Matched Three-Dimensional Organoids and Two-Dimensional Cell Lines of Metastatic Melanoma to the Brain Mirrors Response to Targeted Molecular Therapy

Abstract

INTRODUCTION: Metastatic melanoma to the brain (MMB) represents a significant treatment challenge. Limitations in preclinical models, namely the lack of heterogeneity in cell lines and low throughput of xenograft models, have thwarted novel therapeutic development. Surgically eXplanted Organoids (SXOs) are ex vivo, three-dimensional models that recapitulate genotypic and phenotypic features of parent tumors with minimal extracranial processing. We aim to develop the first known matched patient-derived SXO and cell line of BRAF-mutant MMB to investigate responses to targeted therapy. METHODS: MMB SXOs were created by a novel protocol incorporating techniques for creating glioma and cutaneous melanoma organoids. A BRAFV600K-mutant MMB sample was collected and processed within 30 minutes of resection. SXOs were created using a 1.0 mm biopsy punch. Organoids were cultured in low-adherence plates in an optimized culture medium under continuous orbital rotation without an artificial extracellular scaffold. Organoid media was changed every 48 hours. Concurrently, matched patient-derived cell lines were created and passaged weekly. Drug screens were conducted with the CellTiter-Glo Luminescence Viability Assay (cell line) and ReadyProbe Cell Viability Imaging Kit (SXOs). RESULTS: After an initial shedding period, organoid growth was observed within 3-4 weeks. MMB SXOs retained histological phenotypes of the parent tissue, including pleomorphic epithelioid cells with abundant cytoplasm, large nuclei, focal melanin accumulation, strong Sox10 positivity, and HMB45 positivity. After sufficient growth, organoids could be manually parcellated to increase the number of replicates. Matched SXOs and cell lines demonstrated sensitivity to BRAF-V600 and MEK inhibitors. CONCLUSIONS: Here, we outline the creation of the first known scaffold-free organoid model of MMB. Further study using SXOs will improve the translational relevance of preclinical studies and enable the study of the melanoma tumor microenvironment.