The ability to study drug distribution in real time, without disrupting the biological milieu and structure, has become a cornerstone of modern pharmaceutical development. GSK has developed a portfolio of these methods and this work is often performed in collaboration with academic partners within our Bioimaging Expertise Network (BEN). This presentation will review the successes achieved through this approach, highlighting recent advances; outline project specific case studies and demonstrate the integration of the data into drug development decisions. Finally, the current major challenges will be discussed as new opportunities for imaging collaboration.

This Conference Presentation, “A resurgence in nanoparticles: accelerating the clinical translation of nano-based imaging contrast agents,” was recorded at SPIE Photonics West held in San Francisco, California, United States

Our research aims to demonstrate that Raman spectroscopy and imaging can provide a non-invasive, sensitive, and reproducible determination of the rate and extent at which a topically administered drug becomes available at its site of action in the skin. A key objective is to establish experimental methods, combining Raman spectroscopic tools with complementary calibration techniques and mathematical modelling, to prove this hypothesis ex vivo. The ultimate goal is to provide evidence that the routine, facile and non-invasive measurement of drug pharmacokinetics in the skin in vivo is achievable and has considerable potential for application in regulatory science and decision-making.

Ablative fractional laser (AFL) treatment prior to application of a topical drug enhances drug uptake and the resultant coagulation zone (CZ) surrounding each fractional photothermal injury may provide a reservoir for sustained drug release. In this work, we evaluate how morphological changes in the skin after AFL affect the uptake of an intrinsically fluorescent topical antibiotic. Brightfield images of NBTC stained histopathological slides were evaluated using a deep learning approach for semantic segmentation of fractional laser patterns for automatic assessment of laser hole diameter and CZ morphology. Last, collagen denaturation and drug uptake were quantified via polarization and fluorescence microscopy, respectively.

Systemic drug delivery for dermatological conditions yields little drug to the intended site of action resulting in adverse effects. Topical drug delivery is a viable alternative yet the local cutaneous pharmacokinetics (cPK) is relatively under-explored. Product dosing is dependent upon the knowledge of the dose-cPK relationship, which coincides with the pharmacodynamic (PD) activity. Coherent Raman imaging (CRI) can quantify tissue-specific drug localization and elucidate micro-scale cPK estimates, affording a clinically relevant cPK-PD relationship. This demonstration of a dose-cPK relationship utilizing CRI offers a stepping stone for additional formulation evaluation ex vivo.

Macroscopic fluorescence lifetime FRET imaging (MFLI-FRET) presents a much-needed analytical tool to non-invasively quantify drug-receptor engagement in tumors and other organs in preclinical studies. We demonstrate the specificity and sensitivity of MFLI-FRET for direct and robust measurement of trastuzumab-HER2 target engagement in various types of breast and ovarian cancer tumor xenograft models. Simultaneous metabolic imaging with IRDye 800 CW 2-DG reveals that intracellular delivery of drug is associated with 2-DG lifetime and likely reflects tumors’ microenvironment and perfusion state.

Self-amplifying mRNA (SAM), a synthetic RNA vaccine which self-replicates upon delivery into the cytoplasm encapsulated with lipid nanoparticles (LNPs), leads to a strong and sustained immune response. In this study, we investigated SAM-LNP uptake and subsequent SAM release and distribution in baby hamster kidney (BHK-21) cells using coherent anti-Stokes Raman scattering (CARS) and multiphoton imaging techniques. This work demonstrates the significance of multimodal imaging techniques to capture the successful delivery of SAM and the subsequent production of proteins within cells. Our study can be further extended to label-free detection techniques to investigate targeted drug-delivery.

Ex vivo distribution studies of macromolecular drugs, targets, and biomarkers have been typically limited to bulk sampling methods requiring tissue homogenization or 2D imaging. Bulk sampling cannot capture the complex spatial heterogeneity in tumors and 2D imaging techniques limit analysis to a few representative tissue sections. In this work we explored a tissue clearing, immunolabeling, and lightsheet imaging methods to visualize the ex vivo distribution of biologics in a preclinical models. Biologic distribution in intact tumors was assessed at sub-cellular resolution. A data handling and analysis workflow was developed to assess biologic penetration in tumors and influence of vascular heterogeneity. This work demonstrates potential applications to examine biologic drug distribution-efficacy relationships for drug discovery and development.

Cutaneous pharmacokinetics (cPK) can be quantified utilizing a myriad of in vitro and in vivo approaches. However, these techniques provide macroscale cPK for long experimental durations and do not provide cPK information from the moments directly after a topical formulation application, thus missing critical product dosing insight. Furthermore, formulations are typically applied for 5-10 minutes in the clinic and then removed (purposely or accidentally), which requires a high temporal and spatial resolution estimate of the early timepoint cPK. We have developed a 3-D printed applicator to address the unmet need for early time-point cPK quantification pertinent to the clinic.

The primary goal of this study was to track PS-ASO and GalNAc-PS-ASO uptake in two cell cultures as the first step to understand the observations from the clinical studies. The multimodal imaging setup of CARS and 2PF modalities in conjunction with the image analysis pipeline made it uniquely possible to address these challenges. We report here the time-dependent uptake, internalization, and localization differences between GalNAc-PS-ASOs and PS-ASOs in liver cells. We believe our findings will help us form the basis for further investigations with more complex cellular co-cultures and with tissue and animal models.

Stimulated Raman scattering (SRS) is a rapid, nondestructive technique for imaging biomedical specimens with label-free chemical specificity. Here, we demonstrate a sparse spectral sampling SRS imaging approach where a rapidly-tunable dual-output all-fiber optical parametric oscillator is automatically tuned into specific vibrational modes across >1400cm-1. We demonstrate this system via simultaneously monitoring an active pharmaceutical ingredient and excipient within a single formulation, tracking penetration and permeation through distinct layers of ex vivo mouse skin over time with Raman fingerprint and high wavenumber spectral contrast. This novel method is ideally suited to monitor spatiotemporal dynamic changes during topical drug delivery.