Antitumor effects of vaccination with dendritic cells transfected with modified receptor for hyaluronan-mediated motility mRNA in a mouse glioma model

OBJECT: The receptor for hyaluronan-mediated motility (RHAMM) is frequently overexpressed in brain tumors and was recently identified as an immunogenic antigen by using serological screening of cDNA expression libraries. In this study, which was conducted using a mouse glioma model, the authors tested the hypothesis that vaccination with dendritic cells transfected with RHAMM mRNA induces strong immunological antitumor effects. METHODS: The authors constructed a plasmid for transduction of the mRNAs transcribed in vitro into dendritic cells, which were then used to transport the intracellular protein RHAMM efficiently into major histocompatibility complex class II compartments by adding a late endosomal-lysosomal sorting signal to the RHAMM gene. The dendritic cells transfected with this RHAMM mRNA were injected intraperitoneally into the mouse glioma model 3 and 10 days after tumor cell implantation. The antitumor effects of the vaccine were estimated by the survival rate, histological analysis, and immunohistochemical findings for immune cells. Mice in the group treated by vaccination therapy with dendritic cells transfected with RHAMM mRNA survived significantly longer than those in the control groups. Immunohistochemical analysis revealed that greater numbers of T lymphocytes containing T cells activated by CD4+, CD8+, and CD25+ were found in the group vaccinated with dendritic cells transfected with RHAMM mRNA. CONCLUSIONS: These results demonstrate the therapeutic potential of vaccination with dendritic cells transfected with RHAMM mRNA for the treatment of malignant glioma.     

Identification of target antigens in specific immunotherapy for renal cell carcinoma

PURPOSE: Effective immunotherapy against renal cell carcinoma has not yet been established despite recent advances in specific immunotherapy for various malignancies. A plausible reason is limited information about target antigens of renal cell carcinoma. We searched for useful cancer antigens applicable to immunotherapy for renal cell carcinoma by examining antigen expression in renal cell carcinoma cell lines and testing the ability to induce renal cell carcinoma reactive cytotoxic T lymphocytes. MATERIALS AND METHODS: mRNA expression of a panel of cancer associated antigens was examined using 5 renal cell carcinoma cell lines. Thereafter antigen derived peptides reported to induce cancer reactive cytotoxic T lymphocytes from human leukocyte antigen-A24+ patients with cancer were examined for their potential to induce cytotoxic T lymphocytes from peripheral blood mononuclear cells of human leukocyte antigen-A24+ patients with renal cell carcinoma. RESULTS: Three candidate antigens, including multidrug resistance-associated protein 3, polycomb group protein enhancer of zeste homologue 2 and Her2/neu, were expressed in all 5 renal cell carcinoma cell lines. Six peptides derived from these antigens, including multidrug resistance-associated protein 3(503-511), multidrug resistance-associated protein 3(1293-1302), polycomb group protein enhancer of zeste homologue 2(291-299), polycomb group protein enhancer of zeste homologue 2(735-743), Her2/neu342-350 and Her2/neu485-493, efficiently induced peptide specific and renal cell carcinoma reactive cytotoxic T lymphocytes from human leukocyte antigen-A24+ patients with renal cell carcinoma. Blocking and cold inhibition assays revealed that cytotoxicity against renal cell carcinoma depended on human leukocyte antigen class I restricted and peptide specific CD8+ T cells. CONCLUSIONS: This information could facilitate the development of effective immunotherapy against renal cell carcinoma.     

