Tunable control of CAR T cell activity through tetracycline mediated disruption of protein-protein interaction.

Chimeric antigen receptor (CAR) T cells are a promising form of cancer immunotherapy, although they are often associated with severe toxicities. Here, we present a split-CAR design incorporating separate antigen recognition and intracellular signaling domains. These exploit the binding between the tetracycline repressor protein and a small peptide sequence (TIP) to spontaneously assemble as a functional CAR. Addition of the FDA-approved, small molecule antibiotic minocycline, acts as an "off-switch" by displacing the signaling domain and down-tuning CAR T activity. Here we describe the optimization of this split-CAR approach to generate a CAR in which cytotoxicity, cytokine secretion and proliferation can be inhibited in a dose-dependent and reversible manner. Inhibition is effective during on-going CAR T cell activation and inhibits activation and tumor control in vivo. This work shows how optimization of split-CAR structure affects function and adds a novel design allowing easy CAR inhibition through an FDA-approved small molecule.

Intratumoral IL-12 delivery empowers CAR-T cell immunotherapy in a pre-clinical model of glioblastoma.

Glioblastoma multiforme (GBM) is the most common and aggressive form of primary brain cancer, for which effective therapies are urgently needed. Chimeric antigen receptor (CAR)-based immunotherapy represents a promising therapeutic approach, but it is often impeded by highly immunosuppressive tumor microenvironments (TME). Here, in an immunocompetent, orthotopic GBM mouse model, we show that CAR-T cells targeting tumor-specific epidermal growth factor receptor variant III (EGFRvIII) alone fail to control fully established tumors but, when combined with a single, locally delivered dose of IL-12, achieve durable anti-tumor responses. IL-12 not only boosts cytotoxicity of CAR-T cells, but also reshapes the TME, driving increased infiltration of proinflammatory CD4+ T cells, decreased numbers of regulatory T cells (Treg), and activation of the myeloid compartment. Importantly, the immunotherapy-enabling benefits of IL-12 are achieved with minimal systemic effects. Our findings thus show that local delivery of IL-12 may be an effective adjuvant for CAR-T cell therapy for GBM.

In vivo safety profile of a CSPG4-directed IgE antibody in an immunocompetent rat model.

IgE monoclonal antibodies hold great potential for cancer therapy. Preclinical in vivo systems, particularly those in which the antibody recognizes the host species target antigen and binds to cognate Fc receptors, are often the closest approximation to human exposure and represent a key challenge for evaluating the safety of antibody-based therapies. We sought to develop an immunocompetent rat system to assess the safety of a rodent anti-tumor IgE, as a surrogate for the human therapeutic candidate. We generated a rat IgE against the human tumor-associated antigen chondroitin sulfate proteoglycan 4 (CSPG4) and cross-reactive for the rat antigen. We analyzed CSPG4 distribution in normal rat and human tissues and investigated the in vivo safety of the antibody by monitoring clinical signs and molecular biomarkers after systemic administration to immunocompetent rats. Human and rat CSPG4 expression in normal tissues were comparable. Animals receiving antibody exhibited transient mild to moderate adverse events accompanied by mild elevation of serum tryptase, but not of angiotensin II or cytokines implicated in allergic reactions or cytokine storm. In the long term, repeated antibody administration was well tolerated, with no changes in animal body weight, liver and kidney functions or blood cell counts. This model provides preclinical support for the safety profiling of IgE therapeutic antibodies. Due to the comparable antigen tissue distribution in human and rat, this model may also comprise an appropriate tool for proof-of-concept safety evaluations of different treatment approaches targeting CSPG4.

Combining Immune Checkpoint Inhibitors: Established and Emerging Targets and Strategies to Improve Outcomes in Melanoma.

The immune system employs several checkpoint pathways to regulate responses, maintain homeostasis and prevent self-reactivity and autoimmunity. Tumor cells can hijack these protective mechanisms to enable immune escape, cancer survival and proliferation. Blocking antibodies, designed to interfere with checkpoint molecules CTLA-4 and PD-1/PD-L1 and counteract these immune suppressive mechanisms, have shown significant success in promoting immune responses against cancer and can result in tumor regression in many patients. While inhibitors to CTLA-4 and the PD-1/PD-L1 axis are well-established for the clinical management of melanoma, many patients do not respond or develop resistance to these interventions. Concerted efforts have focused on combinations of approved therapies aiming to further augment positive outcomes and survival. While CTLA-4 and PD-1 are the most-extensively researched targets, results from pre-clinical studies and clinical trials indicate that novel agents, specific for checkpoints such as A2AR, LAG-3, IDO and others, may further contribute to the improvement of patient outcomes, most likely in combinations with anti-CTLA-4 or anti-PD-1 blockade. This review discusses the rationale for, and results to date of, the development of inhibitory immune checkpoint blockade combination therapies in melanoma. The clinical potential of new pipeline therapeutics, and possible future therapy design and directions that hold promise to significantly improve clinical prognosis compared with monotherapy, are discussed.

Subsurface radar sounding of the south polar layered deposits of Mars.

The ice-rich south polar layered deposits of Mars were probed with the Mars Advanced Radar for Subsurface and Ionospheric Sounding on the Mars Express orbiter. The radar signals penetrate deep into the deposits (more than 3.7 kilometers). For most of the area, a reflection is detected at a time delay that is consistent with an interface between the deposits and the substrate. The reflected power from this interface indicates minimal attenuation of the signal, suggesting a composition of nearly pure water ice. Maps were generated of the topography of the basal interface and the thickness of the layered deposits. A set of buried depressions is seen within 300 kilometers of the pole. The thickness map shows an asymmetric distribution of the deposits and regions of anomalous thickness. The total volume is estimated to be 1.6 x 10(6) cubic kilometers, which is equivalent to a global water layer approximately 11 meters thick.