Tolerance to self-antigens prevents the elimination of cancer by the immune system1,2. We used synthetic chimeric antigen receptors (CARs) to overcome immunological tolerance and mediate tumor rejection in patients with chronic lymphocytic leukemia (CLL). Remission was induced in a subset of subjects, but most did not respond. Comprehensive assessment of patient-derived CAR T cells to identify mechanisms of therapeutic success and failure has not been explored. We performed genomic, phenotypic and functional evaluations to identify determinants of response. Transcriptomic profiling revealed that CAR T cells from complete-responding patients with CLL were enriched in memory-related genes, including IL-6/STAT3 signatures, whereas T cells from nonresponders upregulated programs involved in effector differentiation, glycolysis, exhaustion and apoptosis. Sustained remission was associated with an elevated frequency of CD27+CD45RO- CD8+ T cells before CAR T cell generation, and these lymphocytes possessed memory-like characteristics. Highly functional CAR T cells from patients produced STAT3-related cytokines, and serum IL-6 correlated with CAR T cell expansion. IL-6/STAT3 blockade diminished CAR T cell proliferation. Furthermore, a mechanistically relevant population of CD27+PD-1CD8+ CAR T cells expressing high levels of the IL-6 receptor predicts therapeutic response and is responsible for tumor control. These findings uncover new features of CAR T cell biology and underscore the potential of using pretreatment biomarkers of response to advance immunotherapies.

T he prototypical vaccine used in postexposure settings was developed over a century ago for use following exposure to a potentially rabid animal (9, 14). Postexposure vaccination remains the worldwide standard for the prevention of rabies infections of humans. Despite the long history of rabies vaccine use as a postexposure treatment and the facts that over two-thirds of the world population live in regions where rabies is endemic and that over 55,000 people die every year due to RV infections, little information is available regarding the development of effective B cell responses early postvaccination, which may influence the outcome of postexposure vaccine efficacy. Current vaccines for human use are based on inactivated RV strains. Therefore, vaccineinduced protection against RV has long been described as being dependent solely on CD4 ϩ T cell help for the induction of protective antibody responses (9). These observations are supported by experiments carried out in RV-vaccinated nude mice that did not develop anti-RV antibody responses and were not protected against pathogenic challenge in preexposure settings (29). Similarly, Mifune et al. (16) showed that vaccinated athymic nude mice did not produce anti-RV antibodies in postexposure experiments. Furthermore, neutralizing antibodies directed against RV antigens were shown to be produced only with the assistance of Thelper cells (1). Finally, T cell depletion studies further showed the importance of CD4 ϩ T cells in generating neutralizing, protective antibodies in mice after RV infection (21). Together, these experiments showed that current inactivated RV-based vaccines rely on T cell help for the elicitation of effective antibody responses associated with protection against pathogenic challenge.Vaccine-induced immunity is a complex process involving innate and adaptive immune responses. During the development of typical vaccine-induced immunity, antigen-primed T cells migrate to the T and B cell borders of secondary lymphoid organs, where they interact with their cognate antigen-specific B cells. After activation, B cells differentiate into early, short-lived extrafollicular plasma cells, germinal center (GC) B cells, or early unswitched memory B cells that can recirculate (7). Within GCs, B cells differentiate into memory B cells or long-lived plasma cells that secrete high-affinity, postswitched antibodies. In most cases, fully formed GC-derived memory B cells and plasma cells can take from days to weeks to develop. From a traditional vaccination standpoint, this lag time between vaccination and GC B cell development is required and usually acceptable, since GC B cells are critical for long-term B cell responses to protect against future exposure to the pathogen. However, vaccines are also used in postexposure settings after exposure to pathogens, such as for RV. Since the lag time between vaccination and the generation of effective GC B cell responses might be too long, administration of postexposure vaccines for the prevention of rabies infection may...

Over two-thirds of the world's population lives in regions where rabies is endemic, resulting in over 15 million people receiving multi-dose post-exposure prophylaxis (PEP) and over 55,000 deaths per year globally. A major goal in rabies virus (RABV) research is to develop a single-dose PEP that would simplify vaccination protocols, reduce costs associated with RABV prevention, and save lives. Protection against RABV infections requires virus neutralizing antibodies; however, factors influencing the development of protective RABV-specific B cell responses remain to be elucidated. Here we used a mouse model of IL-21 receptor-deficiency (IL-21R−/−) to characterize the role for IL-21 in RABV vaccine-induced immunity. IL-21R−/− mice immunized with a low dose of a live recombinant RABV-based vaccine (rRABV) produced only low levels of primary or secondary anti-RABV antibody response while wild-type mice developed potent anti-RABV antibodies. Furthermore, IL-21R−/− mice immunized with low-dose rRABV were only minimally protected against pathogenic RABV challenge, while all wild-type mice survived challenge, indicating that IL-21R signaling is required for antibody production in response to low-dose RABV-based vaccination. IL-21R−/− mice immunized with a higher dose of vaccine produced suboptimal anti-RABV primary antibody responses, but showed potent secondary antibodies and protection similar to wild-type mice upon challenge with pathogenic RABV, indicating that IL-21 is dispensable for secondary antibody responses to live RABV-based vaccines when a primary response develops. Furthermore, we show that IL-21 is dispensable for the generation of Tfh cells and memory B cells in the draining lymph nodes of immunized mice but is required for the detection of optimal GC B cells or plasma cells in the lymph node or bone marrow, respectively, in a vaccine dose-dependent manner. Collectively, our preliminary data show that IL-21 is critical for the development of optimal vaccine-induced primary but not secondary antibody responses against RABV infections.

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