Academic literature on the topic 'CD8-Positive T-Lymphocytes Immunotherapy'

Pneumocystispneumonia (PCP) is a major cause of morbidity and mortality in patients with HIV infection. CD4+T lymphocytes are critical for host defense against this infection, but in the absence of CD4+T lymphocytes, CD8+T lymphocytes may provide limited host defense. The cytokine interleukin-7 (IL-7) functions to enhance lymphocyte proliferation, survival, and recruitment of immune cells to sites of infection. However, there is little known about the role of IL-7 in PCP or its potential use as an immunotherapeutic agent. We hypothesized that treatment with recombinant human IL-7 (rhIL-7) would augment host defense againstPneumocystisand accelerate pathogen clearance in CD4-depleted mice. Control and CD4-depleted mice were infected withPneumocystis, and rhIL-7 was administered via intraperitoneal injection. Our studies indicate that endogenous murine IL-7 is part of the normal host response toPneumocystis murinaand that administration of rhIL-7 markedly enhanced clearance ofPneumocystisin CD4-depleted mice. Additionally, we observed increased recruitment of CD8+T lymphocytes to the lungs and decreased apoptosis of pulmonary CD8+T lymphocytes in rhIL-7-treated animals compared to those in untreated mice. The antiapoptotic effect of rhIL-7 was associated with increased levels of Bcl-2 protein in T lymphocytes. rhIL-7 immunotherapy in CD4-depleted mice also increased the number of gamma interferon (IFN-γ)-positive CD8+central memory T lymphocytes in the lungs. We conclude that rhIL-7 has a potent therapeutic effect in the treatment of murinePneumocystispneumonia in CD4-depleted mice. This therapeutic effect is mediated through enhanced recruitment of CD8+T cells and decreased apoptosis of lung T lymphocytes, with a preferential action on central memory CD8+T lymphocytes.

Abstract Cytolytic T lymphocytes play an important role in host defense against viral infections, including human immunodeficiency virus (HIV). In a phase I clinical trial (protocol 080 of the AIDS Clinical Trials Group), generation of CD8+ effector cells from peripheral blood of patients with acquired immunodeficiency syndrome (AIDS)-related complex (ARC) or AIDS and safety of autologous adoptive transfer of these cells were evaluated. For therapeutic infusions, CD8+ T cells were purified by positive selection on anti-CD8 monoclonal antibody-coated flasks from leukapheresed peripheral blood of seven patients. These CD8+ T cells were cultured in the presence of interleukin-2 and phytohemagglutinin for up to 3 weeks to obtain cells sufficient for therapeutic infusions (10(8) to 10(10)). All 31 cell cultures established from the seven patients and used for therapy were highly enriched in CD8+ (mean, 97%), CD8+HLA-DR+ (50%), cytotoxic CD8+CD11b- (82%), and memory CD29+ (78%) T lymphocytes. In vitro expanded CD8+ cells had excellent cytotoxic function at the time they were used for therapy, including HIV-specific activity against autologous targets infected with vaccinia vectors expressing HIV-IIIb antigens, gag, pol, and env. Anti-HIV activity of cultured CD8+ cells was significantly higher than that of autologous fresh peripheral blood lymphocytes. Our results show that CD8+ T lymphocytes obtained from peripheral blood of symptomatic HIV-infected patients can be purified, cultured to obtain large numbers of cells with enhanced anti-HIV activity, and safely infused into patients with AIDS as a form of immunotherapy.

