Interleukin-1 alpha and -1 beta (IL-1α and IL-1β) are cytokines that participate in the regulation of immune responses, inflammatory reactions, and hematopoiesis. A direct expression strategy was used to clone the receptor for IL-1 from mouse T cells. The product of the cloned complementary DNA binds both IL-1α and IL-1β in a manner indistinguishable from that of the native T cell IL-1 receptor. The extracellular, IL-1 binding portion of the receptor is 319 amino acids in length and is composed of three immunoglobulin-like domains. The cytoplasmic portion of the receptor is 217 amino acids long.
Abstract Introduction: Sipuleucel-T is an autologous cellular immunotherapy FDA approved for men with asymptomatic or minimally symptomatic metastatic castrate resistant prostate cancer (mCRPC). Previous clinical data have shown that antibodies specific for the immunizing antigen, PA2024, develop within two weeks post-end of sipuleucel-T treatment. Here, we sought to determine whether B lymphocytes were activated during the course of treatment and to evaluate their phenotypic profile in two ongoing clinical studies (NEoACT and PRoACT). Materials and Methods: Sipuleucel-T is manufactured from peripheral blood mononuclear cells (PBMCs) isolated every two weeks by leukapheresis. The PBMCs are cultured with PA2024, an antigen composed of prostatic acid phosphatase (PAP) fused to granulocyte macrophage-colony stimulating factor (GM-CSF), washed, and then infused into the patient as sipuleucel-T. Each patient receives three infusions. The activation status of B lymphocytes was evaluated by multi-color flow cytometry during the manufacture of sipuleucel-T, before and after culture at each of the three treatment times. B lymphocyte subsets were identified by their expression of CD20, IgD and CD27: CD20+IgD+CD27− naive, CD20+IgD+CD27+ mature activated, and CD20+IgD−CD27+ memory. Activation of each of these subsets was determined by their expression of CD80 and CD96. Antigen-specific B lymphocyte function, in the form of antibody production, was evaluated in the serum of sipuleucel-T-treated subjects before and after treatment. Summary: In the mature activated B lymphocyte subset, an increase in the proportion of lymphocytes expressing CD80+ and CD86+ was detected following culture with PA2024, with the increase in CD86+ mature B lymphocytes being the highest after the second infusion of sipuleucel-T. An increase in mature activated B lymphocytes directly ex vivo, pre-culture, was not observed prior to the second or third infusion. Additionally, CD80 and CD86 expression was upregulated on memory B lymphocytes in response to culture with PA2024 and following the first infusion. Furthermore, PA2024- and PAP-specific antibody responses were detected and maintained 2-22 weeks after the completion of sipuleucel-T treatment.Conclusions: These data demonstrate that B lymphocyte activation is a consequence of ex vivo culture with PA2024 and precedes an antigen-specific humoral response, as detected in the patients’ serum after treatment with sipuleucel-T. Collectively, these findings provide evidence of therapy-induced humoral immune activation in men with mCRPC treated with sipuleucel-T. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5508. doi:10.1158/1538-7445.AM2011-5508
The precise molecular characteristics and the mode of action of the T cell derived lymphokines which augment antibody production in vitro remain uncertain. The use of ill-defined culture supernatants to dissect the cellular interactions in vitro involved in antibody production can lead to ambiguous results as the factors may act either on a contaminating non-B-lymphoid population or directly on the B lymphocyte. We report herein the development of a system for measuring in vitro primary antibody responses by murine spleen cells in which endogenous lymphokine production has been minimized by the in vivo administration of cytotoxic antibodies to deplete T lymphocytes and the addition of the glucocorticosteroid, dexamethasone, throughout the culture period. Using such an assay, a lymphokine activity was detected which was capable of augmenting the plaque forming cell response. This lymphokine was present in culture supernatant derived from the lectin activation of the T cell lymphoma, LBRM-33 and was distinct from other known B cell activators, notably IL-2 and IFN gamma. Biochemical purification of this activity indicated that it might be identical to granulocyte-macrophage colony stimulating factor (GM-CSF). The use of recombinant-derived GM-CSF protein unambiguously showed the role of this lymphokine in antibody production. These experiments demonstrated for the first time, the involvement of a hematopoietic factor in antigen-specific immune responses. Moreover, these results demonstrated an important regulatory circuit in the generation of antibody producing B cells in which GM-CSF, derived from activated T cells, stimulates macrophage function.
A method utilizing reversed-phase high-performance liquid chromatography has been developed for the purification to homogeneity of interleukin 2 (IL-2) isolated from a human T-cell leukemia. A final purification of 500,000-fold was obtained with a specific activity of pure IL-2 of 10(9) units/mg. The amino acid analysis of natural IL-2 is strikingly similar to the composition deduced from sequence analysis of a cDNA coding for human IL-2. Protein sequence analysis of CNBr-derived peptides yields data consistent with the sequence proposed from cloned cDNA. The availability of homogeneous IL-2 will allow accurate biological studies of its activity free from the contamination of the numerous lymphokine species that are known to be co-produced with IL-2 during the induction procedure.
