Integrated immune profiling approach for immuno-oncology: Unveiling biomarkers and therapeutic targets with mass cytometry.

Abstract

e14575 Background: Over the last decade, immunotherapy has revolutionized cancer treatment, offering transformative outcomes for many patients. However, its effectiveness in treating solid tumors remains limited due to challenges such as T cell exhaustion, immunosuppressive tumor microenvironment (TME), lack of tumor-specific antigens and a complex immune landscape. Solid tumors also induce systemic immune changes that evolve dynamically with tumor progression. To improve patient outcome, it is essential to understand the interplay between localized and systemic immune responses. Methods: To address these challenges, we employed an integrative approach leveraging CyTOF and Imaging Mass Cytometry (IMC) technologies. Matched samples, including peripheral blood mononuclear cells (PBMCs), tumor-derived cells (TDCs) and formalin-fixed, paraffin-embedded (FFPE) tumor tissues from cancer patients, were analyzed. A custom CyTOF panel with over 40 markers was designed to profile immune subsets, checkpoint molecules, cytokines and transcription factors. Samples were barcoded, stained and analyzed using a CyTOF XT system. For spatial immune profiling, FFPE tumor tissues were stained with a 40-plus-marker IMC panel to characterize immune, stromal and tumor cells and assess immune activation states. Data was acquired with the Hyperion XTi Imaging System, enabling high-resolution imaging of tumor microenvironments. Results: CyTOF analysis revealed heterogeneity in T cell exhaustion and activation profiles in PBMCs and TDCs, highlighting the functional diversity of immune responses. IMC technology provided spatial insights into immune cell interactions within the TME, identifying distinct patterns of immune infiltration and congregation around tumor cells and stromal elements. Comparative analysis of CyTOF and IMC data uncovered shared and divergent phenotypes between peripheral and tumor-resident immune compartments, identifying biomarkers that link systemic immune profiles with TME dynamics. Conclusions: These findings offer invaluable insights into the immunological mechanisms driving antitumor responses. By integrating functional and spatial immune profiling, our approach identifies predictive biomarkers and potential therapeutic targets, guiding the development of personalized immunotherapies. This work highlights the importance of understanding systemic and localized immune responses to improve cancer treatment outcomes and optimize treatment strategies. For Research Use Only. Not for use in diagnostic procedures.