Early identification of immunotherapy resistance through integrated multiparameter imaging

PROJECT SUMMARY Immunotherapies for cancer have revolutionized oncology, but most patients do not experience durable responses. Immune checkpoint inhibitors targeting PD-1 and PD-L1 are now FDA-approved for a wide variety of tumors, but non-invasive strategies to monitor response and identify mechanisms of resistance have not yet been developed. Such approaches would allow responsive patients to remain on therapy and potentially avoid the toxicity associated with other therapies, such as chemotherapy and radiation. In addition, early identification of resistance would allow for new precision medicine-based strategies to block those mechanisms and improve patient outcomes. We recently identified a novel mechanism of inflammation-mediated resistance to immune checkpoint inhibitors. Tumor cell production of the inflammatory cytokine IL-1a drives a suppressive circuit, resulting in the downstream production of G-CSF and CXCL1, which expand and recruit immunosuppressive neutrophils to the tumor microenvironment. In these tumors, immune checkpoint inhibitors do increase CD8 T cell infiltration in the tumor, but these CD8 T cells have poor cytotoxic function. Paradoxically, TNFa produced by T cells potentiates the resistance circuit by driving the production of higher levels of G-CSF and CXCL1, resulting in more neutrophil infiltration into the tumor. Blocking key elements of this circuit can synergize with checkpoint inhibitors to sensitize otherwise resistant tumors. Non-invasive imaging strategies to identify this mechanism of resistance early after therapy would allow patients to receive these combinations rapidly without waiting for evidence of disease progression by a radiographic scan. In this proposal, we will continue developing and will test such imaging strategies, including PET to measure granzyme B, which contributes to response, and neutrophil levels in the tumor, which contribute to resistance, early after treatment. We will also test novel implantable fluorescence sensors capable of measuring these parameters and reporting out their levels continuously in vivo. We will pair these molecular imaging approaches with high-parameter multiplexed ion beam imaging (MIBI) to measure up to 50 proteins simultaneously at subcellular resolution in tissue sections. We will utilize these techniques in established mouse models of response and resistance (Aim 1) and in a mouse model that exhibits variable responses (Aim 2). We will then determine whether early measurements after therapy enable rapid treatment modification for multiple tumor models with differing mechanisms of resistance, establishing imaging-based precision medicine for immunotherapy (Aim 3). These studies will collectively elucidate a novel mechanism of inflammation-mediated resistance to checkpoint inhibitors while developing molecular imaging approaches to measure these in real time early after treatment initiation. This study is in direct response to PAR-21-294, Molecular Imaging of Inflammation in Cancer, and will achieve the high-priority goals established by the NCI.