ICB Seminar


The metastatic melanoma ecosystem during RAF/MEK inhibition in patients

Denis Schapiro, UZH


Monday, December 17th, 2018, 11:30 am, Seminar Room 121, Building 58a

Invited by Prof. Dr. Dr. Fabian Theis


RAF/MEK inhibitors produce rapid responses in patients with BRAFV600E/K mutated advanced/metastatic melanoma. However, virtually all patients experience disease progression and while resistance mechanisms have been described, in most patients the mechanism of resistance is unclear. The effect of therapy on the tumor-microenvironment (TME) and how the architecturally preserved tumor ecosystem may contribute to the development of drug resistance remains poorly understood. This is in large part due to difficulties accessing patient material through sequential biopsy programs and due to technical challenges routed in the lack of technologies enabling measurements of dozens of molecular markers with single-cell spatial resolution.

Here we present a study including eight BRAFV600E/K melanoma patients treated with RAF/MEK inhibitors, who underwent pre-treatment, on-treatment and post-progression biopsies. Formalin-fixed, paraffin-embedded (FFPE) tissue slides from these patients were characterized using tissue-cyclical immunofluorescence (t-CyCIF) to generate 30-plex staining of the malignant and non-malignant components of the tumors at single-cell resolution. Hence, the here presented open-source protocols and analysis pipelines successfully solve the previously described technical challenges.

In total, we have analyzed over 14.7 million single cells and determined the phenotypic and spatial evolution of cell autonomous signaling (p-ERK output), immune infiltrates, and stroma. This analysis recapitulated previously described findings from single-plex immunohistochemistry (IHC) studies and provided several new insights into the ecosystem evolution, potential adaptive resistance mechanisms, and evidence for rationale combination of immune checkpoint inhibitors and RAF/MEK therapies that may delay or prevent the emergence of drug resistance. Along with the analyses, we provide single cell high-resolution images as a rich resource for additional discovery by the broader single-cell community (www.CyCIF.org).

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