Antigen specific T cell migration to sites of infection or cancer is critical for an effective immune response. In mouse models of cancer, the number of lymphocytes reaching the tumor is typically only a few hundred, yet technology capable of imaging these cells using bioluminescence has yet to be achieved. A combination of codon optimization, removal of cryptic splice sites and retroviral modification was used to engineer an enhanced firefly luciferase (ffLuc) vector. Compared with ffLuc, T cells expressing our construct generated >100 times more light, permitting detection of as few as three cells implanted s.c. while maintaining long term coexpression of a reporter gene (Thy1.1). Expression of enhanced ffLuc in mouse T cells permitted the tracking of <3 ؋ 10 4 adoptively transferred T cells infiltrating sites of vaccination and preestablished tumors. Penetration of light through deep tissues, including the liver and spleen, was also observed. Finally, we were able to enumerate infiltrating mouse lymphocytes constituting <0.3% of total tumor cellularity, representing a significant improvement over standard methods of quantitation including flow cytometry.bioluminescence ͉ immunology ͉ molecular biology

Background: Bahia grass pollen (BaGP) is a major cause of allergic rhinitis. Subcutaneous allergen-specific immunotherapy is effective for grass pollen allergy, but is unsuitable for patients with moderate to severe asthma due to the risk of anaphylaxis. T cell-reactive but IgE nonreactive peptides provide a safer treatment option. This study aimed to identify and characterize dominant CD4⁺ T cell epitope peptides of the major BaGP allergen, Pas n 1. Methods: Pas n 1-specific T cell lines generated from the peripheral blood of BaGP-allergic subjects were tested for proliferative and cytokine response to overlapping 20-mer Pas n 1 peptides. Cross-reactivity to homologous peptides from Lol p 1 and Cyn d 1 of Ryegrass and Bermuda grass pollen, respectively, was assessed using Pas n 1 peptide-specific T cell clones. MHC class II restriction of Pas n 1 peptide T cell recognition was determined by HLA blocking assays and peptide IgE reactivity tested by dot blotting. Results: Three Pas n 1 peptides showed dominant T cell reactivity; 15 of 18 (83%) patients responded to one or more of these peptides. T cell clones specific for dominant Pas n 1 peptides showed evidence of species-specific T cell reactivity as well as cross-reactivity with other group 1 grass pollen allergens. The dominant Pas n 1 T cell epitope peptides showed HLA binding diversity and were non-IgE reactive. Conclusions: The immunodominant T cell-reactive Pas n 1 peptides are candidates for safe immunotherapy for individuals, including those with asthma, who are allergic to Bahia and possibly other grass pollens.

The introduction of targeted agents has substantially improved treatment of metastatic clear-cell renal cell carcinoma (RCC). However, complete responses are rare and therapy is not curative. Moreover, information on the latest generation of potent and selective vascular endothelial growth factor receptor (VEGFR) tyrosine kinase inhibitors (TKI) suggests that a plateau has been reached in terms of efficacy. Recent data reveal that targeted agents are involved in modulating immune responses in RCC. In addition, current research adds to our understanding of how RCC escapes an effective anti-tumor response with the potential to modulate these processes by drug development. This review provides specific insight into targeted therapy induced changes in the immunological microenvironment of RCC, summarizes the available evidence, and discusses potential therapeutic implications.

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Adoptive cell transfer of expanded, autologous tumor-infiltrating lymphocytes (TIL) into lymphodepleted melanoma patients can induce the regression of bulky, metastatic disease. To generate the large numbers of T cells needed for infusion, TIL undergo a rapid expansion protocol (REP) in vitro using anti-CD3 antibody, interleukin-2, and irradiated peripheral blood feeder cells that typically results in an approximately 1000-fold expansion over 14 days. However, we have found that the conventional REP (C-REP) often favors the expansion of CD4+ T cells at the expense of tumor antigen-specific CD8+ T cells, which are the most potent cytolytic effector cells. In this study, we demonstrate that addition of transforming growth factor (TGF)-beta1 to the TIL culture at the onset of rapid expansion (T-REP ) maintained the percentage of CD8+ T cells while not inhibiting overall T-cell expansion. Of T cells expanded from different melanoma patient tumors, 13 of 15 TIL demonstrated improved yields and percentages of both CD8+ and MART-1 melanoma antigen-specific T cells after 14 days of expansion in TGF-beta1 compared with the C-REP. This was associated with a marked improvement in the antitumor activity of the resulting bulk TIL culture in terms of interferon-gamma production and melanoma tumor-specific cytotoxic T-lymphocyte activity. In addition, T-REP T cells demonstrated a higher potential for continued expansion in vitro for up to 3 weeks after the expansion compared with C-REP T cells, suggesting that they may also be capable of increased persistence after adoptive cell transfer. Our results suggest that TGF-beta1-expanded TIL have attributes that might predict efficacy superior to that of conventional TIL.

In the human leucocyte antigen (HLA)-matched haematopoietic stem cell transplantation (HSCT) setting, minor histocompatibility antigen (mHA) disparities between recipient and donor can lead to graft-vs-host disease (GVHD) or graft rejection. Graft-vs-leukaemia (GVL) effect is a beneficial T-cell-mediated immune response that can also occur following HLA-matched HSCT. mHAs with tissue expression restricted to cells of the haematopoietic system are particularly relevant as immunotherapeutic targets for destroying malignant cells without inducing GVHD. Therefore, it is important to identify further haematopoietic-restricted polymorphic mHAs, which may have the potential to be used clinically for adoptive immunotherapy. Polymorphic mismatching of minor antigens, such as the B-cell-specific protein, the kappa immunoglobulin light chain (kappa) may play a role in the incidence of GVL and therefore the survival of transplant recipients following transplantation for B-cell malignancies. Polymorphisms in the constant region of the immunoglobulin kappa polypeptide chain have been defined involving single amino acid changes at positions 153 and 191. In this study, 51 HLA-matched B-cell malignancy transplant pairs were kappa typed by polymerase chain reaction and restriction enzyme digestion to investigate the association between kappa allotype disparity and outcome after transplantation. Kappa allotype disparity between transplant pairs may be associated with an increased survival compared with pairs not mismatched for kappa, as kappa mismatched recipients had a higher percentage of complete remissions and a decreased level of relapse in comparison with the nonmismatched recipients. HLA peptide prediction software was used to determine which HLA types were the best binders for kappa peptides. It was observed that patients with tissue types predicted to bind the kappa Km(1,2) peptides had better survival outcomes and no relapse compared with those with tissue types not predicted to bind the kappa Km(1,2) peptides. This study may contribute to the assessment of the clinical role of kappa with regard to the outcome of allogeneic transplantation for B-cell malignancies.

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