Ho Wan (Howard) Chan

In vitro NSCLC organoid efficacy screening platform for inhaled chemotherapeutics

Lung cancer is the deadliest and 2nd most common cancer type worldwide, accounting for 1.8 million deaths annually. Non-small cell lung cancer (NSCLC) constitutes 85% of lung cancer cases and displays a poor response to chemotherapy. Novel treatment strategies are urgently required to improve NSCLC treatment efficacy and survival rates.

Effective chemotherapy requires efficient delivery to the primary lung tumor. Yet, conventional intravenous infusion can only deliver 5 – 10% of the administered drug dose to the lungs, with even lower penetration into primary tumors. Direct aerosol inhalation enables localized pulmonary chemotherapy accumulation and reduces systemic adverse effects by reducing off-target distribution. However, clinical translation of inhaled chemotherapeutics remains sluggish due to insufficient preclinical models that appropriately predict treatment efficacy. Sub-confluent 2D monolayers or 3D spheroids commonly used for in vitro drug testing do not fully represent human tumor architecture and function, while animal models can be challenging to construct and pose significant animal ethics concerns. Organoids have garnered considerable interest in disease modeling and drug screening as they preserve tumor morphology and genetics after long-term culture while being relatively easy to construct. They are also considered valuable alternatives to animal studies for seeking regulatory approval with growing public sentiment against animal research. However, organoids cultured using standard protocols do not mimic direct contact of the lung epithelia with gas, a key feature of respiratory tract structure and function.
The proposed project aims to develop an NSCLC organoid efficacy screening platform for inhaled chemotherapeutics using inhalable paclitaxel nanoagglomerate dry powder (PTX-NADP; Chan et al., Int J Pharm, 2024;653:123877) as the model formulation. Organoid lines will be established from surgically resected NSCLC tissues by Matrigel-embedded 3D culture and used to generate 3D air-liquid interface (ALI) cultures by seeding organoids onto collagen gel-containing Transwell® inserts to mimic the direct contact of the lung epithelium with the gas surface. Upon successful ALI culture, organoids will be treated with different doses of PTX solution by direct PTX inoculation, nebulized PTX/redispersed PTX-NADP using an Aerogen Solo® nebulizer or aerosolized PTX-NADP by dispersing PTX-NADP using a syringe enclosed by a cone to minimize environmental aerosol contamination. The organoids’ growth curve will be monitored by brightfield microscopy, and staining will be conducted for confocal laser scanning microscopy and flow cytometry evaluation of live/dead cells. Western blots will be performed to probe the molecular antitumor mechanism. If successful, the organoid platform will be extended to evaluate other novel inhaled NSCLC treatments, such as targeted therapies and immunotherapies, ultimately expediting their clinical translation.

My interest in respiratory pharmaceutical research stemmed during my pre-registration pharmacy internship in the early wave of COVID-19. I encountered first-hand the unmet medical needs of patients with various respiratory conditions, including COVID-19, lung cancer, and chronic obstructive pulmonary disease. I was highly fascinated by the application of nanotechnologies in medicine when taking my undergraduate drug delivery courses and my final-year research project during my BPharm studies at the University of Hong Kong (HKU). My interests in inhaled drug delivery and nanotechnology have motivated me to continue my research journey as a PhD student at my alma mater. My PhD research focuses on developing nanoagglomerate dry powder formulations for advanced pulmonary drug delivery. The ongoing research work has already led to the successful development of inhalable remdesivir and paclitaxel nanoagglomerate dry powders with promising therapeutic potential, with two first-author original research articles published in the International Journal of Pharmaceutics and two related first-author review articles published in AAPS PharmSciTech and Journal of Pharmaceutical Sciences.

Despite considerable efforts to develop inhalable nanotherapeutics, very few inhalable nanomedicine products have been approved by regulatory authorities, indicating a translational gap between research and pharmaceutical development. Challenges in clinical translation of inhalable nanomedicines, as discussed in our recently published review article (Chan et al., AAPS PharmSciTech, 2023;24(4):98), include manufacturing scale-up and process control and articulation of preclinical data and clinical consideration. To tackle the challenge of manufacturing scale-up, my PhD research work has led to the successful development of an integrated continuous manufacturing platform for inhalable nanoagglomerate dry powders, which eliminates batch-to-batch variations and provides easier process control and greater scalability (Chan et al., Int J Pharm, 2023;644:123303). Several accolades were won based on this invention, including the HKU Research Postgraduate Student Innovation Award and a Silver Medal at the recent 49th International Exhibition of Inventions Geneva.

This proposed project aims to address the difficulty in articulating preclinical data with clinical considerations for inhalable (nano)chemotherapeutics. We have previously demonstrated the superior in vitro antitumor efficacy of inhalable paclitaxel nanoagglomerate dry powders (PTX-NADP) compared to unformulated paclitaxel in 2D A549 lung adenocarcinoma cell monolayers (Chan et al., Int J Pharm, 2024;653:123877), and studies are ongoing to demonstrate their in vivo efficacy in orthotopic lung cancer models. However, significant challenges are encountered before the successful development of these models due to the surgical skills required for orthotopic tumor inoculation and endotracheal intubation for intratracheal drug administration. These procedures may also cause unintentional pain or discomfort to animals, which poses concerns in animal ethics. Indeed, the use of animals for pharmaceutical research has been an active topic of debate. With the US FDA recently announcing that animal studies are no longer a prerequisite for premarket applications, this ignites my strong research interest in developing a novel analytical method evaluating in vitro performance for inhaled chemotherapeutics with a close correlation to clinical efficacy. To this end, an NSCLC organoid efficacy screening platform for inhaled chemotherapeutics is proposed in my research proposal.

As the proposed work involves clinical specimens of NSCLC tissues, we will collaborate with Prof. David CL Lam, Clinical Associate Professor and Chief of the Respiratory Medicine Division in the Department of Medicine, School of Clinical Medicine, HKUMed and Honorary Consultant in the Department of Medicine in Queen Mary and HKU-Shenzhen Hospitals. Prof. Lam is a respiratory physician with a research interest in translational research in lung cancer and has extensive experience in establishing new lung cancer organoid models in his laboratory. He and his team’s expert insights into the design of the proposed 3D air-liquid interface-cultured organoid models will bring synergy to the project’s success in providing novel clinically relevant models for inhaled chemotherapeutic treatment screening.

The award of the DDL Career Development Grant will allow me to gain exposure to the field of organoid research, which I had not been exposed to in my current studies and are currently not available in my department and inspire me to integrate organoid and organ-on-a-chip technologies in my future research. Organoid antitumor performance results from the proposed project will complement my ongoing in vivo efficacy studies on PTX-NADP to further enrich the quality of the manuscript in preparation for publication in prestigious academic journals. At the same time, through dissemination of the research outcomes of the proposed project at the DDL Conference and other oncology or drug delivery conferences, I aim to polish my presentation skills and initiate further collaborations with other members of the scientific community to enhance translational research of inhaled chemotherapeutics further. I will also explore establishing commercial opportunities upon the successful development of the NSCLC organoid platform, such as inhaled chemotherapeutic screening services or platform licensing via opening a start-up. In the long run, the Grant will assist me in establishing an independent and comprehensive research portfolio in inhaled drug delivery from formulation, manufacturing, and physicochemical characterization to preclinical evaluation, such that I can continue to pursue my initiatives to improve the quality of life of patients suffering from respiratory conditions and allow them to “Inhale the future.”