Digital spatial profiling of melanoma shows CD95 expression in immune cells is associated with resistance to immunotherapy

ABSTRACT

Although immune checkpoint inhibitor (ICI) therapy has dramatically improved outcome for metastatic melanoma patients, many patients do not benefit. Since adverse events may be severe, biomarkers for resistance would be valuable, especially in the adjuvant setting. We performed high-plex digital spatial profiling (DSP) using the NanoString GeoMx® on 53 pre-treatment specimens from ICI-treated metastatic melanoma cases. We interrogated 77 targets simultaneously in four molecular compartments defined by S100B for tumor, CD68 for macrophages, CD45 for leukocytes, and nonimmune stromal cells defined as regions negative for all three compartment markers but positive for SYTO 13. For DSP validation, we confirmed the results obtained for some immune markers, such as CD8, CD4, CD20, CD68, CD45, and PD-L1, by quantitative immunofluorescence (QIF). In the univariable analysis, 38 variables were associated with outcome, 14 of which remained significant after multivariable adjustment. Among them, CD95 was further validated using multiplex immunofluorescence in the Discovery immunotherapy (ITX) Cohort and an independent validation cohort with similar characteristics, showing an association between high levels of CD95 and shorter progression-free survival. We found that CD95 in stroma was associated with resistance to ICI. With further validation, this biomarker could have value to select patients that will not benefit from immunotherapy.

KEYWORDS:

• Biomarkers
• DSP
• immunotherapy
• melanoma

Acknowledgments

This work was supported by the Yale SPORE in Skin Cancer P50 CA121974 (M. Bosenberg and H. Kluger, PIs) and by a sponsored research agreement from Navigate Biopharma (DLR). The authors also acknowledge the expert assistance of Lori Charette and the staff of the Yale Tissue Microarray Facility division of Yale Pathology Tissue Services for construction of the TMAs used in the study.

Disclosure statement

SMM is a current employee at Boehringer Ingelheim Inc. PD is a current employee and stockholder at NanoString Technologies; PFW currently works at Verily Life Sciences, Alphabet Inc. (CA, USA); H.M. Kluger reports receiving commercial research grants from Merck, Bristol-Myers Squibb, and Apexigen, and is a consultant/advisory board member for Alexion, Corvus, Nektar, Biodesix, Genentech, Merck, Celldex, Pfizer, Iovance, Array Biopharma, Clinigen, Bristol-Myers Squibb, Instil Bio and Immunocore. D.L. Rimm declares that in the last two years he has served as a consultant to Astra Zeneca, Amgen, BMS, Cell Signaling Technology, Cepheid, Daiichi Sankyo, Danaher, GSK, Konica/Minolta, Merck, NanoString, Novartis, PAIGE.AI, Perkin Elmer/Akoya, and Roche/Ventana. No potential conflict of interest was reported by the author(s).

Authors’ contributions

SMM and DLR participated in the concept, design, and planning of the study. SMM and MM developed the methodology. SMM, SJ, PFW, and HMK constructed the discovery and validation ITX cohorts and conducted clinical data collection. SMM and MM acquired the data. SMM, AF, MM, PD, and RGM analyzed the data. SMM and DLR interpreted the results and drafted the manuscript. All authors critically reviewed, edited the manuscript, and approved the final draft for submission.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/2162402X.2023.2260618

Additional information

Funding

This work was funded by a sponsored research agreement from Navigate Biopharma and in part by the Yale SPORE in Skin Cancer, P50 CA121974 (M. Bosenberg and H. Kluger, PIs).