evaluated for the cytotoxicity by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) based colorimetric assay and in-vitro qualitative cellular uptake studies using confocal microscopy. The in-vivo site specific delivery of DCX was analyzed by the brain distribution study of DCX in comparison with marketed formulation (Docel™). A sustained drug release of about 70% was observed from liposomes in the span of 72 h. The in-vivo results demonstrated that targeted gold liposomes were able to deliver DCX into the brain by 3.70, 2.74 and 4.08-folds higher than Docel™ after 30, 120 and 240 min of the treatment, respectively. Besides, the results of these studies have suggested the feasibility of Tf decorated AuGSH and DCX co-loaded liposomes as a promising platform for targeted nano-theranostics.

Theranostic verteporfin- loaded lipid-polymer liposome for photodynamic applications

In this study we report a novel theranostic lipid-polymer liposome, obtained from DPPC and the triblock copolymer F127 covalently modified with 5(6)-carboxyfluorescein (CF) for photodynamic applications. Due to the presence of F127, small unilamellar vesicle (SUV) liposomes were synthesized by a simple and fast thin-film hydration method without the need for an extrusion process. The vesicles have around 100 nm, low polydispersity and superb solution stability. The clinically used photosensitizer verteporfin (VP) was entrapped into the vesicles, mostly in monomeric form, with 90% loading efficiency. Stern-Volmer and fluorescence lifetime assays showed heterogeneous distribution of the VP and CF into the vesicles, ensuring the integrity of their individual photophysical properties. The theranostic properties were entirely photoactivatable and can be trigged by a unique wavelength (470 nm). The feasibility of the system was tested against the Glioblastoma multiforme cell line T98G. Cellular uptake by time-resolved fluorescence microscopy showed monomerized VP (monoexponential decay, 6.0 ns) at nucleus level, while CF was detected at the membrane by fluorescence microscopy. The strategy's success was supported by the reduction of 98% in the viability of T98G cells by the photoactivated lipid-polymer liposome with [VP] = 1.0 μmol L−1. Therefore, the novel theranostic liposome is a potential system for use in cancer and ocular disease therapies.

Formulation and invitro evaluation of upconversionnanoparticle-loaded liposomes for brain cancer.

Aim: This work focused on the development of transferrin-conjugatedtheranostic liposomes consisting of docetaxel (DXL) and upconversion nanoparticlesfor the diagnosis and treatment of gliomas. Materials & methods:Upconversion nanoparticles and docetaxel-loaded theranostic liposomes were prepared by a solvent injection method. Formulations were analyzed for physicochemical properties, encapsulation efficiency, drug release, elemental analysis, cytotoxicity and fluorescence. Results: The particle size was around 200nm with spherical morphology and an encapsulation efficiency of up to 75.93%, was achieved for liposomes with an invitro drug release of 71.10%. The IC50 values demonstrated enhanced cytotoxicity on C6 glioma cells with targeted liposomes in comparison with nontargeted liposomes. Conclusion: Prepared theranostic liposomes may be promising for clinical validation after an invitro and invivo evaluation on cell lines and animals, respectively.

Design and in vitro evaluation of folate-targeted, co-drug encapsulated theranostic liposomes for non-small cell lung cancer

Lung cancer is the type that has the highest incidence and mortality rate. Several methods or agents, for the diagnosis and treatment of lung cancer, fall short of when it comes to early and accurate diagnosis and eradication of tumor tissue. In this study, nanosized, folate-targeted, co-drug encapsulated (paclitaxel and vinorelbine), radiolabeled liposomal formulations were designed for effective diagnosis and therapy of Non-Small Cell Lung Cancer. Theranostic liposomes were appropriately observed according to their particle size (about 150–180 nm) and encapsulation efficiency of 15% for PCX and 20% for VNB as well as their zeta potential of about −10 mV. The uptake of actively targeted liposomes was higher than passively targeted ones in H1299 cells, as well as actively targeted formulations were uptaken with a higher rate in H1299 cells than A549 cells. Empty liposomal formulations exhibited biocompatible profile on cells, as well as the anti-cancer efficacy of co-drug encapsulated liposomes was found to be significantly higher than only-one drug encapsulated liposomal formulations. Therefore, theranostic nanosized, radiolabeled, co-drug encapsulated liposomes were found to be potential for the diagnosis and therapy of Non-Small Cell Lung Cancer. Further in vivo studies are surely needed, and our studies are continuing.

DNA biodots based targeted theranostic nanomedicine for the imaging and treatment of non-small cell lung cancer

The light absorption and emission characteristics of DNA biodots (DNA-BD), along with biocompatibility, give them a high potential for use in various medical applications, particularly in diagnostic purpose. DNA, under high pressure and temperature, condenses to form luminescent biodots. The objective of this research is to develop DNA-biodots (BD) loaded and cetuximab conjugated targeted theranostic liposomes of etoposide for lung cancer imaging and therapy. Theranostic liposomes were prepared by using the solvent injection method and characterized for their particle size, polydispersity, zeta potential, encapsulation efficiency, and pH-dependent in-vitro release, SEM, TEM AFM, EDX, and XRD. The t50% (time at which 50% of the drug releases from the preparation) of the formulations was pH-dependent, with a significant increase in the release at lower pH (5.5). To kill A549 adenocarcinoma cells, the etoposide (control) required significantly (p < 0.05) higher drug concentrations in comparison to non-targeted and; the non-targeted formulation required more concentrations in comparison to targeted liposomes. The in-vivo results demonstrated that CTX-TPGS decorated theranostic liposomes could be a promising carrier for lung theranostics due to their nano-size and selectivity towards EGFR overexpressed cells which provided an improved NSCLC targeted delivery of ETP in comparison to the non-targeted and control formulations.

