Immunohistochemical analysis of lymphocyte subsets infiltrating gastric carcinoma after mitomycin C administration

Summary The intensity of lymphoid cell infiltration and distribution of lymphocyte subsets in tumors were investigated immunohistochemically on tumor tissues obtained from 11 patients with gastric carcinoma, who had been treated with mitomycin C (MMC), 12 mg/m², i.v. 5 days before operation. The results were compared with those obtained from 24 untreated patients as controls. In the tumor tissues from pretreated patients, the intensity of lymphoid infiltration was not significantly different from that of untreated patients. However, high-grade infiltration of CD4⁺ cells was observed in 55% of pretreated patients, whereas only 8% of control patients exhibited the high-grade infiltration (P <0.02). Since the CD8⁺ cell infiltration was not significantly altered, the ratio of CD4⁺ to CD8⁺ cells was more frequently estimated to be more than 1 in patients pretreated with MMC, as compared to untreated controls (P <0.02). Further, CD25⁺ cells in pretreated tumor tissues were more predominant than those in control tumor tissues (P <0.05). These results suggest that MMC administration induces these alterations in lymphocyte subsets in tumor tissue in patients with gastric carcinoma.     

Ontak reduces the immunosuppressive tumor environment and enhances successful therapeutic vaccination in HER-2/<Emphasis Type="Italic">neu</Emphasis>-tolerant mice

Disrupting tumor-mediated mechanisms suppressing host immunity represents a novel approach to tumor immunotherapy. Depletion of regulatory T cells (Tregs) increases endogenous anti-tumor immunity and the efficacy of active immunotherapy in experimental tumor models. HLA-A2.1/HLA-DR1 (A2.1/DR1) × BALB- neuT ⁺ (neuT ⁺) triple transgenic mice represent an improvement over neuT ⁺ mice for evaluating vaccination regimens to overcome tolerance against HER-2/neu. We questioned whether depletion of Tregs with Denileukin diftitox (Ontak) enhances the efficacy of a therapeutic vaccine consisting of HER-2(85–94) (p85) CTL and HER-2(776–790) (p776) Th peptides against the growth of TUBO.A2 transplantable tumor in male A2.1/DR1 × neuT ⁺ Tg mice. While the therapeutic vaccine primed the tumor-reactive CD8⁺ CTLs and CD4⁺ effector T lymphocytes (Teffs) compartment, inducing activation, tumor infiltration, and tumor rejection or delay in tumor growth, treatment with Ontak 1 day prior to vaccination resulted in enhanced CD4⁺ and CD8⁺ T-cell-mediated vaccine-specific immune responses in the periphery. This was closely associated with greater infiltration and a striking change in the intratumor balance of Tregs and vaccine-specific CTLs/Teffs that directly correlated with markedly enhanced antitumor activity. The data suggest that Tregs control both CD4⁺ and CD8⁺ T-cell activity within the tumor, emphasize the importance of the intratumor ratio of vaccine-specific lymphocytes to Tregs, and demonstrate significant inversion of this ratio and correlation with tumor rejection during Ontak/vaccine immunotherapy.     

肺癌患者外周血T淋巴细胞分型与抗核抗体之间的关系研究

摘要 目的：研究肺癌患者外周血T淋巴细胞分型与抗核抗体之间的关系。方法：选择2019年1月到2021年6月在我院接受治疗的肺癌患者81例作为研究组,并选择同期健康志愿者81例作为对照组,检测并比较两组患者外周血CD4⁺、CD8⁺和CD4⁺/CD8⁺淋巴细胞比例,以及抗核抗体血清滴度。比较不同抗核抗体、年龄、性别、TNM分期、肿瘤分化程度以及病理类型肺癌患者外周血CD4⁺、CD8⁺和CD4⁺/CD8⁺淋巴细胞比例。结果：（1）肺癌患者外周血CD4⁺和CD4⁺/CD8⁺淋巴细胞比例显著低于对照组,而CD8⁺淋巴细胞比例显著高于对照组（P<0.05）;（2）III+IV肺癌患者外周血CD4⁺、和CD4⁺/CD8⁺淋巴细胞比例均显著低于I+II肺癌患者,而CD8⁺淋巴细胞比例均显著高于I+II肺癌患者（P<0.05）;（3）小细胞肺癌患者外周血CD4⁺、和CD4⁺/CD8⁺淋巴细胞比例均显著低于非小肺癌患者,而CD8⁺淋巴细胞比例均显著高于非小肺癌患者（P<0.05）;（4）肺癌患者抗核抗体血清滴度显著高于对照组（P<0.05）;（5）抗核抗体阳性患者CD4⁺和CD4⁺/CD8⁺淋巴细胞亚群比例均显著低于抗核抗体阴性患者,而CD8⁺淋巴细胞亚群比例显著高于抗核抗体阴性患者（P<0.05）。结论：肺癌患者外周血T淋巴细胞亚群表达异常,并且其表达水平可能与抗核抗体滴度有关。     

