Photo-triggerable liposomes based on lipid-porphyrin conjugate and cholesterol combination: Formulation and mechanistic study on monolayers and bilayers.

Lipid-porphyrin conjugates are considered nowadays as promising building blocks for the conception of drug delivery systems with multifunctional properties such as photothermal therapy (PTT), photodynamic therapy (PDT), phototriggerable release, photoacoustic and fluorescence imaging. For this aim, we have recently synthesized a new lipid-porphyrin conjugate named PhLSM. This was obtained by coupling pheophorbide-a (Pheo-a), a photosensitizer derived from chlorophyll-a, to egg lyso-sphingomyelin. The pure PhLSMs were able to self-assemble into vesicle-like structures that were however not stable and formed aggregates with undefined structures due to the mismatch between the length of the alkyl chain in sn-1 position and the adjacent porphyrin. Herein, stable PhLSMs lipid bilayers were achieved by mixing PhLSMs with cholesterol which exhibits a complementary packing parameter. The interfacial behavior as well as the fine structures of their equimolar mixture was studied at the air/buffer interface by the mean of Langmuir balance and x-ray reflectomerty (XRR) respectively. Our XRR analysis unraveled the monolayer thickening and the increase in the lateral ordering of PhLSM molecules. Interestingly, we could prepare stable vesicles with this mixture that encapsulate hydrophilic fluorescent probe. The light-triggered release kinetics and the photothermal conversion were studied. Moreover, the obtained vesicles were photo-triggerable and allowed the release of an encapsulated cargo in an ON-OFF fashion.

Photo-triggerable liposomal drug delivery systems: from simple porphyrin insertion in the lipid bilayer towards supramolecular assemblies of lipid-porphyrin conjugates.

Light-responsive liposomes are considered nowadays as one of the most promising nanoparticulate systems for the delivery and release of an active pharmaceutical ingredient (API) in a spatio-temporal manner. Several strategies can be used to design photo-triggered liposomes. One of them consists in the incorporation of a photosensitizer (PS) in the lipid matrix of a liposomal bilayer that induces the release of the cargo either via a photochemical or a photophysical process. Among the described photosensitizers, porphyrin derivatives have appeared as the most potent ones. This review describes the state-of-the-art of photo-triggerable liposomes based on the combination of lipids and porphyrin derivatives either free or conjugated. It focuses on the different light-triggered release mechanisms and the requirements for the development of such systems. It also details the different strategies for the synthesis of lipid-porphyrin conjugates, their self-assembling properties and their biomedical applications.

Newly Synthesized Lipid-Porphyrin Conjugates: Evaluation of Their Self-Assembling Properties, Their Miscibility with Phospholipids and Their Photodynamic Activity In Vitro.

Lipid-porphyrin conjugates are considered nowadays as promising building blocks for the conception of supramolecular structures with multifunctional properties, required for efficient cancer therapy by photodynamic therapy (PDT). The synthesis of two new lipid-porphyrin conjugates coupling pheophorbide-a (Pheo-a), a photosensitizer derived from chlorophyll-a, to either chemically modified lyso-phosphatidylcholine (PhLPC) or egg lyso-sphingomyelin (PhLSM) is reported. The impact of the lipid backbone of these conjugates on their self-assembling properties, as well as on their physicochemical properties, including interfacial behavior at the air/buffer interface, fluorescence and absorption properties, thermotropic behavior, and incorporation rate in the membrane of liposomes were studied. Finally, their photodynamic activity was evaluated on esophageal squamous cell carcinoma (ESCC) and normal esophageal squamous epithelium cell lines. The liposome-like vesicles resulting from self-assembly of the pure conjugates were unstable and turned into aggregates with undefined structure within few days. However, both lipid-porphyrin conjugates could be efficiently incorporated in lipid vesicles, with higher loading rates than unconjugated Pheo-a. Interestingly, phototoxicity tests of free and liposome-incorporated lipid-porphyrin conjugates demonstrated a better selectivity in vitro to esophageal squamous cell carcinoma relative to normal cells.

