卵巢癌浸润淋巴细胞的免疫学特性研究

目的 研究卵巢癌浸润淋巴细胞（ＴＩＬ）在体外扩增后免疫学特性,评估ＴＩＬ用于晚期卵巢癌治疗的前景。方法 利用流式细胞仪分析ＴＩＬ的细胞表型,使用分子生物学和免疫学方法研究ＴＩＬ分泌细胞因子的能力和杀伤肿瘤细胞的活性。结果 ＴＩＬ细胞表型的差异可能与肿瘤的种类,性质,取材部位有关,结缔组织,基质中来源的ＴＩＬ以ＣＤ＾＋３,ＣＤ＾＋４为主,肿瘤组织和小血管周围以ＣＤ＾＋３,ＣＤ＾＋８为主,体外ＩＬ－２     

Ex vivo expansion of tumor-infiltrating lymphocytes from nasopharyngeal carcinoma patients for adoptive immunotherapy

Establishing Epstein-Barr virus (EBV)-specific cytolytic T lymphocytes (EBV-CTLs) from peripheral blood mononuclear cells (PBMCs) for adoptive immunotherapy has been reported in EBV-associated malignancies including Hodgkin's lymphoma and nasopharyngeal carcinoma (NPC). In the current study,we performed ex vivo expansion of tumor-infiltrating lymphocytes (TILs) obtained from NPC biopsy specimens with a rapid expansion protocol using anti-CD3 monoclonal antibody (OKT3), recombinant human interleukin (IL)-2, and irradiated PBMCs from healthy donors to initiate the growth of TILs. Young TIL cultures comprised of more than 90% of CD3+T cells, a variable percentage of CD3+CD8+and CD3+CD4+T cells, and less than 10% of CD3-CD16+natural killer cells, a similar phenotype of EBV-CTL cultures from PBMCs. Interestingly, TIL cultures secreted high levels of the Th1 cytokines, interferon gamma (IFNγ) and tumor necrosis factor-alpha (TNF-α), and low levels of the Th2 cytokines, IL-4 and IL-10. Moreover, young TILs could recognize autologous EBV-transformed B lymphoblast cell lines, but not autologous EBV-negative blast cells or allogeneic EBV-negative tumor cells. Taken together, these data suggest that ex vivo expansion of TILs from NPC biopsy tissue is an appealing alternative method to establish T cell-based immunotherapy for NPC.     

Characterization of tumor infiltrating lymphocytes in paired primary and metastatic renal cell carcinoma specimens

Renal cell carcinoma (RCC) is one of the most chemo- and radio-resistant malignancies, with poor associated patient survival if the disease metastasizes. With recent advances in immunotherapy, particularly with PD-1/PD-L1 blockade, outcomes are improving, but a substantial subset of patients does not respond to the new agents. Identifying such patients and improving the therapeutic ratio has been a challenge, although much effort has been made to study PD-1/PD-L1 status in pre-treatment tumor. However, tumor infiltrating lymphocyte (TIL) content might also be predictive of response, and our goal was to characterize TIL content and PD-L1 expression in RCC tumors from various anatomic sites. Utilizing a quantitative immunofluorescence technique, TIL subsets were examined in matched primary and metastatic specimens. In metastatic specimens, we found an association between low CD8+ to Foxp3+ T-cell ratios and high levels of PD-L1. High PD-L1-expressing metastases were also found to be associated with tumors that were high in both CD4+ and Foxp3+ T-cell content. Taken together these results provide the basis for combining agents that target the PD-1/PD-L1 pathway with agonist of immune activation, particularly in treating RCC metastases with unfavorable tumor characteristics and microenvironment. In addition, CD8+ TIL density and CD8:Foxp3 T-cell ratio were higher in primary than metastatic specimens, supporting the need to assess distant sites for predictive biomarkers when treating disseminated disease.     