Multifunctional Theranostic Liposomes Loaded with a Hypoxia-Activated Prodrug for Cascade-Activated Tumor Selective Combination Therapy.

Photodynamic therapy (PDT) is becoming a promising therapeutic regimen but is limited by the hypoxic microenvironment in solid tumors and the undesirable post-treatment phototoxicity side effects on normal tissues. To overcome these restrictions and enhance the antitumor therapeutic effect, near-infrared (NIR) light-activated, cancer cell-specific, hypoxia prodrug-loaded chlorin e6 liposomes were developed for tumor selective combination therapy guided by multimodal imaging. The photothermal agent indocyanine green (ICG) and hypoxia-activated prodrug tirapazamine (TPZ) were coencapsulated into the liposomes, followed by modification with cRGD and conjugation with GdIII to form ICG/TPZ@Ce6-GdIII theranostic liposomes (ITC-GdIII TLs). In the ITC-GdIII TLs, both the fluorescence and photodynamic effect of Ce6 were quenched by ICG via fluorescence resonance energy transfer. The ITC-GdIII TLs can effectively reach the tumor site through the enhanced permeability and retention effect as well as the cRGD-mediated active targeting ability. The fluorescence and photodynamic effect of Ce6 can be activated by the photothermal effect of ICG under NIR light. Upon subsequent irradiation with a 660 nm laser, the released Ce6 could kill cancer cells by generating cytotoxic singlet oxygen. Furthermore, the PDT process would induce hypoxia, which in turn activated the antitumor activity of the codelivered hypoxia-activated prodrug TPZ for a combination antitumor effect. The TLs could be utilized for multimodal imaging (fluorescence/photoacoustic/magnetic resonance imaging)-guided cascade-activated tumor inhibition with optimized therapeutic efficiency and minimized side effects, holding great potential for constructing intelligent nanotheranostics.

Theranostic liposomes as nanodelivered chemotherapeutics enhanced the microwave ablation of hepatocellular carcinoma.

Aim: This study aimed to develop indocyanine green- and doxorubicin-loaded liposomes (DILPs) as theranostic nanoplatform for the detection of hepatocellular carcinoma (HCC) and as an efficient chemotherapeutic to enhance microwave ablation. Materials & methods: DILPs were synthesized and thoroughly characterized. Biocompatibility, tumor uptake and accumulation, and synergistic ablation-chemotherapeutic efficiency were systematically explored in them. In addition, human HCC surgical samples were used to test the affinity of DILPs for HCC. Results: The combination of microwave ablation and DILPs enhanced the ablation efficiency of HCC with apparent tumor inhibition. DILPs exhibited excellent diagnostic ability and could detect 2.5-mm HCC lesions via optoacoustic tomography imaging. DILPs had better affinity for human HCC surgical samples compared with normal liver tissue. Conclusion: Theranostic DILPs could serve as promising nanoparticles for treatment and management of HCC in the clinic.

Design and Testing of Dual-Targeted Gd3N@C80-Containing Glioblastoma Theranostics

The majority of the malignant brain tumors are gliomas with glioblastoma being the most common and aggressive type. Due to its extreme infiltrative nature and higher occurrence frequency, finding new glioblastoma therapeutics and diagnostics is a high priority. In line with this, we previously synthesized a glioblastoma-targeting theranostic that images and delivers a therapeutic payload of doxorubicin (DXR) to human xenograft tumors. Herein, a dual-targeted theranostic is synthesized on the hypothesis that targeting multiple receptors would enhance delivery precision. Doxorubicin and endo-fullerene (Gd3N@C80) were encapsulated within liposomes and conjugated with transferrin (Tf) and lactoferrin (Lf). The hydrodynamic size, zeta potential, encapsulation efficiency, and tagging with proteins of the Gd3N@C80- and DXR-loaded liposomes were characterized using dynamic light scattering (DLS), inductively coupled plasma (ICP) mass spectrometer, Fourier-transform infrared spectroscopy (FTIR), and ultraviolet-visible (UV-Vis) spectrophotometry analysis. As verification of efficacy, the Tf and Lf dual-tagged theranostic liposomes were able to significantly induce cell death compared to Lf monotagged theranostics.

Single-particle characterization of theranostic liposomes with stimulus sensing and controlled drug release properties

Stimuli-responsive nanotheranostic systems, integrated with diagnosis and treatment features, have recently emerged and attracted much interest. However, most of the research mainly focuses on the novelty of nanomaterials, and undervalues the significance of single-particle characterization which can provide detailed physical and biochemical information for performance evaluation and heterogeneity assessment. Due to the small particle size and low content of functional modules, high throughput and multiparameter analysis of individual stimuli-responsive nanoparticles still remains challenging. Here, fabricating a reactive oxygen species (ROS)-responsive liposome (Lipo@BODIPY11) and taking it as an example, we report the development of a strategy for theranostic nanoparticle characterization by a laboratory-built nano-flow cytometer (nFCM). Coincident detection of light scatter and fluorescence intensity provided a measure for liposome quality assessment. Theranostic performance referred to stimuli-responsive capability and drug release behavior upon ROS treatment were obtained in minutes. Besides, the dissociation of functional modules from liposomes and the formation of aggregates under high modification degree were revealed, which was otherwise masked by ensemble-averaged methods. At last, consistent results were also observed in intracellular studies. This nFCM-based method provides a comprehensive approach for the proof-of-principle study, heterogeneity assessment and quality control of biochemical nanosensors and theranostic nanomaterials.