Optimization of the Production of Covalently Circularized Nanodiscs and Their Characterization in Physiological Conditions.

Lipid nanodiscs are widely used platforms for studying membrane proteins in a near-native environment. Lipid nanodiscs made with membrane scaffold proteins (MSPs) in the linear form have been well studied. Recently, a new kind of nanodisc made with MSPs in the circular form, referred to as covalently circularized nanodiscs (cNDs), has been reported to have some possible advantages in various applications. Given the potential of nanodisc technology, researchers in the field are very interested in learning more about this new kind of nanodisc, such as its reproducibility, production yield, and the possible pros and cons of using it. However, research on these issues is lacking. Here, we report a new study on nanodiscs made with circular MSPs, which are produced from a method different from the previously reported method. We show that our novel production method, detergent-assisted sortase-mediated ligation, can effectively avoid high-molecular-weight byproducts and also significantly improve the yield of the target proteins up to around 80% for larger circular MSP constructs. In terms of the application of circular MSPs, we demonstrate that they can be used to assemble nanodiscs using both synthetic lipids and native lipid extract as the source of lipids. We also show that bacteriorhodopsin can be successfully incorporated into this new kind of cND. Moreover, we found that cNDs have improved stability against both heat and high-concentration-induced aggregations, making them more beneficial for related applications.

Impact of lipid composition and photosensitizer hydrophobicity on the efficiency of light-triggered liposomal release.

Photo-triggerable liposomes are considered nowadays as promising drug delivery devices due to their potential to release encapsulated drugs in a spatial and temporal manner. In this work, we have investigated the photopermeation efficiency of three photosensitizers (PSs), namely verteporfin, pheophorbide a and m-THPP when incorporated into liposomes with well-defined lipid compositions (SOPC, DOPC or SLPC). By changing the nature of phospholipids and PSs, the illumination of the studied systems was shown to significantly alter their lipid bilayer properties via the formation of lipid peroxides. The system efficiency depends on the PS/phospholipid association, and the ability of the PS to peroxidize acyl chains. Our results demonstrated the possible use of these three clinically approved (or under investigation) PSs as potential candidates for photo-triggerable liposome conception.

Quantitative proteomics of protein complexes and their implications for cell reprograming and pluripotency.

Pluripotency of embryonic stem cells (ESCs)/induced pluripotent stem cells (iPSCs) and reprograming of somatic cells (SCs) to pluripotency are governed by known and unknown factors. These factors, including protein complexes, are poorly described at the proteome level. Here, we established the quantitative proteomic profiles across three types of cells (iPSCs, ESCs, and SCs) using OFFGEL fractionation coupled with LTQ-Orbitrp analysis. Additionally, we utilized the previously published proteomic profiles of iPSCs, ESCs, and SCs. By integrating these proteomic profiles with protein-protein interaction resources, we identified numerous protein complexes in iPSCs and/or ESCs, which include known and novel chromatin remodeling complexes that facilitate cell reprograming. The identified protein complexes also include the previously unreported ones that are associated with the imperfect aspects of iPSCs or cell reprograming process. Further, we performed a comparison between our study and previously published studies and highlighted a partial conservation of the identified protein complexes across the iPSCs generated by different laboratories and iPS cell-type specific protein complexes. The identified protein complexes were validated by integrated in silico analysis of microarray repository data related to ESCs differentiation into embryoid bodies. A majority of the protein complexes exhibited significant (p < 0.005) co-regulation of their components upon ESC differentiation, suggesting their role in the maintenance of the pluripotent state. Finally, we showed a link between the components of the protein complexes and embryonic development using the existing loss-of-function phenotype data. Together, our integrated approach provides the first comprehensive view of the protein complexes that may have implications for cell reprograming and pluripotency.