Tumor infiltrating lymphocytes in ovarian cancer

The accumulation of tumor infiltrating lymphocytes (TILs) in ovarian cancer is prognostic for increased survival while increases in immunosuppressive regulatory T-cells (Tregs) are associated with poor outcomes. Approaches that bolster tumor-reactive TILs may limit tumor progression. However, identifying tumor-reactive TILs in ovarian cancer has been challenging, though adoptive TIL therapy in patients has been encouraging. Other forms of TIL immunomodulation remain under investigation including Treg depletion, antibody-based checkpoint modification, activation and amplification using dendritic cells, antigen presenting cells or IL-2 cytokine culture, adjuvant cytokine injections, and gene-engineered T-cells. Many approaches to TIL manipulation inhibit ovarian cancer progression in preclinical or clinical studies as monotherapy. Here, we review the impact of TILs in ovarian cancer and attempts to mobilize TILs to halt tumor progression. We conclude that effective TIL therapy for ovarian cancer is at the brink of translation and optimal TIL activity may require combined methodologies to deliver clinically-relevant treatment.     

肿瘤浸润淋巴细胞与lncRNA在肿瘤发展中作用的研究进展

肿瘤浸润淋巴细胞在肿瘤微环境中扮演着重要的角色,不同的细胞亚型在肿瘤的进展中发挥抗肿瘤或促肿瘤作用.肿瘤浸润淋巴细胞主要包括T淋巴细胞、B淋巴细胞和自然杀伤细胞,这些细胞可在多种因素的刺激下发生活化成熟及产生效应,参与肿瘤的免疫应答.长链非编码RNA(long non-codingRNA)是一类多效的调控因子,可参与机...     

膀胱移行细胞癌肿瘤浸润免疫细胞表型及临床意义

背景与目的:对肿瘤浸润免疫细胞分型有助于了解肿瘤局部的细胞免疫状态,对指导肿瘤免疫治疗具有一定意义。关于肾癌、卵巢癌、乳腺癌、结肠癌等肿瘤的免疫细胞表型已有报道。因此本研究旨在探讨膀胱移行细胞癌肿瘤浸润免疫细胞表型及临床意义。方法:采用免疫组化法对58例膀胱癌标本肿瘤浸润免疫细胞进行分型,测定细胞密度及癌巢进入率,与肿瘤分期、分级进行相关性分析。结果:肿瘤组织内存在CD3 T细胞(CD3 )、CD8 T细胞(CD8 )、巨噬细胞(Mφ)、树突状细胞(DC)和自然杀伤细胞(NK),正常膀胱黏膜未见DC和NK。浸润肿瘤和低分化肿瘤CD3 、CD8 和Mφ密度分别高于浅表肿瘤和高分化肿瘤,差异有显著性。浸润肿瘤NK密度显著高于浅表肿瘤,与肿瘤分化程度无相关性。DC密度与肿瘤分期、分化程度无相关性。浅表肿瘤CD3 癌巢进入率显著高于浸润肿瘤,与肿瘤分化程度无相关性。浅表肿瘤、高分化肿瘤CD8 、Mφ和DC癌巢进入率分别高于浸润肿瘤、低分化肿瘤,差异有显著性。NK癌巢进入率与肿瘤分期、分级无相关性。结论:DC和NK是伴随膀胱癌发生特异性出现的免疫细胞。随着肿瘤进展,多种免疫细胞进入肿瘤组织,数量呈上升趋势,提示机体免疫系统对肿瘤细胞发生积极的反应。但是多种免疫细胞的癌巢进入率却呈下降趋势,提示肿瘤进展可能使多种免疫细胞进入癌巢受阻。     