Control of lymphocyte infiltration of lung tumors in mice by host’s genes: mapping of four Lynf (lymphocyte infiltration) loci

Tumor infiltration by lymphocytes is essential for cell-mediated immune elimination of tumors in experimental systems and in immunotherapy of cancer. Presence of lymphocytes in several human cancers has been associated with a better prognosis. We present evidence that individual propensity to tumor infiltration is genetically controlled. Infiltrating lymphocytes are present in 50% of lung tumors in O20/A mice, but in only 10% of lung tumors in OcB-9/Dem mice. This difference has been consistent in experiments conducted over 8 years in two different animal facilities. To test whether this strain difference is controlled genetically, we analyzed the presence of infiltrating lymphocytes in N-ethyl-N-nitroso-urea (ENU) induced lung tumors in (O20 × OcB-9) F₂ hybrids. We mapped four genetic loci, Lynf1 (Lymphocyte infiltration 1), Lynf2, Lynf3, and Lynf4 that significantly modify the presence and intensity of intra-tumoral infiltrates containing CD4⁺ and CD8⁺ T lymphocytes. These loci appear to be distinct from the genes encoding the molecules that are presently implicated in lymphocyte infiltration. Our findings open a novel approach for the assessment of individual propensity for tumor infiltration by genotyping the genes of the host that influence this process using DNA from any normal tissue. Such prediction of probability of tumor infiltration in individual cancer patients could help considerably to assess their prognosis and to decide about the application and the type of immunotherapy.     

Tumor‐Targeting Salmonella typhimurium A1‐R Promotes Tumoricidal CD8+ T Cell Tumor Infiltration and Arrests Growth and Metastasis in a Syngeneic Pancreatic‐Cancer Orthotopic Mouse Model

The present study determined the effect of the tumor‐targeting strain Salmonella typhimurium A1‐R (S. typhimurium A1‐R) on CD8⁺ tumor‐infiltrating lymphocytes (TILs) in a syngeneic pancreatic‐cancer orthotopic mouse model. The effect of tumor‐targeting S. typhimurium A1‐R on CD8⁺ TILs was determined on the Pan02 murine pancreatic‐adenocarcinoma implanted orthotopically in the pancreatic tail of C57BL/6 immunocompromised mice. Three weeks after orthotopic implantation, mice were randomized as follows G1: untreated control group (n = 8); and G2: S. typhimurium A1‐R‐treatment group (n = 8, 1 × 10⁷ colony forming units [CFU]/body, iv, weekly, 3 weeks). On the 22nd day from initial treatment, all mice were sacrificed and tumors were harvested. The tumor‐volume ratio was defined as ratio of tumor volume on the 22nd day relative to the 1st day. The tumor volume ratio was significantly lower in the S. typhimurium A1‐R‐treated group (G2) (3.0 ± 2.8) than the untreated control (G1) (39.9 ± 30.7, P < 0.01). Hematoxylin and easin (H&E) staining on tumor sections was performed to evaluate tumor destruction which was classified according to the Evans grading system and found to be much greater in the S. typhimurium A1‐R‐treated mice (G2). Six mice in G1 had peritoneal dissemination, whereas no mice showed peritoneal dissemination in G2 (P < 0.01). Immunohistochemical staining with anti‐mouse CD8⁺ antibody was performed in order to detect TILs determined by calculating the average number of CD8⁺ cells in three high power fields (200×) in the treated and untreated tumors. The TIL score was significantly higher in G2 (133.5 ± 32.2) than G1 (45.1 ± 19.4, P < 0.001). The present study demonstrates that S. typhimurium A1‐R promotes CD8⁺ T cell infiltration and inhibition of tumor growth and metastasis. J. Cell. Biochem. 119: 634–639, 2018. © 2017 Wiley Periodicals, Inc.     

