Double-Multiplex Immunostainings for Immune Profiling of Invasive Breast Carcinoma: Emerging Novel Immune-Based Biomarkers.

The role of tumor microenvironment in invasive breast cancer prognosis and treatment is highly appreciated. With the advent of immunotherapy, immunophenotypic characterization in primary tumors is gaining attention as it can improve patient stratification. Here, we discuss the benefits of spatial analysis employing double and multiplex immunostaining, allowing the simultaneous detection of more than one protein on the same tissue section, which in turn helps us provide functional insight into infiltrating immune cells within tumors. We focus on studies demonstrating the prognostic and predictive impact of distinct tumor-infiltrating lymphocyte subpopulations including different CD8(+) T subsets as well as CD4(+) T cells and tumor-associated macrophages in invasive breast carcinoma. The clinical value of immune cell topography is also appreciated. We further refer to how the integration of digital pathology and artificial intelligence in routine practice could enhance the accuracy of multiplex immunostainings evaluation within the tumor microenvironment, maximizing our perception of host immune response, improving in turn decision-making towards more precise immune-associated therapies.