肝细胞性肝癌组织浸润淋巴细胞表型与分布研究

目的:肝细胞性肝癌组织浸润淋巴细胞与外周血T 细胞表型可能与肿瘤进展及预后相关,本研究检测肝癌患者组织及外周血T 细胞表型与分布,分析淋巴细胞表型变化与预后的关系。方法:分析2007年10月至12月中山医院147 例肝癌及癌旁组织浸润淋巴细胞表型（T 细胞或B 细胞表面标志物:CD3、CD8、CD4、CD20、CD19、Foxp 3）,表型与临床病理特征及预后的关系;检测26例肝癌外周血CD3、CD8、CD4 +T细胞数量并其比例变化。结果:癌巢内肿瘤浸润细胞明显少于癌周组织（P < 0.01）,癌周淋巴细胞主要分布于癌旁正常肝组织、汇管区,其与患者肝炎病史及肝硬化相关,表型以CD3 +T细胞为主,其中又以CD8 + 细胞毒性T 细胞为主;CD4 染色在多数病例为阴性,Foxp 3 仅在个别病例（15/ 109）呈阳性。肿瘤浸润淋巴细胞B 细胞标志CD20、CD19均为阴性。肿瘤组织内CD8 +T细胞浸润数量与预后正相关,而癌周浸润淋巴细胞数目与患者转移及复发无显著关系。结论:肝癌肿瘤浸润细胞在癌巢内明显少于癌周组织,肿瘤及癌周浸润细胞以CD8 + 细胞毒性T 细胞为主。肿瘤组织内CD8 +T细胞浸润数量与预后相关,而癌周浸润淋巴细胞数量与患者转移及复发无显著关系。      

The effects of selenium on tumor growth in epithelial ovarian carcinoma

Objective

Epidemiological studies suggest that selenium protects against the development of several cancers. Selenium (sodium selenite) has been reported to interfere with cell growth and proliferation, and to induce cell death. In this study, we tested whether selenium could have growth-inhibiting effect in ovarian cancer cells and an orthotopic animal model.

Methods

Cell growth in selenium-treated cells was determined in human ovarian cancer cells, A2780, HeyA8, and SKOV3ip1 using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assay. Animal experiment of selenium with paclitaxel was performed using SKOV3ip1 cells in nude mice to evaluate their inhibiting effect for tumor growth. In addition, another animal experiment of paclitaxel with or without selenium was performed to assess the effect of survival and food intake in mice.

Results

The in vitro growth of selenium-treated cells was significantly decreased dose-dependently in A2780, HeyA8, and SKOV3ip1 cells. Therapy experiment in mice was started 1 week after injection of the SKOV3ip1 cells. Treatment with selenium (1.5 mg/kg, 3 times/week) and paclitaxel injection showed no addictive effect of the inhibition of tumor growth. However, combination of selenium and paclitaxel showed the slightly increased food intake compared with paclitaxel alone.

Conclusion

Although selenium has growth-inhibiting effect in ovarian carcinoma cells in vitro, there is no additive effect on tumor growth in mice treated with combination of paclitaxel and selenium. However, food intake is slightly higher in selenium-treated mice during chemotherapy.     

Tumor-infiltrating lymphocytes and prognosis of hepatocellular carcinoma.

Tumor-infiltrating lymphocytes (TIL) were isolated from 17 human hepatocellular carcinomas (HCC). The proliferation of TIL cultured with recombinant interleukin-2 (rIL-2) was evaluated. We also examined prognosis in relation to TIL. Successful expansion of TIL was achieved in 16 of the 17 lesions. In 10 of the 16, TIL increased more than 100-fold. Cytotoxicity to the allogeneic HCC cell line, HC-4, was demonstrated in all 13 TIL cultures tested. Maximum cytotoxicity was noted two to four weeks after culture. No statistical difference was observed either with respect to prognosis based upon growth rate or the cytotoxicity demonstrated in vitro. The initial number of TIL per unit weight of tumor was, however, significantly greater in the group for which the prognosis was good (19.0 +/- 5.1 x 10(6) vs. 5.6 +/- 1.6 x 10(6), P < 0.05). It would appear that greater lymphocytic tumor infiltration is a prognostic marker.     

