A destructive feedback loop mediated by CXCL10 in central nervous system inflammatory disease.

OBJECTIVE: Paraneoplastic neurologic disorders (PND) are autoimmune diseases associated with cancer and ectopic expression of a neuronal antigen in a peripheral tumor. Patients with PND harbor high-titer antibodies and T cells in their serum and cerebrospinal fluid (CSF) that are specific to the tumor antigen, and treatment with the immunosuppressant FK506 (tacrolimus) decreases CSF white blood cell counts. The objective of this study was to determine the effect of FK506 on CSF chemokine levels in PND patients. METHODS: CSF samples before and after FK506 treatment were tested by multiplex assay for the presence of 27 cytokines. Follow-up in vitro experiments aimed to determine whether T cells secrete CXCL10 in response to cognate antigen. RESULTS: Here we report that PND patients harbor high levels of the chemokine CXCL10 in their CSF. CXCL10 is a cytokine that recruits CXCR3(+) cells such as activated T cells, and we found that FK506 treatment specifically decreased CSF CXCL10 from among 27 cytokines tested. In vitro, CXCL10 was only produced during antigen-specific cognate interactions between T cells and antigen-presenting cells (APCs) when interferon-γ (IFNγ) receptors were present on the T cell. INTERPRETATION: These results support a model in which antigen-specific T cell stimulation by PND APCs triggers IFNγ, followed by CXCL10 production and further lymphocyte recruitment, suggesting that treatments targeting T cells or CXCL10 in the central nervous system (CNS) may interrupt a destructive positive feedback loop present in CNS inflammation.

A T-cell receptor associated with naturally occurring human tumor immunity.

The onconeural antigens appear to serve as tumor rejection antigens in the paraneoplastic neurologic disorders. Here, we used an unbiased peptide binding screen, followed by studies in HLA-A2.1 transgenic mice to identify naturally processed HLA-A2.1 restricted epitopes of the paraneoplastic cerebellar degeneration breast/ovarian cancer antigen cdr2. These mice were used to clone high-avidity cdr2-specific CD8(+) T cells that recognize human tumor cells presenting endogenously loaded MHC class I-cdr2 peptide. T cells with this specificity were detected in the peripheral blood of two HLA-A2.1(+) paraneoplastic cerebellar degeneration patients. We cloned T cell receptor (TCR) alpha and beta genes from cdr2-specific T cells; electroporation of RNA encoding this TCR turned nonreactive donor T cells into efficient killers of human cdr2-expressing tumor cells. Cloned cdr2-specific TCR genes provide a clinically relevant means for immunologic targeting of human gynecologic cancers.

Neuroimmunology of the paraneoplastic neurological degenerations.

The paraneoplastic neurological degenerations (PNDs) are remarkable examples of naturally occurring tumor immunity in humans. In PND patients, common tumors such as breast, ovarian and lung tumors express proteins normally made exclusively in the brain, eliciting an immune response that successfully suppresses growth of the tumor. This successful anti-tumor response would be expected to go clinically unnoticed by the patient, but the immune cells mediating the response somehow cross into the brain, resulting in an autoimmune attack on neurons and neurological symptoms. Pieces of the mystery of this tumor immune response and neuronal autoimmunity have been assembled, but much more needs to be learned.

CD8+ T-cell-dependent immunity following xenogeneic DNA immunization against CD20 in a tumor challenge model of B-cell lymphoma.

The CD20 B-cell differentiation antigen is an attractive target for immunotherapy of B-cell lymphomas. In an experimental lymphoma model, BALB/c mice were immunized with mouse or human CD20 cDNA (mCD20 and hCD20, respectively) or their extracellular domains (minigenes). IFNgamma secretion by CD8+ T cells against CD20 was detected in mice vaccinated with hCD20 or human minigene, indicating that hCD20-primed CD8+ T cells recognize syngeneic CD20. Systemic challenge with syngeneic A20 cells, an aggressive lymphoma, resulted in long-term survival in a subset of immunized mice. Overall long-term survival was 14% in groups vaccinated with the human minigene versus 4% in control groups (P < 0.001). CD8+ T-cell depletion during the effector phase completely abrogated this effect. Antibodies against a recombinant mouse CD20 protein produced in insect cells were detected in mice immunized with hCD20 DNA and human and mouse minigene, but not in mice receiving mCD20 DNA. These results show that active immunization with xenogeneic DNA vaccines can induce CD8+ T cell-dependent immunity against CD20.

Patients with lung cancer and paraneoplastic Hu syndrome harbor HuD-specific type 2 CD8+ T cells.

Paraneoplastic neurologic disorders (PNDs) offer an uncommon opportunity to study human tumor immunity and autoimmunity. In small cell lung cancer (SCLC), expression of the HuD neuronal antigen is thought to lead to immune recognition, suppression of tumor growth, and, in a subset of patients, triggering of the Hu paraneoplastic neurologic syndrome. Antigen-specific CTLs believed to contribute to disease pathophysiology were described 10 years ago in paraneoplastic cerebellar degeneration. Despite parallel efforts, similar cells have not been defined in Hu patients. Here, we have identified HuD-specific T cells in Hu patients and provided an explanation for why their detection has been elusive. Different Hu patients harbored 1 of 2 kinds of HuD-specific CD8+ T cells: classical IFN-gamma-producing CTLs or unusual T cells that produced type 2 cytokines, most prominently IL-13 and IL-5, and lacked cytolytic activity. Further, we found evidence that SCLC tumor cells produced type 2 cytokines and that these cytokines trigger naive CD8+ T cells to adopt the atypical type 2 phenotype. These observations demonstrate the presence of an unusual noncytotoxic CD8+ T cell in patients with the Hu paraneoplastic syndrome and suggest that SCLC may evade tumor immune surveillance by skewing tumor antigen-specific T cells to this unusual noncytolytic phenotype.

Vaccination with CD20 peptides induces a biologically active, specific immune response in mice.

CD20 is a 33-kD B-cell antigen that is expressed from the early pre-B-cell stage of development and is lost on differentiation of B cells into plasma cells. Because CD20 is expressed strictly by B cells, it is an attractive target for B-cell lymphoma therapy. Monoclonal antibodies to CD20 have been used successfully in the treatment of B-cell lymphomas. We hypothesized that a vaccine consisting of CD20 peptide sequences might be capable of inducing an active, specific, humoral immune response to the protein. Vaccine therapy would have the advantage of generating a polyclonal response to the antigen in contrast to the monoclonal response of an infused antibody. Balb/c mice were vaccinated with prototype vaccine constructs that consisted of peptides representing the human or mouse CD20 extracellular sequences conjugated to carrier proteins and mixed with QS21 adjuvant. Sera from the vaccinated mice demonstrated high-titer, specific antibodies to various epitopes on the immunizing peptides in enzyme-linked immunosorbent assay, weaker antibody binding to native CD20 on cells by flow cytometry, and antibody-mediated complement killing of CD20(+) cells in some cases. Specific proliferation and secretion of interleukin 4 and interferon gamma by mouse spleen cells in response to the immunizing peptides were also demonstrated. Mice vaccinated with the CD20 peptide keyhole limpet hemocyanin conjugates had a 25% decrease in CD19(+) splenic B cells relative to control mice. These data indicate that a biologically active, specific immune response to CD20 can be elicited in mice vaccinated with CD20 peptide conjugates.