Skin-integrated melanoma organoids to study tumor microenvironment-mediated mechanisms of invasion in vitro.

Summary: Abstract Body: Resistance to targeted therapies remains a challenge in metastatic melanoma. The tumor microenvironment (TME) is a promising therapeutic target, as it influences cancer progression and drug response. The lack of human in vitro 3D models that reproduce the melanoma-TME crosstalk has limited the development of effective therapies. We generated skin-integrated melanoma-TME organoids (mTMEOs) that incorporate key TME components and can serve as a platform for investigating the TME-mediated mechanisms of metastasis in vitro. Primary human skin cells were co-aggregated with GFP+ melanoma cells (SK-MEL-28 or A375) under low-adhesion conditions, forming self-assembling melanoma-skin organoids (mSOs) that mimic the skin’s structure and cellularity. For the mTMEOs, mSOs were embedded in fibroblast-laden collagen gels simulating the dermal stroma. Tumor invasion rate and migration distances were monitored via fluorescence imaging and quantified using an ImageJ-based pipeline to track GFP+ signals. mSOs recapitulated skin features, including a layered epidermis and dermal-hypodermal core, as well as melanoma-specific traits like pagetoid spread, atypical melanocytes, and spreading. mTMEOs modeled the radial-to-vertical growth phase transition, enabling tumor invasion into the dermis-like gel. Stromal fibroblasts enhanced migration distances and led to fewer but larger melanoma clusters, suggesting the TME’s tumor-supportive role by increasing melanoma movement and shift toward collective cell migration, linked to more aggressive metastases. mTMEOs replicate melanoma TME features, including skin architecture, stromal fibroblasts, and collagenous ECM. They provide a tunable platform for studying TME-driven invasion mechanisms and therapy testing. By incorporating patient-derived melanoma cells, these constructs could become a personalized drug-testing tool to improve patient outcomes. Gemma Nomdedeu-Sancho¹, Nicholas Edenhoffer¹, Anastasiya Gorkun¹, Christine Ahn², Anthony Atala¹, Shay Soker¹ 1. Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, United States. 2. Department of Dermatology, Wake Forest University School of Medicine, Winston-Salem, NC, United States. Pigmentation, Melanoma, and Melanoma Immune Surveillance