Identification of Epstein-Barr virus-specific CD8+ T lymphocytes in the circulation of pediatric transplant recipients

BACKGROUND: Pediatric transplant recipients are at increased risk for Epstein Barr virus (EBV)-related B cell lymphomas. In healthy individuals, the expansion of EBV-infected B cells is controlled by CD8+ cytotoxic T cells. However, immunosuppressive therapy may compromise antiviral immunity. We identified and determined the frequency of EBV-specific T cells in the peripheral blood of pediatric transplant recipients. METHODS: HLA-B*0801 and HLA-A*0201 tetramers folded with immunodominant EBV peptides were used to detect EBV-specific CD8+ T cells by flow cytometry in peripheral blood mononuclear cells from 24 pediatric liver and kidney transplant recipients. The expression of CD38 and CD45RO on EBV-specific, tetramer-binding cells was also examined in a subset of patients by immunofluorescent staining and flow cytometry. RESULTS: Tetramer-binding CD8+ T cells were identified in 21 of 24 transplant recipients. EBV-specific CD8+ T cells were detected as early as 4 weeks after transplant in EBV seronegative patients receiving an organ from an EBV seropositive donor. The frequencies (expressed as a percentage of the CD8+ T cells) of the tetramer-binding cells were HLA-B8-RAKFKQLL (BZLF1 lytic antigen peptide) tetramer, range=0.96 to 3.94%; HLA-B8-FLRGRAYGL (EBNA3A latent antigen peptide) tetramer, range=0.03 to 0.59%; and HLA-A2-GLCTLVAML (BMLF1 lytic antigen peptide) tetramer, range=0.06 to 0.76%. The majority of tetramer reactive cells displayed an activated/memory phenotype. CONCLUSIONS: Pediatric transplant recipients receiving immunosuppression can generate EBV-specific CD8+ T cells. Phenotypic and functional analysis of tetramer cells may prove useful in defining and monitoring EBV infection in the posttransplant patient.     

Clonal T-cell populations and increased risk for cytotoxic T-cell lymphomas in B-CLL patients: clinicopathologic observations and molecular analysis

Chronic lymphocytic leukemia (CLL) is associated with increased risk of malignancy, but the occurrence of other lymphomas, in particular T-cell lymphomas, is rare. We identified 7 cases of peripheral T-cell malignancy associated with B-cell-derived CLL from the files of two institutions over a 20-year period. The presence of both B and T lymphoproliferative disorders was confirmed in all cases by immunophenotype and in 6 cases by gene rearrangements. Six patients developed peripheral T-cell lymphoma (PTCL), unspecified, during the course of CLL (10-168 months). In all 5 evaluable cases, the cells had a cytotoxic T-cell phenotype; the sixth case was CD56+, but TIA-1 and Granzyme B could not be studied. A seventh patient with CLL developed mycosis fungoides, and an aggressive NK cell leukemia. To investigate possible risk factors for the development of PTCL, we screened 100 unselected peripheral blood samples from newly diagnosed CLL patients by PCR for the presence of clonal T cell populations. We found evidence of clonal T-cell expansion in 8 patients and increased lymphocytes with large granular lymphocyte morphology in 7 of 8 cases. The immunophenotype was assessed by multicolor flow cytometry and in 4 cases the T-cell expansion was composed of either CD3+/CD8+ or CD3+/CD4-/CD8- cells. The cytotoxic nature of the clonal T-cell expansions in the peripheral blood correlates with the cytotoxic nature of the PTCLs, but their role in the subsequent development of T-cell lymphomas is still unclear. PTCL following CLL should be distinguished from typical Richter syndrome, which it can mimic clinically.     

Immunological evaluation of neoadjuvant peptide vaccination before radical prostatectomy for patients with localized prostate cancer

BACKGROUND: The purpose of this study was to determine the safety and immune responses of pre-operative personalized peptide vaccine for patients with localized prostate cancer. METHOD: Ten human leukocyte antigen (HLA)-A24(+) patients with localized prostate cancer received weekly personalized peptide vaccine for six times with positive peptides (up to four kinds of peptides) from 16 kinds of vaccine candidates, followed by a retropubic radical prostatectomy (RRP). Eight patients with localized prostate cancer receiving RRP served as the control group. The serum prostate-specific antigen (PSA) level, and peptide-specific cytotoxic T lymphocyte (CTL) precursor analysis by interferon-gamma production, and peptide-reactive immunoglobulin G (IgG) using an enzyme-linked immunosorbent assay were monitored during the treatment. Distributions of CD45RO(+) cells, CD8(+) T cells, and CD20(+) B cells in tissue microarray samples were studied using an immunohistochemical technique. RESULT: The peptide vaccination was safe and well tolerated with no major adverse effects. Increased CTL response and the anti-peptide IgG titer were observed in the post-vaccination samples in 8 of 10 or 8 of 10 patients, respectively. The intensity of CD45RO(+) infiltrating cells in the vaccination group was significantly larger than that in the control group. CD8(+) T cell infiltration was seen only in the vaccinated group. CONCLUSION: Increased immune responses, at both the circulation and tumor sites in the vaccinated group, support the further development of personalized peptide vaccines for patients with localized prostate cancer.     