Analysis of peripheral blood lymphocyte subsets and prognosis in patients with septic shock

The purpose of the study is to study the relationship between peripheral blood lymphocyte subset proportion and prognosis in patients with septic shock. Fifty‐two patients with septic shock, admitted to the intensive care unit between March 2007 and December 2010, were enrolled in this study. Peripheral blood lymphocyte subset proportions were measured using flow cytometry. The percentage of CD3⁺CD4⁺ T lymphocytes and CD19⁺ lymphocytes, CD4⁺/CD8⁺ T cell ratio were substantially lower in patients with septic shock compared to the control group (P < 0.01). The percentage of CD3⁺CD8⁺ T lymphocytes did not differ significantly between the two groups (P > 0.05). The percentage of CD16⁺CD56⁺ lymphocytes was higher in patients with septic shock than in the control group (P < 0.01). Compared with the survivor group, the percentage of CD3⁺CD4⁺ T lymphocytes and CD19⁺ lymphocytes, CD4⁺/CD8⁺ T cell ratio were clearly lower in the non‐survivor group (P < 0.01). There was no difference in the percentage of CD3⁺CD8⁺ T lymphocytes between the non‐survivor and survivor groups (P > 0.05). The percentage of CD16⁺CD56⁺ lymphocytes was higher in the non‐survivor group than in the survivor group (P < 0.05). The total maximum SOFA score and the delta SOFA score were much higher in the non‐survivor group than in the survivor group (P < 0.01). Immune imbalance occurs in patients with septic shock. Peripheral blood lymphocyte subset proportion and SOFA scores can be used to assess the treatment and prognosis of septic shock.     

MSI-H colorectal cancers preferentially retain and expand intraepithelial lymphocytes rather than peripherally derived CD8<Superscript>+</Superscript> T cells

The healthy colorectal mucosa contains many resident intraepithelial lymphocytes (IELs) consisting of partially activated yet hyporesponsive CD8⁺ T cells. A predominant feature of colorectal cancers (CRCs) characterized by high levels of microsatellite instability (MSI-H) is heavy infiltration by an intraepithelial population of tumor infiltrating lymphocytes (iTILs). While it has been assumed that these iTILs originate from tumor infiltration by peripheral CD8⁺ effector T cells, their origin remains unknown. In light of the phenotypic and functional differences exhibited by IELs and peripheral T cells, elucidation of the precursor population of iTILs in MSI-H CRCs could clarify the role played by these lymphocytes in tumor progression. The aim of the present study was to investigate whether MSI-H CRCs interact differently with IEL- versus peripherally-derived CD8⁺ T cells. Using a Transwell assay system to mimic basolateral infiltration of tumor cells by lymphocytes, T cell migration, retention, proliferation and phenotypic alterations were investigated. Results indicate that MSI-H CRCs preferentially retain and expand IEL-derived cells to a greater degree than their microsatellite stable (MSS) counterparts. While MSI-H CRCs also retained more peripherally derived T cells, this number was considerably less than that from the IEL population. While interaction of IELs with either CRC type led to baseline lymphocyte activation, MSS CRCs induced upregulation of additional activation markers on retained IELs compared to MSI-H CRCs. These results suggest that the abundant iTILs present in MSI-H CRCs result from expansion of the preexisting mucosal IEL population and imply a limited prognostic role for iTILs in MSI-H CRC.     