Cytolytic T lymphocytes play an important role in host defense against viral infections, including human immunodeficiency virus (HIV). In a phase I clinical trial (protocol 080 of the AIDS Clinical Trials Group), generation of CD8+ effector cells from peripheral blood of patients with acquired immunodeficiency syndrome (AIDS)-related complex (ARC) or AIDS and safety of autologous adoptive transfer of these cells were evaluated. For therapeutic infusions, CD8+ T cells were purified by positive selection on anti-CD8 monoclonal antibody-coated flasks from leukapheresed peripheral blood of seven patients. These CD8+ T cells were cultured in the presence of interleukin-2 and phytohemagglutinin for up to 3 weeks to obtain cells sufficient for therapeutic infusions (10(8) to 10(10)). All 31 cell cultures established from the seven patients and used for therapy were highly enriched in CD8+ (mean, 97%), CD8+HLA-DR+ (50%), cytotoxic CD8+CD11b- (82%), and memory CD29+ (78%) T lymphocytes. In vitro expanded CD8+ cells had excellent cytotoxic function at the time they were used for therapy, including HIV-specific activity against autologous targets infected with vaccinia vectors expressing HIV-IIIb antigens, gag, pol, and env. Anti-HIV activity of cultured CD8+ cells was significantly higher than that of autologous fresh peripheral blood lymphocytes. Our results show that CD8+ T lymphocytes obtained from peripheral blood of symptomatic HIV-infected patients can be purified, cultured to obtain large numbers of cells with enhanced anti-HIV activity, and safely infused into patients with AIDS as a form of immunotherapy.

Abstract Abstract 2984 T lymphocytes play central roles in cellular immunity, exerting their proliferative and effector activities when they recognize antigens via T-cell receptors (TCRs) in HLA-restricted and antigen-specific manner. Adoptive cell transfer therapy (ACT), the administration of ex vivo-activated and -expanded autologous tumor-reactive T lymphocytes, is currently one of the effective methods for immunotherapy, especially for treatment of metastatic solid tumors including melanoma. However, the successful applications of this method are currently limited for tumor therapies. To broaden the range of the application of ACT, we endeavored to develop easier method to obtain cells that carry antigen-specific TCR genes. For the purpose, generation of induced pluripotent stem (iPS) cells from an antigen-reactive single T lymphocyte is attractive and rewarding way. iPS cells have a capacity for unlimited self-renewal while maintaining pluripotency. These features may enable us to induce an unlimited number of T lymphocytes, especially high proliferative naïve / central memory-type T lymphocytes, showing reactivity to specific antigens. If they retain properties of naïve T lymphocytes, they may proliferate for a longer period and achieve better therapeutic effects than their peripheral blood counterparts expanded in vitro. Peripheral T lymphocytes were isolated from healthy volunteers. Then reprogramming factors (OCT4, SOX2, KLF4, and c-MYC) were transduced into fresh or frozen / thawed T lymphocytes. T lymphocyte-derived iPS-like colonies were observed within 3 weeks and they were isolated and clonally expanded. They exhibited standard ES-like morphology, cell surface marker expression, alkaline phosphatase activity, as well as differentiation potential into various tissues related to all three germ layers. Human TCRs are encoded in four genes (TCRA, TCRB, TCRG, TCRD), which should be genetically assembled in an irreversible manner during T-lymphocyte development. This feature allowed us to retrospectively confirm that the iPS cells were generated from T lymphocyte. The TCR gene rearrangements encoded in an iPS colony were clonal for all iPS lines, indicating that each iPS colony was derived from a single T lymphocyte. Sequence analyses of TCR genes revealed whether the rearrangements were productive, and the productivity might promise the conservation of TCR genes rearrangement during the reprogramming process. Next, we tried to re-differentiate T lymphocyte derived-iPS (T-iPS) cells into T cells by co-culturing them with murine stromal cell layers (OP9 and OP9-DL1). T-cell differentiation was evidenced by the expression of T-cell markers, such as CD5, CD7, CD27, CD4, CD8, TCR α β and CD3. We obtained 33.5 ± 17.9% CD4+ CD8+ double positive (DP) cells, 6.51 ± 5.40% CD4+ CD8− single positive (SP) cells and 3.80 ± 1.28% CD4− CD8+ SP cells. They could be activated via TCR stimulation, and produce cytokines as functionally matured T lymphocytes do. The re-differentiation efficiency of T-iPS cells was higher than those of other pluripotent stem cells, such as embryonic stem (ES) cells, fibroblasts derived-iPS cells, or cord blood derived-iPS cells. Transcribed TCR mRNA sequences in re-differentiated T cells were analyzed, and they were revealed to be identical to that engraved in the pre-differentiated T-iPS cells genome in CD4+ CD8+ DP phase. However, fully matured CD4+ CD8− or CD4− CD8+ SP phase cells had several TCRA gene rearrangement patterns distinct from the original T-iPS cell's. On the other hand, TCRB gene maintained identity with the original. The variance of the sequences, especially antigen-recognition site sequences, indicated that the antigen-specificity in the original T lymphocyte might be converted during DP to SP transition process in vitro. These data indicate that functionally matured T cells were generated by re-differentiating T-iPS cells in vitro, and that re-assemble of TCRA genes could take place during SP T cell maturation process. In order to fulfill the T-iPS-mediated immunotherapy, we need to overcome the obstacle of further TCRA gene rearrangements. We think the solution lies in refinement of the re-differentiation method for controlling the expression of RAG1 and RAG2 recombinases or for inhibiting their activities. Disclosures: No relevant conflicts of interest to declare.

