{
  "abstract": "Background Profiling of tumor resistance is critical for advancing precision oncology in breast cancer. Current immunohistochemistry protocols are constrained by the number of markers that can be assessed concurrently, and reliance on multiple rounds of antigen retrieval and sequential staining. These issues present a high risk of tissue loss and poor signal quality. To address these limitations, we developed a new 17-plex fluorescence immunohistochemistry (fIHC) assay designed to characterize the cell cycle and resistance profiles across diverse breast cancer subtypes. This assay offers a robust, streamlined methodology to provide deeper molecular insights on the pathogenesis of breast cancer and provides support for both patient stratification and therapeutic development.Methods Navigate BioPharma designed a 17-plex fIHC assay with fluorescence-conjugated antibodies targeting key markers involved in cell cycle regulation, tumor resistance, and immune interaction, including AR, CCNE, CD20, CD3e, ER, FAP, HER2, Ki67, p53, PDL1, PR, pRb, Rb, TK1, and Cytokeratin, with a nuclear counterstain (Hoechst). We successfully validated a clinical grade multiplex fIHC assay using automated staining (intelliPath FLX), imaging (Orion) and analysis (HALO and HALO AI). The assay is validated across multiple breast cancer subtypes.Results Sensitivity and specificity were confirmed on positive and negative controls. Assay reproducibility was seen across instruments, operators, and independent experiment runs for all markers. There are over 65,000 unique phenotypes that are possible to characterize using this assay, including the ability to understand co-expression of diagnostic breast biomarkers at a cellular level.Conclusions We have developed and validated a 17-plex fIHC assay to explore cell cycle dynamics and tumor resistance across diverse breast cancer indications. Implementation of this assay may support applications in clinical trials to inform patient stratification, monitoring therapeutic efficacy, and unraveling mechanisms of resistance in breast cancer. By delivering high-dimensional phenotyping, this assay addresses key gaps in understanding tumor biology and immune cell interactions, supporting the development of next-generation breast cancer therapies.",
  "authors": [
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Jacob Levy"
    },
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Sophia Kekchidou"
    },
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Richard Van Krieken"
    },
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Xun Li"
    },
    {
      "affiliations": [
        "Novartis Biomedical Research, Cambridge, MA, USA"
      ],
      "name": "David Yang"
    },
    {
      "affiliations": [
        "Novartis Biomedical Research, Cambridge, MA, USA"
      ],
      "name": "LaSalette Charette"
    },
    {
      "affiliations": [
        "Novartis Biomedical Research, Cambridge, MA, USA"
      ],
      "name": "Ying Huang"
    },
    {
      "affiliations": [
        "Novartis Biomedical Research, Cambridge, MA, USA"
      ],
      "name": "Lisa Kattenhorn"
    },
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Naveen Dakappagari"
    },
    {
      "affiliations": [
        "Navigate BioPharma Services, Inc., a Novartis subsidiary, Carlsbad, CA, USA"
      ],
      "name": "Jennifer Bordeaux"
    }
  ],
  "title": "100 Cell cycle and tumor resistance profiling in breast cancer by a 17-plex immunofluorescence assay on the Orion platform",
  "uid": "c5e73f70-0af0-5416-8908-de8a134e7cba"
}