辅助性T细胞和细胞毒性T淋巴细胞双表位修饰的树突状细胞肿瘤疫苗治疗胃癌的实验研究

目的研究辅助性T细胞（Th）表位和细胞毒性T淋巴细胞（CTL）双表位修饰的树突状细胞（DCs）肿瘤疫苗用于胃癌免疫治疗的效果。方法用CTL表位MAGE-341-49和Th表位MAGE-322-36混合多肽冲击DCs，每周刺激脾脏T细胞1次，4周后收集多肽特异性T细胞。流式仪分析T细胞亚群分布，测定CD4^＋T细胞识别抗原细胞因子分泌及CD8^＋T细胞杀伤肿瘤细胞效能，观察双表位修饰的DCs肿瘤疫苗治疗胃癌的保护性免疫效应。结果双表位致敏的DCs体外可同时活化CD4^＋和CD8^＋T细胞，其中CD4^＋T细胞识别肿瘤细胞小鼠前胃癌细胞株MFC后分泌大量Th1型细胞因子[干扰素（IFN）-γ，白介素（IL）-2]，CD8^＋T细胞强效杀伤MFC。双表位修饰的DCs肿瘤疫苗小鼠体内免疫治疗获得抵抗后继胃癌细胞MFC的免疫保护能力，并显著高于单一表位（CTL或Th）修饰的DC8疫苗。结论Th和CTL双表位修饰的DCs肿瘤疫苗可同时激活CD4^＋Th1细胞和CD8^＋CTL抗肿瘤免疫，有效清除胃癌细胞。     

Analysis of T cell subsets and beta chemokines in patients with pulmonary sarcoidosis

BACKGROUND: Sarcoidosis is a systemic granulomatous disorder of unknown origin characterised by accumulation of T lymphocytes and macrophages in multiple organs. Several cytokines and adhesion molecules may contribute to the accumulation of T lymphocytes in pulmonary sarcoidosis. The distribution of T lymphocyte subsets, T cell bearing CD11a and beta chemokines such as regulated on activation normal T expressed and secreted (RANTES), macrophage inflammatory peptide 1 alpha (MIP-1 alpha), and macrophage chemoattractant protein 1 (MCP-1) in bronchoalveolar lavage (BAL) fluid and peripheral blood were compared in untreated patients with sarcoidosis and normal subjects. METHODS: Flow cytometric analysis with monoclonal antibodies to cell surface antigens was used to identify T lymphocyte subsets in the BAL fluid of untreated patients with sarcoidosis (n = 40)--either without (group A, n = 12) or with (group B, n = 28) radiological evidence of pulmonary involvement--and in 22 normal subjects. The level of different beta chemokines was estimated by enzyme linked immunosorbent assay (ELISA). RESULTS: A high percentage of CD3+ cells, CD4+ cells expressing HLA-DR antigen, and a high CD4/CD8 ratio were detected in the BAL fluid of patients compared with normal subjects. In particular, CD4+ CD29+ memory T cells were significantly increased in patients with sarcoidosis. Furthermore, these cells were higher in those in group B than group A. The level of RANTES in the BAL fluid of patients was significantly higher than in normal subjects and correlated well with the percentage, number, and expression of CD29 on CD4 cells. The expression of CD11a (alpha chain of lymphocyte function associated antigen-1, LFA-1) on CD3+ cells in the BAL fluid of patients with sarcoidosis was not different from that of normal subjects. However, the expression of CD11a on CD3+ cells in the BAL fluid of patients in group A was significantly lower than that of patients in group B and normal subjects. CONCLUSIONS: These results suggest a possible interaction between activated memory T cells bearing CD11a and RANTES which may contribute to the pulmonary involvement in patients with sarcoidosis.