Prognostic and predictive value of tumor infiltrating lymphocytes in early breast cancer

It is well recognized that the immune system plays an essential role in tumor defense. The presence of tumor-infiltrating lymphocytes reflects an individual immunological response. In early breast cancer, the presence of TILs is associated with a more favorable outcome and response to therapy. In this review, we describe how TILs are assessed. Also, we discuss their role as prognostic and predictive biomarker in the neoadjuvant and adjuvant settings as well as in residual disease. Moreover, we discuss the possible implementation of TILs in daily clinical practice as well as in future clinical trials in order to fine tune prognosis and improve treatments.     

Lymphokine-activated killer cell activity of peripheral blood,spleen, regional lymph node,and tumor infiltrating lymphocytes in gastric cancer patients

Lymphokine-activated killer (LAK) cell activity of peripheral blood mono-nuclear cells (PBM), spleen cells (SPC), regional lymph node cells (LNC), and tumor-infiltrating lymphocytes (TIL), induced by activation with in-terleukin 2 (IL 2) for 4 days, was evaluated in patients with gastric carcinoma. TIL exhibited the lowest LAK activity and the cytotoxicity of LNC was significantly lower than that of either PBM or SPC. There was no difference between PBM and SPC. Then, there were significant correlations of LAK activity among PBM, SPC, and LNC, whereas poor correlations were observed in the cytotoxicity between TIL and PBM, SPC, or LNC. Phenotypic analysis of each cell population was performed before and after activation with IL 2. Before culture, the cells mediating natural killer (NK) activity such as CD16⁺, CD56⁺, and CD57⁺ cells were few in LNC and TIL. However, CD56⁺ and CD57⁺ cells in TIL were increased after culture. Then, CD4⁺ Leu8⁺ and CD8⁺ CD11⁺ cells, which identify suppressor cell function, were not elevated in LNC or TIL, as compared to that in PBM or SPC. Further, the proportions of OKIa1⁺ and CD25⁺ cells expressing T-cell activation and IL 2 receptor were uniformly increased in all cell populations after culture. These results indicate the differential reactivity of each lymphocyte population to IL 2 and fundamental dysfunction of LNC and, especially TIL, suggesting the specific influence of the local tumor environment on the lymphocyte function in the area in patients with gastric carcinoma. © Wiley-Liss, Inc.     

Endometrial carcinoma in women under 40 years of age: incidence of ovarian carcinoma and ovarian metastasis

Background. Patients under 40 years of age with endometrial carcinoma were compared with patients aged 40 years or more in terms of clinical and histopathological characteristics. Methods. One hundred and fifty-three patients with endometrial carcinoma who had their initial treatment in our hospital between 1980 and 1996 were divided into two groups; those under 40 years of age (group A) and those 40 years or more (group B). They were compared in terms of clinical stage, histological differentiation, degree of myometrial invasion, existence of lymph-vascular invasion, incidence of ovarian carcinoma with endometrial carcinoma, incidence of ovarian metastasis, treatment methods, and prognosis. Results. Fourteen patients (9.2%) were in group A and 139 patients (90.8%) in group B. There were no significant differences in the proportion of stage I patients in the two groups, but the proportion of stage IV patients was significantly higher in group A (P < 0.005). There were no significant differences in histological differentiation, degree of myometrial invasion, and the existence of lymph-vascular invasions. The incidence of ovarian carcinoma with endometrial carcinoma was 21.4% for group A and 2.2% for group B, being significantly higher in group A (P < 0.005). The incidence of simultaneous ovarian carcinoma with endometrial carcinoma was 14.2% for group A and 1.4% for group B, being significantly higher in group A (P < 0.05). The incidence of ovarian metastasis of endometrial carcinoma in groups A and B was 14.2% and 2.9% respectively, showing no significant difference. The incidence of either ovarian carcinoma with endometrial carcinoma or ovarian metastasis of endometrial carcinoma was significantly higher in group A (35.7%) than in group B (5%; P < 0.0005). There were no significant differences in the 5-year survival rates. Conclusion. Women aged under 40 years had a significantly higher incidence of ovarian carcinoma associated with endometrial carcinoma and a significantly higher incidence of either ovarian carcinoma with endometrial carcinoma or metastasis of endometrial carcinoma to the ovary than women aged 40 years or more. The 5-year survival rate showed no difference between the groups. Received: June 17, 1999 / Accepted: November 26, 1999     

