Simple SummaryTriple-negative breast cancer is characterized by a lack of targetable treatment receptors and current standard of care options, such as radiation therapy and chemotherapy, are associated with acute patient toxicity. Noninvasive imaging of PD-L1, with [89Zr]-Atezolizumab-PET imaging, has the potential to identify tumors with increased susceptibility to immunotherapy. In this work, we have optimized the labeling conditions of [89Zr]-Atezolizumab and used noninvasive [89Zr]-Atezolizumab PET imaging to characterize the longitudinal changes in PD-L1 expression in TNBC treated with standard of care treatment options. The goal of this study is to understand how [89Zr]-Atezolizumab PET imaging can characterize changes in intratumoral molecular biology and how this can inform eventual response and combination therapy.AbstractTriple-negative breast cancers (TNBCs) currently have limited treatment options; however, PD-L1 is an indicator of susceptibility to immunotherapy. Currently, assessment of PD-L1 is limited to biopsy samples. These limitations may be overcome with molecular imaging. In this work, we describe chemistry development and optimization, in vitro, in vivo, and dosimetry of [89Zr]-Atezolizumab for PD-L1 imaging. Atezolizumab was conjugated to DFO and radiolabeled with89Zr. Tumor uptake and heterogeneity in TNBC xenograft and patient-derived xenograft (PDX) mouse models were quantified following [89Zr]-Atezolizumab-PET imaging. PD-L1 expression in TNBC PDX models undergoing therapy and immunohistochemistry (IHC) was used to validate imaging. SUV from PET imaging was quantified and used to identify heterogeneity. PET/CT imaging using [89Zr]-Atezolizumab identified a significant increase in tumor:muscle SUVmean1 and 4 days after niraparib therapy and revealed an increased trend in PD-L1 expression following other cytotoxic therapies. A preliminary dosimetry study indicated the organs that will receive a higher dose are the spleen, adrenals, kidneys, and liver. [89Zr]-Atezolizumab PET/CT imaging reveals potential for the noninvasive detection of PD-L1-positive TNBC tumors and allows for quantitative and longitudinal assessment. This has potential significance for understanding tumor heterogeneity and monitoring early expression changes in PD-L1 induced by therapy.