     

Active immunotherapy for advanced intracranial murine tumors by using dendritic cell-tumor cell fusion vaccines

OBJECT: Immunotherapy for malignant brain tumors by active immunization or adoptive transfer of tumor antigen-specific T lymphocytes has the potential to make up for some of the limitations of current clinical therapy. In this study, the authors tested whether active immunotherapy is curative in mice bearing advanced, rapidly progressive intracranial tumors. METHODS: Tumor vaccines were created through electrofusion of dendritic cells (DCs) and irradiated tumor cells to form multinucleated heterokaryons that retained the potent antigen processing and costimulatory function of DCs as well as the entire complement of tumor antigens. Murine hosts bearing intracranial GL261 glioma or MCA 205 fibrosarcoma were treated with a combination of local cranial radiotherapy, intrasplenic vaccination with DC/tumor fusion cells, and anti-OX40R (CD134) monoclonal antibody (mAb) 7 days after tumor inoculation. Whereas control mice had a median survival of approximately 20 days, the treated mice underwent complete tumor regression that was immunologically specific. Seven days after vaccination treated mice demonstrated robust infiltration of CD4+ and CD8+ T cells, which was exclusively confined to the tumor without apparent neurological toxicity. Cured mice survived longer than 120 days with no evidence of tumor recurrence and resisted intracranial tumor challenge. CONCLUSIONS: These data indicate a strategy to achieve an antitumor response against tumors in the central nervous system that is highly focused from both immunological and anatomical perspectives.     

Effective antigen cross-presentation by prostate cancer patients' dendritic cells: implications for prostate cancer immunotherapy

Despite the potency with which dendritic cells (DCs) are able to utilize the exogenous MHC I antigen cross-presentation pathway to cross-present antigen for the activation of killer T cells in model systems, concern about defects in immune function in cancer patients has led to uncertainty regarding whether immune cells derived from patients can effectively be used to generate tumor vaccines. We have undertaken a careful analysis of the potency of using DCs obtained from prostate cancer patients to cross-present antigen derived from human prostate tumor cells for the activation of antigen-specific T cells. Such DCs can be matured ex vivo into functionally active cells and are capable of cross-presenting influenza antigen derived from internalized apoptotic prostate tumor cells. Importantly, we demonstrate effective stimulation of both CD4+ and CD8+ T cells, as evident by production of IFN-gamma, and the ability of CD8+ T cells to differentiate into effector CTLs. These results, defining conditions in which prostate cancer patient DCs can efficiently utilize the cross-presentation pathway and in which apoptotic tumor can serve as a source of antigen for DCs to activate T cells, demonstrate that this system warrants clinical study as a potential immunotherapy.     