Relationship between the expressions of PD-L1 and tumor-infiltrating lymphocytes in oral squamous cell carcinoma

Tumor-infiltrating lymphocytes (TILs) are considered to represent immune reactions of the host to a malignant tumor. Programmed death receptor ligand-1 (PD-L1) is a surface protein that blocks the function of T lymphocytes and is expressed on cancer cells. Tumor-associated fibroblasts (TAFs), which influence tumor growth have also been reported to express PD-L1 and thus inhibit TILs. In the present study, we investigated the densities of CD4⁺/CD8⁺ TILs, PD-L1 expression of tumor cells and TAFs in oral squamous cell carcinoma (OSCC). Forty-five cases of OSCC were selected. We evaluated PD-L1 expression and the infiltration degree of each lymphocyte by immunohistochemical examination. These data were analyzed in connection with clinicopathological factors. Peritumoral CD8⁺ TILs were observed in every patient with OSCC, and their densities were correlated with lymph node metastasis (P < 0.001), tumor size (P = 0.003), and clinical stage (P < 0.001). PD-L1 expression on OSCC cells was observed in 39 cases and was associated with the lower density of intratumoral CD8⁺ TILs (P = 0.047). PD-L1 expression of tumors <4 cm in size was correlated with the histological grade of the tumor (P = 0.022). TAFs were positive for PD-L1 in 18 cases. Peritumoral TILs were significantly associated with tumor size, lymph node metastasis and clinical stage. Though PD-L1 expressed by OSCC cells did not affect patients’ survival, its correlation with decreased number of intratumoral TILs suggests that the development of a strategy to block the interactions of PD-L1 with TIL would be a useful tool for inhibiting tumor growth.     

卵巢癌患者外周血T淋巴细胞的表达分析

目的探讨卵巢癌患者外周血T淋巴细胞的表达变化情况,分析CD+4CD+25表达与预后的关系。方法选择2009年2月到2012年11月收治的35例卵巢癌患者作为试验组,另选择35例同期体检排除肿瘤及免疫相关性疾病的健康人作为对照组,检测两组患者外周血中T淋巴细胞的表达水平。试验组同期分离肿瘤淋巴细胞进行免疫染色分析。结果对照组和试验组外周血淋巴细胞中CD+4CD+25的表达量分别为(5.36±0.63)%和(44.12±6.23)%,差异有统计学意义(P<0.05)。外周血淋巴细胞CD+4CD+25比例与卵巢癌患者年龄、NCCN分期、组织学类型和淋巴结转移均无关(P>0.05)。CD+4CD+25高表达组卵巢癌患者总生存期低于CD+4CD+25低表达组(P<0.05)。结论卵巢癌患者外周血CD+4CD+25淋巴细胞多呈现高表达,与预后密切相关,可成为卵巢癌预后指标和免疫治疗的生物靶点。     

Prognostic significance of tumor‐infiltrating lymphocytes in nondisseminated nasopharyngeal carcinoma: A large‐scale cohort study

