Soluble HLA/peptide monomers cross-linked with co-stimulatory antibodies onto a streptavidin core molecule efficiently stimulate antigen-specific T cell responses.

Soluble MHC-peptide complexes, commonly referred to as tetramers, have been shown to induce strong cross-linking of TCR and CD8, resulting in a vigorous activation followed by a rapid non-apoptotic CD8(+) T cell death. This has limited tetramer use for antigen-specific T cells isolation and cloning, as sorted tetramer positive cells were shown to possess compromised functional integrity. Here we show that the cross-linking of a secondary co-stimulatory signal into oligomeric MHC:peptide complexes prevents such cell death, and in contrast strongly stimulates antigen-specific T cell responses. Such soluble antigen-presenting complexes (sAPCs) containing MHC:peptide complexes linked to either anti-CD27 or anti-CD28 antibodies were capable of priming and expanding HLA-A*0201 restricted CMV specific T cells and also of generating functional HLA-A*0301 restricted BCR/ABL-specific T cell responses. These sAPCs constitute an encouraging alternative method for generating antigen-specific T cells that could be applied to a variety of antigens.

BCR/ABL-specific CD8+ T cells can be detected from CML patients, but are only expanded from healthy donors.

The BCR/ABL p210 fusion protein has long been considered an ideal target antigen for the development of immunotherapeutic strategies in chronic myeloid leukaemia (CML) due to its central role in malignant transformation and to its unique novel amino acid sequence solely expressed in leukaemia cells. However, the feasibility to expand BCR-ABL-specific T cells remains still controversial. Using BCR/ABL peptide/MHC tetramers, significantly higher frequencies of tetramer positive cells were detected in the peripheral blood of HLA-A*0301 (mean 0.38%) and HLA-B*0801 (mean 0.28%) CML patients than in healthy donors (P = 0.0025 and 0.0026, respectively). However, following stimulation with autologous peptide-pulsed DCs, BCR/ABL-specific T cells were only expanded from some healthy donors, suggesting that CML patients may have a specific immune deficit with respect to the BCR/ABL antigen.

Differential pathways govern CD4+ CD28- T cell proinflammatory and effector responses in patients with coronary artery disease.

Patients with acute coronary syndromes experience circulatory and intraplaque expansion of an aggressive and unusual CD4(+) lymphocyte subpopulation lacking the CD28 receptor. These CD4(+)CD28(-) cells produce IFN-gamma and perforin, and are thought to play an important role in coronary atheromatous plaque destabilization. Aberrant expression of killer Ig-like receptors (KIRs) in CD4(+)CD28(-) cells is broadly thought to be responsible for their cytotoxicity, but the mechanisms involved remain poorly defined. We therefore sought to investigate the mechanism and regulation of CD4(+)CD28(-) cell functionality using T cell clones (n = 536) established from patients with coronary artery disease (n = 12) and healthy volunteers (n = 3). Our functional studies demonstrated that KIR2DS2 specifically interacted with MHC class I-presenting human heat shock protein 60 (hHSP60) inducing cytotoxicity. Further investigations revealed the novel finding that hHSP60 stimulation of TCR alone could not induce a cytotoxic response, and that this response was specific and KIR dependent. Analysis of CD4(+)CD28(-)2DS2(+) clones (n = 162) showed that not all were hHSP60 cytotoxic; albeit, their prevalence correlated with coronary disease status (p = 0.017). A higher proportion of clones responded to hHSP60 by IFN-gamma compared with perforin (p = 0.008). In this study, for the first time, we define the differential regulatory pathways involved in CD4(+)CD28(-) cell proinflammatory and effector responses. We describe in this study that, contrary to previous reports, CD4(+)CD28(-) cell recognition and killing can be specific and discriminate. These results, in addition to contributing to the understanding of CD4(+)CD28(-) cell functionality, may have implications for the monitoring and management of coronary artery disease progression.

Large numbers of dysfunctional CD8+ T lymphocytes bearing receptors for a single dominant CMV epitope in the very old.

Longitudinal studies suggest that a set of immune parameters including high percentages of peripheral CD8+, CD28-, CD57+ T lymphocytes, low CD4 and B cell counts, and poor T cell proliferative responses to mitogens is associated with decreased remaining longevity in the free-living very elderly (> 85 years). This combination of immune parameters was also significantly associated with an inverted CD4/CD8 ratio and cytomegalovirus seropositivity. Here, using tetramer technology, we show markedly increased numbers of CD8+ T cells bearing receptors for one single CMV epitope in the very elderly. Moreover, the fraction of these tetramer-reactive cells secreting interferon-gamma after specific antigenic stimulation was significantly lower in the old than in the young, as was the percentage of CD28-positive cells in this population. Therefore, we conclude that marked expansions of CMV-specific CD8+ T cells have occurred and that the obsession of a large fraction of the entire CD8+ T cell subset with one single viral epitope may contribute to the increased incidence of infectious disease in the elderly by shrinking the T cell repertoire available for responses to other antigens.