Current strategies for the immunotherapy of melanoma include augmentation of the immune response to tumor antigens represented by melanosomal proteins such as tyrosinase, gp100, and MART-1. The possibility that intentional targeting of tumor antigens representing normal proteins can result in autoimmune toxicity has been postulated but never demonstrated previously in humans. In this study, we describe a patient with metastatic melanoma who developed inflammatory lesions circumscribing pigmented areas of skin after an infusion of MART-1–specific CD8+ T cell clones. Analysis of the infiltrating lymphocytes in skin and tumor biopsies using T cell–specific peptide–major histocompatibility complex tetramers demonstrated a localized predominance of MART-1–specific CD8+ T cells (>28% of all CD8 T cells) that was identical to the infused clones (as confirmed by sequencing of the complementarity-determining region 3). In contrast to skin biopsies obtained from the patient before T cell infusion, postinfusion biopsies demonstrated loss of MART-1 expression, evidence of melanocyte damage, and the complete absence of melanocytes in affected regions of the skin. This study provides, for the first time, direct evidence in humans that antigen-specific immunotherapy can target not only antigen-positive tumor cells in vivo but also normal tissues expressing the shared tumor antigen.

4544 Background: Superficial transitional cell carcinoma (TCC) is an immune-responsive tumor evidenced by immunotherapy trials with BCG demonstrating improved survival. In contrast, more advanced muscle-invasive TCC is not considered an immunologically active tumor. Yet, host immune functions that may have a clinical impact on the biologic activity of these more invasive tumors have not been systemically evaluated. CD8+ T-cells are responsible for cytotoxicity and potential tumor eradication by interaction with antigen plus human leukocyte antigens (HLA). A clear association between intratumoral CD8+ T-cells and clinical outcome has not been established in TCC. Methods: We performed pathological, immunohistochemical and RT-PCR analyses of 69 TCC patient samples that were obtained with appropriate informed consent on an Institutional Review Board (IRB)-approved protocol. The samples were studied for pathological stage, tumor-associated antigen expression, class I HLA expression, and CD8+ intratumoral T-cells. Systemic CD8+ T-cells from one patient with positive CD8+ intratumoral T-cells were studied by tetramer analyses for reactivity against the NY-ESO-1 tumor antigen expressed on the patient’s tumor. Results: In a subset analysis, advanced TCC (pT2, pT3 and pT4) patients who had higher numbers of CD8+ tumor infiltrating lymphocytes (TILs) had a greater disease-free survival (p = 0.0002) and overall survival (p = 0.011) than similarly staged TCC patients with lower numbers of CD8+ TILs. In the multivariate analyses, CD8+ TILs (p = 0.04) and tumor stage (p < 0.001) were significant risk factors to predict overall survival. Furthermore, a CD8+ T-cell clone derived from one patient demonstrated strong recognition of the tumor antigen NY-ESO-1. Conclusions: This is the first report, to our knowledge, that CD8+ TILs is an important prognostic indicator for patients with advanced TCC. Investigational immunotherapy strategies to evoke CD8+ T-cell responses are warranted in patients with advanced TCC. [Table: see text]

