1165P - Single cell spatial features of in-transit melanoma associated with patient outcome to immunotherapy

Background

Melanoma in-transit metastasis (ITM) occurs in up to 20% of patients with cutaneous melanoma. The 10-year melanoma-specific survival for patients with ITM varies from 75% to 43%. Anti-PD-1-based immunotherapy is used when patients develop unresectable stage III disease or distant metastasis (stage IV). The tumour microenvironment (TME) of ITM melanoma remains poorly understood, where distinct cellular and spatial features can influence patients’ outcome to therapy. This study aims to characterise the spatial TME in ITM melanoma patients.

Methods

We performed 40-plex PhenoCycler imaging on FFPE whole-tissue sections and CITE-Seq using tumour dissociates from 20 ITM patients treated with immunotherapy, 10 biopsied at baseline and 10 biopsied at the time of progression. Using bioinformatic analysis including spatial neighbourhood characterisation and receptor-ligand analyses, we investigated the TME features in ITM melanoma.

Results

Spatial neighbourhood analysis identified tertiary lymphoid structures (TLSs) consisting of B cells, CD8⁺ T cells and dendritic cells, and these immune cells are interfacing HLA-A^high melanoma at the tumour margin. An enrichment for TLSs in pre-treatment ITM was associated with good immunotherapy outcome. Distinct subtypes of macrophages were stratified by phenotypic marker expression and spatial location, and intratumoural enrichment of M1-like macrophages (CD68⁺HLA-DR⁺CD107a⁺CD163^-) was associated with immunotherapy response. Intra-tumoural melanoma heterogeneity was demonstrated in ITMs from patients with primary resistance to treatment and at progression. The resistant TME subtypes displayed a multitude of immune evasive phenotypes, including the upregulation of immune checkpoint receptors (LAG3, VISTA, IDO1), high proliferation, collagen deposition, and low immune recruitment. Single cell sequencing analysis demonstrates cellular programs correlated with response and resistance features in ITM patients.

Conclusions

Taken together, these findings provide insights into the spatial TME interactions associated with immunotherapy response, facilitating the development of biomarkers and therapeutic targets.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Melanoma Institute Australia.

Disclosure

All authors have declared no conflicts of interest.