利用RNA干扰诱导产生的CD8+CD28-抑制性T淋巴细胞的免疫学特性

目的 探讨通过RNA干扰技术诱导产生的CD8+ CD28-抑制性T淋巴细胞(Ts细胞)的免疫学特性.方法 取SD大鼠骨髓,培养分离树突状细胞(DC),设计、合成主要组织相容性复合物(MHC)Ⅰ类小片段干扰RNA(siRNA),以MHC Ⅰ siRNA转染DC.先以Wistar大鼠肠系膜淋巴组织液刺激转染MHCI siRNA的DC,然后将DC与从SD大鼠脾脏分离得到的CD8+T淋巴细胞共同培养,通过磁珠法分离出Ts细胞.分别在由SD大鼠脾脏淋巴细胞(反应细胞)和Wistar大鼠肠系膜淋巴组织细胞(刺激细胞)组成的混合淋巴细胞培养体系中加入数量不等的Ts细胞,检测反应细胞增殖情况;分别以Wistar大鼠肠系膜淋巴组织细胞和卵白蛋白(OVA)刺激SD大鼠脾脏淋巴细胞,然后再按不同比例加入Ts细胞,检测各组脾脏淋巴细胞的增殖情况;在由SD大鼠脾脏淋巴细胞、Wistar大鼠肠系膜淋巴组织液和Ts细胞组成的混合淋巴细胞培养体系中加入可溶性重组白细胞介素2(rrIL-2),观察IL-2对Ts细胞功能的影响;采用实时定量聚合酶链反应(PCR)测定Ts细胞中转化生长因子β(TGF-β和γ干扰素(IFN-γ)mRNA的表达,流式细胞仪和实时PCR检测Ts细胞上CD25分子的表达.结果 Ts细胞对SD大鼠脾脏淋巴细胞和Wistar大鼠肠系膜淋巴组织细胞之问的混合淋巴细胞反应(MLR)具有抑制作用,但对于SD大鼠脾脏淋巴细胞和OVA之间的MLR则无抑制作用.在SD大鼠脾脏淋巴细胞、Wistar大鼠肠系膜淋巴组织液和Ts细胞组成的混合淋巴细胞培养体系中加入rrIL-2后,SD大鼠脾脏细胞的增殖并无明显增加(P＞0.05).与CD8+CD28+T淋巴细胞和CD8+ T淋巴细胞比较,Ts细胞的TGF-β和IFN-γ mRNA的表达量明显升高(P＜0.01,P＜0.05),而CD25的表达量明显降低(P＜0.05).结论 采用经MHC I siRNA干扰的DC能够诱导CD8+T淋巴细胞产生CD8+ CD28-Ts细胞;Ts细胞在体外具有免疫抑制特性,其免疫抑制作用不被外源性IL-2所逆转,且其免疫调节作用具有抗原特异性.     

膀胱癌免疫治疗相关肿瘤微环境及其在人和小鼠肿瘤中的组成特征初探

目的 应用生物信息学分析鉴定膀胱癌免疫治疗中起关键作用的肿瘤微环境（TME）组成特征,初步分析这些组分在人膀胱癌组织和BBN（N-butyl-N-（4-hydroxybutyl）-nitrosamine）诱导的自发小鼠膀胱肿瘤中表型的相似性。方法 分析晚期膀胱癌PD-L1阻断治疗IMvigor210数据集,利用DESeq2筛选与疗效相关的差异基因,通过GO和KEGG通路富集分析找出差异富集的通路;采用IOBR分析TME中的细胞浸润特征。通过TCGA数据库分析差异浸润细胞基因集与膀胱癌预后和分期的关系,在我院32例膀胱癌组织标本中进行免疫组化验证。建立BBN诱导的自发小鼠膀胱肿瘤模型,通过H&E染色和免疫组化对BBN诱导的自发小鼠膀胱肿瘤和人膀胱癌进展相似性及其TME主要组分进行比较。结果 在IMvigor210数据集中存在521个与免疫治疗疗效相关的差异基因,其中44个差异基因富集在含胶原细胞外基质上,提示成纤维细胞与免疫治疗疗效相关。TME分析发现细胞周期、组蛋白等肿瘤细胞基因集与NK细胞、CD8⁺ T细胞基因集在膀胱癌免疫治疗有效组中富集,而炎症性肿瘤相关成纤维细胞基因集在免疫治疗无效组中富集。TCGA分析显示：NK细胞和CD8⁺ T细胞基因集与膀胱癌患者总体生存时间呈正相关,CD56表达与膀胱癌病理分期呈正相关;PDGFRB表达与膀胱癌病理分期呈正相关。相比于低分期膀胱癌,PDGFRB⁺成纤维细胞在高分期膀胱癌中数量更多。在BBN小鼠模型中,BBN诱导的自发小鼠膀胱肿瘤发生率随着诱导时间逐渐升高,最终发展为肌层浸润性膀胱癌。免疫组化染色显示：小鼠和人膀胱肿瘤的TME中NK细胞和CD8⁺ T细胞以及成纤维细胞的浸润特征相似。结论 本研究结合生信分析和免疫组化评分发现：膀胱癌中NK细胞、CD8⁺ T细胞和成纤维细胞与免疫治疗疗效和预后具有相关性;这些细胞在BBN诱导的自发小鼠膀胱肿瘤与人膀胱癌组织中的浸润表型相似。这些结果可为研究膀胱癌免疫治疗的机制提供理论依据和动物模型。     

