Modeling micro-invasion of 3D head and neck carcinoma and efficacy of ALA-PDT

Significance: Oral cavity cancers are common worldwide, with over 389,000 new cases and nearly 188,000 deaths reported in 2022. About 70% of cases are diagnosed at Stage III or IV. Oral squamous carcinoma (OSCC) invasion is an early step in the metastatic process, in which tumor cells breach surrounding tissue and eventually disseminate through the vascular system. OSCC is often accompanied by oral submucosal fibrosis (OSMF), and this fibrotic microenvironment creates a permissive niche that promotes invasive progression and increased epithelial-mesenchymal transition, both of which are associated with chemoresistance. Aim: We aim to develop 3D-OSCC culture models of early invasion that mimic in-vivo physiological conditions and use these models to study response to photodynamic therapy (PDT) using 5-aminolevulinic acid (ALA) in problematic ECM-infiltrating invasive cell populations. Approach: TR146 OSCC spheroids were transplanted into extracellular matrix (ECM; supplemented with type I collagen). Imaging of invasive OSCC spheroids was performed before and after ALA photosensitization and following light delivery using a 635 nm laser source. Images were segmented to analyze differential responses to PDT in the primary spheroid and in surrounding ECM-invasive cells. Results: ECM-invading cells exhibited phenotypic traits consistent with increased EMT and also exhibited higher sensitivity to PDT than cells in the primary nodule. Conclusion: ALA PDT is effective against ECM-invading cells that are associated with invasive progression and poor outcomes. PDT may also serve as an effective adjuvant to surgery, destroying residual invasive cells in the resection bed that would otherwise lead to recurrence.