V36 - Polymer Lipid Nanoparticles Using Drug-Dye Interactions for Improved Cancer Theranostics

Introduction: : Lapatinib is an FDA-approved dual tyrosine kinase inhibitor for solid tumors, including breast cancer, but its clinical use is limited by poor solubility, low bioavailability, and off-target effects. Lipid nanoparticles (LNPs) have risen to address this issue due to their promising drug delivery system. However, they still suffer from drawbacks such as low drug loading efficiency and susceptibility to immune system recognition. Moreover, the real-time monitoring of the delivery of Lapatinib using LNPs is infeasible without significant modification to the LNP system, hence the need for an imagining agent. This study amplifies the delivery of Lapatinib using next-generation LNPs through a two-step strategy. First, semiconducting polymer dyes are used to boost drug loading via intermolecular interactions and enable near-infrared imaging (Figure 1). Second, CD47-rich red blood cell membrane proteins are integrated onto LNPs to create biomimetic LNPs (bLNPs), promoting immune evasion and prolonged circulation (Figure 1). Our future work includes testing immune evasion with murine macrophages and evaluating toxicity and uptake in breast 2D and 3D cancer models. This approach demonstrates how molecular design and biomimicry can improve LNP-based cancer theranostics.

Materials and

Methods: : All materials were purchased from Sigma-Aldrich unless otherwise specified. Polymer-Lipid NPs were synthesized using a nanoprecipitation technique, followed by ultracentrifugation to purify the nanoprobes from any unreacted dyes, drugs, or surfactants. The nanoprobes were characterized using dynamic light scattering, zeta potential measurements, UV-Vis-NIR absorption spectra, and NIR fluorescence spectra. Red blood cell membrane (RBCm) proteins were extracted using a Mem-PER Plus Membrane Protein Extraction Kit (Thermo Fisher) and quantified using BCA assay. For imparting red blood cell mimicry, LNPs were coated with RBCm (to form bLNPs) under stirring overnight and washed to remove unbounded RBCm proteins. RBCm on bLNPs were quantified with a BCA assay.

Results, Conclusions, and Discussions:: Polymer-lipid LNPs demonstrated stable UV-Vis-NIR absorption spectral and fluorescence emission profiles, confirming the successful formation of Lapatinib-dye LNPs (Figure 2). Due to the formation of intermolecular interactions, the drug and dye formed a complex, and subsequent dynamic light scattering results revealed a single peak corresponding to the drug-dye-loaded LNPs, indicating the creation of a homogeneous nanoparticle population. Zeta potential analysis showed colloidal stability at 4 °C and in 10% human plasma solution. IVIS imaging (Ex: 745 nm, Em: 840 nm) identified SPN-Lap500 as having an optimal candidate where the drug-dye interaction was the maximum (Figure 2) and thus, subsequently used for further experimentations. Protein nano-sprinkling on SPN-Lap500 with RBCm proteins yielded stable, fully incorporated bLNPs, with increased hydrodynamic diameter indicating successful membrane protein integration (Figure 2). A BCA calibration curve confirmed a correlation between protein concentration and absorbance at 562 nm and further quantified the amount of RBCm proteins loaded onto SPN-Lap500.

To address the clinical limitations of Lapatinib, including poor aqueous solubility, limited bioavailability, and off-targeted effects, we developed a next-generation biomimetic lipid nanoparticle (bLNP) delivery system. By leveraging specific intermolecular interactions, Lapatinib was co-formulated with a NIR-I fluorescent dye to create a stable drug-dye complex within the LNPs. Protein nano-sprinkling comprised of LNPs coated with RBCm to induce red blood cell mimicry, resulting in prolonged blood circulation, lowered macrophage uptake, and active targeting of tumor cells through CD47 protein interactions. This biomimetic strategy not only improves the physical properties of Lapatinib but also enables real-time monitoring of nanoparticle distribution, representing a significant advancement in the precision delivery of anticancer therapeutics.

Acknowledgements and/or References (Optional):: The authors want to thank the Edward E. Whitacre Jr. College of Engineering at Texas Tech University.