A prostate stem cell antigen-derived peptide immunogenic in HLA-A24- prostate cancer patients

BACKGROUND: We attempted to identify prostate stem cell antigen (PSCA)-derived peptides immunogenic in HLA-A24+ prostate cancer patients. METHODS: Peripheral blood mononuclear cells (PBMCs) were stimulated in vitro with each of three different PSCA-derived peptides, which were prepared based on the HLA-A24 binding motif, and their peptide-specific and HLA-A24-restricted anti-tumor responses were examined. Plasma levels of immunoglobulin G (IgG) against PSCA peptides were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Among three PSCA peptides, the PSCA 76-84 peptide most effectively induced peptide-specific cytotoxic T lymphocytes (CTLs) from PBMCs of HLA-A24+ prostate cancer patients. Cytotoxicity was dependent on peptide-specific and CD8+ T cells. The PSCA 76-84 peptide-stimulated PBMCs showed a significant level of cytotoxicity against prostate cancer cells in an HLA-A24-restricted manner. IgG reactive to the PSCA 76-84 peptide was detected in half of patients. CONCLUSIONS: The PSCA 76-84 peptide should be considered for use in clinical trials of immunotherapy for HLA-A24+ patients.     

Phenotypic definition of primed T cells in human renal allografts. Use of the CD45R marker

Immunohistological studies indicate that T cells and macrophages are the major components of human kidney allograft infiltrates. Recent work has demonstrated a division of T lymphocytes into 2 subpopulations with distinct functions on the basis of their expression of the CD45R antigen (CD45R+ "naive" and CD45R- "memory" T cells). This study analyzes CD45R expression on circulating T cells and T cells infiltrating renal allografts in patients undergoing rejection and/or cyclosporine nephrotoxicity. The percentage of circulating T cells that expressed CD45R in patients with rejecting (63 +/- 4) or stable grafts (66 +/- 3) was not different from values obtained for normal donors (62 +/- 3). In contrast, the percentage of T cells expressing CD45R infiltrating rejecting grafts was 21 +/- 2 and was not affected by the stage of rejection; in patients with CsA toxicity the value was 22 +/- 6. The reduced proportion of T cells that expressed CD45R in the allograft may reflect a change in status from the naive state due to alloantigenic stimulation (which can be demonstrated in vitro) and/or a propensity of memory T cells to enter or be retained in an allograft.     

Advances in specific immunotherapy for prostate cancer

OBJECTIVES: The absence of effective therapies for advanced prostate cancer has entailed an intensive search for novel treatments. This review presents an overview of specific immunotherapeutic strategies for prostate cancer. METHODS: Current literature was reviewed regarding the identification of tumor antigens and the design of T-cell- and antibody-based immunotherapy for prostate cancer. The PubMed database was searched using the key words antibodies, clinical trials, dendritic cells, immunotherapy, prostate cancer, and T cells. RESULTS: T cells and antibodies are powerful components of the specific antitumor immune response. CD8+ cytotoxic T lymphocytes (CTLs) efficiently destroy tumor cells. CD4+ T cells improve the antigen-presenting capacity of dendritic cells (DCs) and support the stimulation of tumor-reactive CTLs. Monoclonal antibodies exhibit their antitumor effects via antibody-dependent cellular cytotoxicity and complement activation. Consequently, much attention has been given to the identification of tumor antigens that represent attractive targets for specific immunotherapy. Several prostate cancer-related antigens were described and used in clinical trials. Such studies were based on the administration of peptides, proteins, or DNA. Furthermore, men with prostate cancer were vaccinated with peptide-, protein-, or RNA-loaded DCs, which display an extraordinary capacity to induce tumor-reactive T cells. Monoclonal antibodies directed against surface antigens were also used. Clinical trials revealed that immunotherapeutic strategies represent safe and feasible concepts for the induction of immunologic and clinical responses in men with prostate cancer. CONCLUSIONS: Specific immunotherapy represents a promising treatment modality for prostate cancer. Further improvement of the current approaches is required and may be achieved by combining T-cell- and antibody-based vaccination strategies with radio-, hormone-, chemo-, or antiangiogenic therapy.     