The American Joint Committee on Cancer (AJCC) staging system is inadequate for an accurate prognosis in nasopharyngeal carcinoma (NPC). Thus, new biomarkers are under intense investigation. Here, we investigated whether the density of TILs could predict prognosis in NPC. First, we used 1490 cases of nasopharyngeal carcinoma samples from two independent cohorts to evaluate the density and distribution of tumor‐infiltrating lymphocytes (TILs). Second, in one cohort, we assessed associations between TILs and clinical outcomes in 593 randomly selected samples (defined as the training set) and validated findings in the remaining 593 samples (defined as the validation set). Furthermore, we confirmed the prognostic value of TILs in a second independent cohort of 304 cases (defined as the independent set). Based on multivariable Cox regression analysis, we also established an effective prognostic nomogram including TILs to improve accuracy in predicting disease‐free survival (DFS) for patients with nondisseminated NPC. We found that high TILs in the training set were significantly associated with favorable DFS [hazard ratio (HR) 0.41, 95% confidence interval (CI) 0.28–0.58, p < 0.001], overall survival (OS, HR 0.42, 95% CI 0.27–0.64, p < 0.001), distant metastasis‐free survival (DMFS, HR 0.37, 95% CI 0.23–0.58, p < 0.001) and local‐regional recurrent free survival (LRRFS, HR 0.43, 95% CI 0.25–0.73, p = 0.002). Multivariate analysis showed that TILs are an independent prognostic indicator for DFS in all cohorts. In summary, this study indicated that TILs may reflect the immunological heterogeneity of NPC and could represent a new prognostic biomarker.     

晚期上皮性卵巢癌肠肿瘤切除的作用

目的：分析晚期上皮性卵巢癌行肿瘤细胞减灭术时肠道转移瘤行肠道肿瘤切除的临床应用。方法：回顾性分析1998～2003年52例晚期上皮性卵巢癌行肿瘤细胞减灭术时肠道转移瘤行肠道肿瘤切除的患者,与同期未行肠道肿瘤切除的仅行姑息性手术的16例患者进行比较,采用统计学方法进行处理。结果：68例手术治疗患者中,52例完成肠道肿瘤切除手术,其中34例无肉眼呵见残余肿瘤,8例残余肿瘤〈1cm,10例残余肿瘤〉1cm,其中位生存期分别为28个月、23个月和13个月,16例因肿瘤广泛转移未行肠道肿瘤切除仅行姑息性手术的患者中位生存期为7．66个月,肠道肿瘤广泛转移及肠系膜根部广泛种植是手术失败的关键。结论：晚期上皮性卵巢癌行肿瘤细胞减灭术时行肠道转移瘤切除,达到满意手术效果时对生存期提高足有帮助的,而选择恰当的患者是手术治疗的关键。     

Diverse effect of cytokine treatment of tumor cells on specific versus non-specific cytotoxicity

(First submitted 15 Nov 1991;accepted 11 December 1991) The effect of IFN-γ and TNF-α treatment of an ovarian carcinoma line on the sensitivity to lysis by specific CTL clones and non-specific Tumor Associated Lymphocytes (TAL), isolated from the ascites fluid, was analyzed. Thein vitro established TAL line displayed a non-specific lytic activity against the autologous tumor as well as against several allogeneic tumor lines. Pretreatment with IFN-γ alone, or in combination with TNF-α, rendered the carcinoma line less susceptible to lysis by the autologous TAL line. Conversely, susceptibility to lysis by tumor specific T cell clones, isolated from the TAL line, was increased as a result of cytokine pretreatment. Several TCR-α/β⁺, CD8⁺ T-cell clones showing a more specific pattern of lysis against the autologous tumor were isolated. Lysis of the autologous tumor by these clones involved the TCR-α/β via a MHC-class I restricted mechanism dependent on the adhesion molecules ICAM-1 and LFA-3, as inferred from antibody blocking studies. The enhanced sensitivity to specific CTL clones seen after cytokine treatment may be related to theenhanced expression of ICAM-1 molecules on the ovarian carcinoma. These results have implications for cytokine based immunotherapy, where IFN-γ may enhance the effects of tumor associated specific CTL while decreasing that of non-specific effector cells.     

超声诊断卵巢透明细胞癌

目的 探讨术前超声诊断卵巢透明细胞癌(ovarian clear cell carcinoma,OCCC)的临床价值.方法 回顾性分析14例经手术病理检查证实的OCCC患者资料,观察OCCC的临床特征及术前超声表现.结果 3例病灶位于双侧卵巢,11例病灶位于单侧卵巢.超声表现为类圆形或类椭圆形肿块,边界清,有包膜;实性...