Abstract 3923: Ubiquitin ligases implicated as predictive biomarkers for poor outcome to immunotherapy in melanoma patients
Authors: Beaton Nigel , Treiber Tobias , Juncker-Jensen Anna , Tan Xin-Xing , Gomez Julia Martinez , Levesque Mitchell , Vowinkel Jakob 
Published: June 2022

Abstract
Background From a medical standpoint, melanoma is considered the most serious form of skin cancer. Immunotherapy treatment regimes frequently target the PD1 pathway and can be highly effective. Despite successes some patients remain unresponsive and a better understanding of what underlies these non-responders could enable better patient treatment stratification and an overall higher positive response rate. Here using unbiased mass spectrometry (LC-MS) proteomics profiling, we sought to identify protein biomarker signatures in patients with metastatic melanoma that correlate with the patients’ intervention responses. Via this discovery-based approach we identify a panel of protein biomarkers that correlate with an improved treatment response and could serve as a panel for patient stratification prior to commencement of immunotherapy regimes.
Methods Formalin-fixed paraffin-embedded (FFPE) tumor samples were used for unbiased quantification of proteins using data-independent acquisition (DIA) LC-MS technology and Biognosys’ Spectronaut software. Patient samples were analyzed at the start of treatment (PD1 inhibitor) and classified as responders (n = 9) or non-responders (n = 15) at 3 months post-treatment. Significant protein expression changes between the two groups was determined by t-test (p &lt; 0.01, log2 fold-change &gt; 0.58) and further machine learning-assisted analysis was conducted. Additionally, patient’s tumor mutational burden (TMB) was analyzed.
Results Analysis of the FFPE samples from anti-PD1-treated metastatic melanoma patients resulted in the identification and quantification of more than 9,000 proteins in total with an average of nearly 8,000 proteins per sample. Stratification of the patients into responders and non-responders led to the identification of 103 proteins that are significantly up- or down-regulated between the groups. Interestingly, proteins associated with ubiquitination pathways were significantly enriched among the identified regulated proteins with 7 proteins having significantly lower abundance in responders when compared to non-responders including various members of the ubiquitin ligase family. Further, a proteomic signature was identified that characterizes the responder and non-responder groups.
Conclusions In this study we demonstrate the power of unbiased proteomic profiling for the characterization of melanoma patients and their responses to PD1-targeted immunotherapy. We identify a set of biomarkers associated with ubiquitination pathways that correlate with treatment outcome although these findings need to further be confirmed in orthogonal cohorts.
Citation Format: Nigel Beaton, Tobias Treiber, Anna Juncker-Jensen, Xin-Xing Tan, Julia Martinez Gomez, Mitchell Levesque, Jakob Vowinkel. Ubiquitin ligases implicated as predictive biomarkers for poor outcome to immunotherapy in melanoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3923.