HLA class II antigen presentation by prostate cancer cells

Prostate cancer is the second most commonly diagnosed cancer in men. Recent evidence suggests that reduced expression of target protein antigens and human leukocyte antigen (HLA) molecules is the predominant immune escape mechanism of malignant prostate tumor cells. The purpose of this study was to investigate the prospect of antigen specific immunotherapy against prostate cancer via the HLA class II pathway of immune recognition. Here, we show for the first time that prostate cancer cells express HLA class II proteins that are recognized by CD4+ T cells. Prostate tumor cells transduced with class II molecules efficiently presented tumor-associated antigens/peptides to CD4+ T cells. This data suggests that malignant prostate tumors can be targeted via the HLA class II pathway, and that class II-positive tumors could be employed for direct antigen presentation, and CD4+ T-cell mediated tumor immunotherapy.Prostate Cancer and Prostatic Diseases (2008) 11, 334-341; doi:10.1038/sj.pcan.4501021; published online 16 October 2007.     

Foxp3+调节性T细胞与乳腺癌淋巴结转移及增殖的关系

目的:探讨Foxp3~+调节性T细胞(Tregs)与乳腺癌淋巴结转移及增殖的关系。方法:用免疫组化法检测168例女性乳腺癌患者癌组织中CD4和Foxp3(调节性T细胞标志)的表达,以42例女性乳腺良性病变乳腺组织为对照,分析Tregs与乳腺癌淋巴结转移及乳腺癌组织细胞核增殖相关抗原Ki-67表达的关系。结果:乳腺癌组织中CD4~+T细胞与Foxp3~+T细胞(Tregs)的数量均高于乳腺良性病变组织,差异均有统计学意义(均P0.05);有淋巴结转移的乳腺癌组织中CD4~+T细胞与Foxp3~+T细胞数均高于无淋巴结转移的乳腺癌组织,但仅后者差异有统计学意义(P0.05)。乳腺癌组织中Tregs的浸润数量与Ki-67表达之间无关(P0.05)。结论:乳腺癌组织微环境存在免疫抑制,Tregs浸润数量与淋巴结转移关系密切,但与肿瘤增殖无关,提示Tregs可作为判断乳腺癌患者有无淋巴结转移的新指标。     

Immune selection after antigen-specific immunotherapy of melanoma.

BACKGROUND: Melanoma antigen (MA)-specific vaccination strongly enhances antitumor reactivity in vivo and is capable of producing strong cytotoxic T lymphocyte responses in vitro. Furthermore, specific human leukocyte antigen-restricted T cell activation is hypothesized to occur in response to peptide-based immunotherapy, which may lead to the preferential killing of tumor cells bearing the relevant MA. The development of melanoma antigen-loss variants may subsequently occur in vivo. METHODS: Analysis of 532 melanoma lesions from 204 patients was performed on fine-needle aspiration biopsy specimens. Lesions were graded for the expression of the MAs gp100 and MART-1 with use of immunocytochemistry. A total of 351 melanoma lesions were divided into cohorts on the basis of the treatment received. The pretreatment group (n = 175) consisted of lesions obtained before any form of gp100 immunotherapy, with the posttreatment group (n = 176) consisting of lesions obtained after vaccination with a modified gp100 epitope, gp209-2M +/- interleukin 2 (IL-2). RESULTS: The percentage of lesions not expressing the gp100 antigen is greater than the percentage not expressing MART-1 (26% vs 14%). The frequency of lesions with high expression (> 75%) of gp100 significantly decreased with therapy (47% vs 34%) and conversely negative lesions increased (18% vs 29%). Treatment of lesions with peptide alone (no IL-2) revealed a significant decrease in gp100 expression (47% vs 32%), enhanced with the addition of IL-2 to therapy (47% vs 35%). The expression of MART-1 remained essentially unchanged unless IL-2 was added (54% vs 54%, MART-1 peptide alone, 54% vs 43%, MART-1 peptide + IL-2). Of 94 patients (181 lesions) assessed for gp100 expression before treatment, 10 patients responded to therapy. Pretreatment lesions in responding patients expressed some level of gp100 in all cases compared with 27% of nonresponding lesions, which were negative for gp100 expression. CONCLUSIONS. This study indirectly demonstrates that vaccination with an MA-derived peptide can result in immune selection in vivo. Furthermore, it provides strong immunologic evidence for the specificity of MA vaccines and to the relevance of MA expression in predicting the response to vaccination.     