Abstract Background: Cytotoxic T lymphocytes (CTLs) and T-helper type 1 (Th1) cells undoubtedly play a crucial role in the eradication of tumors in vivo. However, the production of Th1 cytokines such as IL-2 and IFN-γ is markedly suppressed in the majority of tumor-bearing hosts. Such defects in Th1-mediated immunity in cancer patients have made it difficult to induce tumor-specific CTLs that promote tumor rejection. Adoptive transfer of tumor-specific CTLs and Th1 cells can overcome the difficulty to induce tumor-specific immune response in cancer patients; however, the generation and expansion of tumor-specific CTLs and Th1 cells in vitro are not easy. In the present study, to overcome this problem, we isolated TCR-α and -β chain genes from a WT1-specific CD8+ CTL clone, which had been shown to exert strong cytotoxicity against hematopoietic malignancies and solid tumors in an HLA-A24-restricted manner, and transduced them into nonspecifically activated human CD8+ and CD4+ T cells. Consequently, both CD8+ and CD4+ T cells appeared to acquire WT1-specific function in an HLA-A24-restricted manner. Methods: A WT1 peptide (CMTWNQMNL)-specific CD8+ CTL clone, TAK-1, was established as reported previously (Blood95:286,2000). TAK-1 exerts cytotoxicity against variety of tumor cells including leukemia, myeloma, and lung cancer cells but not against normal cells in an HLA-A24-restricted manner. cDNAs encoding TCR-α and -β chain genes were amplified from cDNA of TAK-1 by RT-PCR. TCR-α and -β chain cDNAs were inserted into the plasmid vector. Preparation of lentiviral vectors for transduction of TCR-α and -β chain cDNAs was performed as described previously (Cancer Res64:1490,2004). Peripheral blood CD4+ and CD8+ T cells isolated from healthy individuals were cultured with anti-CD3 mAb and retronectin and then infected twice with lentivirus vectors. The infected cells were expanded by culture in the presence of IL-2, IL-12, IFN-γ and anti-IL-4 mAb. Cytotoxicity of CTLs against WT1-peptide-loaded cells and various human tumor cells was examined by a standard 51Cr-release assay. Recognition of tumor cells by Th1 cells was examined by measuring IFN-γ production by ELISA. Results: CD4+ T-cell line (CD4-TCR) and CD8+ T cell line (CD8-TCR) expressing TCR-α and -β chains of TAK-1 were established. Both CD4-TCR and CD8-TCR cells exerted cytotoxicity against WT1 peptide-loaded HLA-A24-positive but not -negative cells. CD8-TCR cells appeared to be cytotoxic against human tumor cells including leukemia, myeloma, and lung cancer cells in an HLA-A24-restricted manner, but did not show any cytotoxicity against HLA-A24-positive normal cells. CD4-TCR cells produced IFN-γ in response to stimulation with HLA-A24-positive but not -negative leukemia cells. Conclusion: The present data demonstrate the functional reconstitution of CD4+ as well as CD8+ T cells by transfer of the αβ TCR complex of a WT1-specific CD8+ CTL clone. Since WT1 is a universal tumor-associated antigen, transfer of TCR genes of WT1-specific CTLs into CD4+ and CD8+ T cells would be useful for Th1-based immunotherapy of various malignancies.

Abstract The Wilms tumor (WT1) gene has been reported to be preferentially expressed in acute leukemia cells, regardless of leukemia subtype and chronic myelogenous leukemia cells in blast crisis, but not in normal cells. This finding suggests strongly that WT1 protein is a potential target of immunotherapy for human leukemia. In this study, we established a CD8+ cytotoxic T-lymphocyte (CTL) clone directed against a WT1-derived peptide and examined its immunologic actions on leukemia cells. A CD8+ CTL clone, designated TAK-1, which lysed autologous cells loaded with a WT1-derived 9-mer peptide consisting of the HLA-A24 (HLA-A*2402)-binding motifs was established by stimulating CD8+ T lymphocytes from a healthy individual repeatedly with WT1 peptide-pulsed autologous dendritic cells. TAK-1 was cytotoxic to HLA-A24–positive leukemia cells expressing WT1, but not to HLA-A24–positive lymphoma cells that did not express WT1, HLA-A24–negative leukemia cells, or HLA-A24–positive normal cells. Treating leukemia cells with an antisense oligonucleotide complementary to the WT1 gene resulted in reduced TAK-1-mediated cytotoxicity, suggesting that target antigen of TAK-1 on leukemia cells is the naturally processed WT1 peptide in the context of HLA-A24. TAK-1 did not inhibit colony formation by normal bone marrow cells of HLA-A24–positive individuals. Because WT1 is overexpressed ubiquitously in various types of leukemia cells, but not in normal cells, immunotherapy using WT1 peptide-specific CTL clones should be an efficacious treatment for human leukemia. (Blood. 2000;95:286-293)

