Systems Imaging of the Immune Synapse

Three-dimensional live cell imaging of the interaction of T cells with antigen presenting cells (APC) visualizes the subcellular distributions of signaling intermediates during T cell activation at thousands of resolved positions within a cell. These information-rich maps of local protein concentrations are a valuable resource in understanding T cell signaling. Here, we describe a protocol for the efficient acquisition of such imaging data and their computational processing to create four-dimensional maps of local concentrations. This protocol allows quantitative analysis of T cell signaling as it occurs inside live cells with resolution in time and space across thousands of cells

Efficacy of anti-CD147 chimeric antigen receptors targeting hepatocellular carcinoma

Chimeric antigen receptor (CAR) therapy is a promising immunotherapeutic strategy for treating multiple refractory blood cancers, but further advances are required for solid tumor CAR therapy. One challenge is identifying a safe and effective tumor antigen. Here, we devise a strategy for targeting hepatocellular carcinoma (HCC, one of the deadliest malignancies). We report that T and NK cells transduced with a CAR that recognizes the surface marker, CD147, also known as Basigin, can effectively kill various malignant HCC cell lines in vitro, and HCC tumors in xenograft and patient-derived xenograft mouse models. To minimize any on-target/off-tumor toxicity, we use logic-gated (log) GPC3–synNotch-inducible CD147-CAR to target HCC. LogCD147-CAR selectively kills dual antigen (GPC3+CD147+), but not single antigen (GPC3-CD147+) positive HCC cells and does not cause severe on-target/off-tumor toxicity in a human CD147 transgenic mouse model. In conclusion, these findings support the therapeutic potential of CD147-CAR-modified immune cells for HCC patients

Cytoskeletal control of B cell responses to antigens.

The actin cytoskeleton is essential for cell mechanics and has increasingly been implicated in the regulation of cell signalling. In B cells, the actin cytoskeleton is extensively coupled to B cell receptor (BCR) signalling pathways, and defects of the actin cytoskeleton can either promote or suppress B cell activation. Recent insights from studies using single-cell imaging and biophysical techniques suggest that actin orchestrates BCR signalling at the plasma membrane through effects on protein diffusion and that it regulates antigen discrimination through the biomechanics of immune synapses. These mechanical functions also have a role in the adaptation of B cell subsets to specialized tasks during antibody responses

Is autoimmunity the Achilles' heel of cancer immunotherapy?

The emergence of immuno-oncology as the first broadly successful strategy for metastatic cancer will require clinicians to integrate this new pillar of medicine with chemotherapy, radiation, and targeted small-molecule compounds. Of equal importance is gaining an understanding of the limitations and toxicities of immunotherapy. Immunotherapy was initially perceived to be a relatively less toxic approach to cancer treatment than other available therapies-and surely it is, when compared to those. However, as the use of immunotherapy becomes more common, especially as first- and second-line treatments, immunotoxicity and autoimmunity are emerging as the Achilles' heel of immunotherapy. In this Perspective, we discuss evidence that the occurrence of immunotoxicity bodes well for the patient, and describe mechanisms that might be related to the induction of autoimmunity. We then explore approaches to limit immunotoxicity, and discuss the future directions of research and reporting that are needed to diminish it