High-Plex Spatial RNA Profiling Reveals Cell Type‒Specific Biomarker Expression during Melanoma Development

Maija Kiuru,Michelle A Kriner,Samantha Wong,Guannan Zhu,Jessica R Terrell,Qian Li,Margaret Hoang,Joseph Beechem,John D Mcpherson

doi:10.1016/j.jid.2021.06.041

Abstract

Early diagnosis of melanoma is critical for improved survival. However, the biomarkers of early melanoma evolution and their origin within the tumor and its microenvironment, including the keratinocytes, are poorly defined. To address this, we used spatial transcript profiling that maintains the morphological tumor context to measure the expression of >1,000 RNAs in situ in patient-derived formalin-fixed, paraffin-embedded tissue sections in primary melanoma and melanocytic nevi. We profiled 134 regions of interest (each 200 μm in diameter) enriched in melanocytes, neighboring keratinocytes, or immune cells. This approach captured distinct expression patterns across cell types and tumor types during melanoma development. Unexpectedly, we discovered that S100A8 is expressed by keratinocytes within the tumor microenvironment during melanoma growth. Immunohistochemistry of 252 tumors showed prominent keratinocyte-derived S100A8 expression in melanoma but not in benign tumors and confirmed the same pattern for S100A8's binding partner S100A9, suggesting that injury to the epidermis may be an early and readily detectable indicator of melanoma development. Together, our results establish a framework for high-plex, spatial, and cell type‒specific resolution of gene expression in archival tissue applicable to the development of biomarkers and characterization of tumor microenvironment interactions in tumor evolution.

Highlights

Melanoma, the deadliest of the common skin cancers, is curable with early diagnosis and treatment (Gershenwald et al, 2017)
To address the challenges outlined above, we took a discovery-based approach for of studying tumor-microenvironment interactions during melanoma evolution within the native ro morphological context of the tumor, including within the keratinocyte microenvironment. -p we examined expression of over 1,000 genes (Supplementary Table S1) in 134 e regions of interest (ROIs) enriched for melanocytes, neighboring keratinocytes or immune cells Pr in patient-derived formalin-fixed, paraffin-embedded (FFPE) tissue sections from 12 l melanocytic tumors, ranging from benign to malignant, using the NanoString GeoMx® Digital na Spatial Profiler (DSP) (Beechem, 2020) (Figure 1)
We discovered that the damage-associated ur molecular pattern (DAMP) S100A8, which is a known melanoma marker (10) thought to be Jo expressed by immune cells (18), is keratinocyte-derived in melanoma and confirmed this finding by immunohistochemistry (IHC) in a cohort of 252 melanomas

Summary

Introduction

The deadliest of the common skin cancers, is curable with early diagnosis and treatment (Gershenwald et al, 2017). Previous studies have revealed the importance of keratinocyte-derived growth factors and cell adhesion molecules in limiting melanocyte proliferation in normal skin and elucidated mechanisms by which malignant melanocytes escape this regulation (Haass et al, 2005, Villanueva and Herlyn, 2008). These experiments relied on the use of co-culture systems or heterologous expression of keratinocyte-derived genes in melanocytes, neither of which capture the spatial element of melanocyte-keratinocyte interactions in situ. Since scRNAseq rely on fresh tissue, studies on benign melanocytic tumors in humans are lacking

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Results