The Wilms tumor (WT1) gene has been reported to be preferentially expressed in acute leukemia cells, regardless of leukemia subtype and chronic myelogenous leukemia cells in blast crisis, but not in normal cells. This finding suggests strongly that WT1 protein is a potential target of immunotherapy for human leukemia. In this study, we established a CD8+ cytotoxic T-lymphocyte (CTL) clone directed against a WT1-derived peptide and examined its immunologic actions on leukemia cells. A CD8+ CTL clone, designated TAK-1, which lysed autologous cells loaded with a WT1-derived 9-mer peptide consisting of the HLA-A24 (HLA-A*2402)-binding motifs was established by stimulating CD8+ T lymphocytes from a healthy individual repeatedly with WT1 peptide-pulsed autologous dendritic cells. TAK-1 was cytotoxic to HLA-A24–positive leukemia cells expressing WT1, but not to HLA-A24–positive lymphoma cells that did not express WT1, HLA-A24–negative leukemia cells, or HLA-A24–positive normal cells. Treating leukemia cells with an antisense oligonucleotide complementary to the WT1 gene resulted in reduced TAK-1-mediated cytotoxicity, suggesting that target antigen of TAK-1 on leukemia cells is the naturally processed WT1 peptide in the context of HLA-A24. TAK-1 did not inhibit colony formation by normal bone marrow cells of HLA-A24–positive individuals. Because WT1 is overexpressed ubiquitously in various types of leukemia cells, but not in normal cells, immunotherapy using WT1 peptide-specific CTL clones should be an efficacious treatment for human leukemia. (Blood. 2000;95:286-293)

PURPOSE: To study distribution and toxicity of cytolytic T lymphocytes (CTLs) against a single melanoma epitope. PATIENTS AND METHODS: CD8+ T cells obtained by leukapheresis from 10 patients with disseminated HLA-A2.1+, tyrosinase-positive melanomas were immunized in vitro against tyrosinase369-377 (YMNGTMSQV). Drosophila cells transduced with HLA-A2.1, CD80, and CD54 (intracellular adhesion molecule-1) were used for priming, followed by two rounds of immunization with mononuclear cells as antigen-presenting cells. 1 × 108 CTL were infused intravenously (IV) on day 1. CTL frequency was measured by limiting dilutions in five patients. 111In labeling and scintigraphy measured distribution of CTL in next five. Five days later, 1 × 108 CTLs were infused on 4 successive days to both groups. Immunohistology of response was judged by biopsies. RESULTS: Infusions were nontoxic. CTLs were undetectable in the blood, going to lungs within 5 minutes. At 4, 24, and 72 hours, they were found in liver and spleen. Lesions were visualized by scintiscans in one responding patient where two subcutaneous nodules were noted at 4 and 24 hours. A second patient had a partial response and remains alive with disease 2 years later. CD8+ T cells were found in lesions of responders, associated with the presence of HLA-A2 molecules and tyrosinase. Two nonresponders without tyrosinase and HLA-A2 molecules had a paucity of CD8+ T cells in their lesions. Whether the CD8+ T cells in lesions of responders were those we had reinfused is uncertain. CONCLUSION: CTLs immunized against a single melanoma epitope were nontoxic but did not specifically localize to tumor sites. Nevertheless, two patients had disease regression. Additional therapeutic studies with specifically immunized CTL seem justified.