Aberrant colonic expression of MHC class II antigens in Hirschsprung's disease.

The major histocompatibility complex (MHC) Class I and II cell surface antigens responsible for the recognition of self vs non-self were studied in patients with documented Hirschsprung's disease. Monoclonal antibodies reactive with monomorphic determinants of human lymphocyte antigen (HLA)-A,B,C (Class I) and HLA-DR (Class II) were used to demonstrate immunohistochemically the expression of MHC antigens in 27 biopsy specimens from a variety of colorectal disorders. The rectal specimens examined from patients with Hirschsprung's disease showed an unexpected, marked elevation of Class II antigens with abnormal localization in the mucosa and lamina propria. This ectopic expression was not seen in any portion of the small or large bowel of patients who did not have Hirschsprung's disease. Furthermore, proximal normal colon of children with Hirschsprung's disease failed to show increased expression of Class II antigen. In an attempt to better define the effector arm at a cellular level, the distribution of helper T cells (CD4+), cytotoxic/suppressor T cells (CD8+) and natural killer cells (NK; CD16+) was examined in 5 cases. In Hirschsprung's disease, rectal infiltration of CD8+ and CD16+ cells was found, but not CD4+ cells. Ectopic expression of Class II antigen with increased numbers of rectal T cells and NK cells suggested that an early immunologic event may be causal in Hirschsprung's disease.     

In vitro study of expression of interleukin-2 receptors in T-lymphocytes from patients with IgA nephropathy

The present study was undertaken to examine the expression of interleukin-2 receptor (IL-2R)/CD25 antigen in cultured T-lymphocyte subsets in IgA nephropathy. Twenty-four IgA nephritic patients, 12 patients with chronic glomerulonephritis (non-IgA nephropathy), and 17 healthy controls were studied in an infection-free interval. T-cell subsets in peripheral blood mononuclear cells (PBMC) and activated T-lymphocyte subsets expressing IL-2R were determined by double immunofluorescence staining with fluorochromes conjugated to monoclonal antibodies against T-helper/inducer (CD4+) cell, T-suppressor/cytotoxic (CD8+) cell, B (CD20+) lymphocytes, and IL-2R. The percentages of CD4+ and CD8+ lymphocytes, CD4/CD8 ratio, and total activated lymphocytes (with IL-2R/CD25 antigen) did not differ between the IgA nephritic patients, patients with chronic glomerulonephritis, and healthy controls in freshly isolated, unstimulated lymphocytes or PBMC cultured with pokeweed mitogen. Following pokeweed mitogen stimulation for 5 days, 17.3 +/- 10.3%, 16.6 +/- 8.4%, and 16.7 +/- 9.4% of PBMC from IgA nephritic patients, patients with chronic glomerulonephritis, and controls respectively expressed IL-2R (p greater than 0.05). However, the individual T-cell subsets bearing IL-2R were distinctly different between the IgA nephritic patients and patients with chronic glomerulonephritis or healthy controls. IgA nephritic patients had increased activated CD4+ lymphocytes (with IL-2R) (p less than 0.025) and reduced activated CD8+ lymphocytes (p less than 0.025). Our study suggests a defective immunoregulation in IgA nephropathy with enhanced T-helper/inducer and reduced T-suppressor/cytotoxic activity when stimulated with mitogen and probably, during clinical exacerbation.