Development and Validation of Characterization Methods for Lipidots® Multifunctional Platform: a Step forward Industrial Transfer

M. Varache, M. Ciancone, F. Caputo, C. Laffont, M. Escudé, D. Jary, P. Boisseau, I. Texier, F.P. Navarro, A.-C. Couffin

Keywords: lipid nanocarriers, nanomedicine, physical-chemical characterizations, regulatory health agencies


For eight years, our research group has been developing lipid nanocarriers, known as Lipidots®, as a multifunctional platform for medical applications. A wide range of active pharmaceutical ingredients (APIs) and contrast imaging agents has been encapsulated into Lipidots® (including Paclitaxel, Camptothecine, Budenoside, Cyclosporine A, photosensitizers and fluorescent dyes), enabling the delivery of therapeutic agents and the monitoring or diagnosis by fluorescence imaging [1-2]. As a nanocarrier, Lipidots® have been proved to possess a high colloidal stability over months. Particle size and viscosity of the core can be tuned, resulting in high payloads, controlled drug release profile and improved drug bioavaibility to appropriate biological sites [3-4]. In addition to size and surface charge of the nanomaterials, as key parameters requested by regulatory health agencies, a more deep understanding of nanoparticles composition, morphology, aggregation, crystallinity, drug loading, drug localization, drug release kinetics and nanoparticles interaction with proteins in biological fluids is crucial to meet the requirements of final medicinal nanoproducts. Complementary physical-chemical characterization methods have been used to determine the morphology, the size, the shape and the surface charge of the Lipidots®. The internal physical state of the particle core has been studied by Differential Scanning Calorimetry (DSC), 1H NMR, and Small-Angle Neutron Scattering (SANS). The quantification and identification of Lipidots® components (e.g. mixture of mono-, di-, and triglycerides, phospholipids and PEGylated surfactants) have been performed by using validated H/UPLC-ELSD methods and mass spectrometry. Moreover, in order to predict the in vivo behavior of the nanotherapeutics, we developed a valuable and transposable tool for separation and quantification of free and entrapped drugs/dyes by combining Solid Phase Extraction and Drug Content Determination (DCD) [5]. Such methods allow studying drug leakage kinetics under storage conditions and the drug release profiles in different biological media. HPLC quantification combined with spectroscopic techniques has been also routinely performed for optimization of payload formulation. Localization of different encapsulated drugs within the nanodroplets has been characterized by NMR spectroscopy and correlated with the loading profiles. Finally, small-angle neutron scattering (SANS) has been performed to study the interaction of the Lipidots® with circulating proteins that can lead to a protein corona as it is well admitted nowadays that bound proteins form the real nano-bio interface governing the properties of the nanoparticles like cell uptake. Such extensive characterization in addition to the ability to scale up the manufacturing process of our carriers, allows identifying characterization criteria and specifications to move towards industrial production and approval of the product for the clinical market. In this communication, we will present an overview of these nanocharacterization techniques, which represent a step forward industrial transfer, and an example of the development of a full cascade of standardized physical-chemical characterization methods for nanomedicines.