Prognostic impact of B-cell density in cutaneous melanoma

Studies on the prognostic importance of tumor-infiltrating lymphocytes have mainly focused on T cells, while little is known about the role of tumor-infiltrating B lymphocytes. We investigated the prevalence of CD20⁺ B cells by immunohistochemistry in primary melanoma samples of 106 patients and analyzed in relation to clinicopathological parameters and patients’ survival. The majority of samples contained a significant amount of B lymphocytes, predominantly dispersed in the stroma surrounding tumor deposits (mean peritumoral and intratumoral densities: 178.7 ± 156.1 vs. 4.9 ± 6.9 cells/mm², respectively). B cells organized in follicle-like aggregates were also observed in 26% of the samples. B-cell density correlated with that of activated (CD25⁺ or OX40⁺) T lymphocytes. Infiltration by CD20⁺ lymphocytes did not correlate with tumor thickness, while the presence of B-cell aggregates was observed more frequently in thick melanomas. On the other hand, B-cell infiltration was more pronounced in nonmetastatic or lymph node metastatic tumors, compared to visceral metastatic ones. Accordingly, high number of these cells provided significant survival advantage (P = 0.0391 and P = 0.0136 for intra- and peritumoral infiltration, respectively). Furthermore, combination of peritumoral B-cell density with the number of activated T lymphocytes identified patient subgroups with different disease outcome, which was most favorable in the case of high density, while very poor in the case of low density of both cell types. Multivariate survival analysis identified tumor thickness and CD20⁺/OX40⁺ cell density combination as significant independent prognostic factors. Taken together, our results show correlation between low number of CD20⁺ B lymphocytes and melanoma progression, indicating a possible role of tumor-infiltrating B cells in antitumoral immune response. It was also reflected in better outcome of the disease since the density of B lymphocytes alone as well as in combination with that of activated T cells proved of prognostic importance in patients with malignant melanoma.     

Polyclonal activation of human lymphocytes and induction of cytotoxic lymphocytes by streptococcal preparations

Polyclonal activation of human peripheral blood lymphocytes (PBLs)in vitro by preparations ofStreptococcus pyogenes Su strain (OK-432) and other heat-killed strains was investigated. The streptococcal preparations tested induce a proliferative response of PBLs via interleukin-2 (IL-2)-independent pathways. The proliferative response is accompanied by the generation of lymphoblastic cells (LBCs), which consist of heterologous lymphocyte populations: CD4⁺ helper type of T cells, and CD4^–CD8^– double-negative (DN) lymphocytes, including both CD3⁺ TcR ⁺ T cells and CD2⁺CD3^– immature type of T or non-T cell type of lymphocytes. Almost all the LBCs express Leu19, TfR (transferrin receptor), LFA-1 and CD38 (OKT10) antigens, which are expressed on activated T cells, NK cells and some other lymphocytes. The proliferative response of human PBLs is also accompanied by the generation of potent cytotoxic activity against NK-sensitive and -resistant targets. C-dependent cytolysis and cell sorting experiments of OK-432-activated LBCs revealed that both CD3⁺ and CD3^– types of CD4^–CD8^– DN lymphocytes, but not CD4⁺ helper T cells, may be major populations responsible for the cytotoxicity induced. On the other hand, CD4^–CD8 T cells may be required for the proliferation of PBLs and generation of cytotoxic effector cells. These results suggest that the OK-432 and other streptococcal preparations stimulate the human PBLsin vitro to induce the proliferation/activation of CD4⁺ T cells, mediating the following generation of DN cytotoxic effector lymphocytes.     

Immortalization of human T lymphocytes by oncogenes

Immortalized human T cell lines were established by cotransfecting c-Ha-ras and c-myc oncogenes to lymph node lymphocytes. The cell lines kept growing for 3 months after establishment without a decrease in growth rate. The cells did not require interleukin-2(IL-2) for their growth, but addition of IL-2 stimulated the growth of these cells. Flow cytometric analysis revealed that these cells were T cells expressing CD4 or CD8 antigens. A CD4 positive (CD4⁺) cell line produced IL-6, indicating that the cell line belongs to helper T cells. The CD8 positive (CD8⁺) cell line possessed cytotoxicity to tumor cells, indicating that the cell line were killer T cells. Both cell lines were able to proliferate in serum-free medium indefinitely.     

Effect of cytokines (IL-2, IL-7, and IL-15) having common γ-chain of receptors on differentiation and maturation of CD4+/CD8+ T-cells in a CD45RA T-lymphocyte population in vitro

The effect of cytokines (IL-2, IL-7, and IL-15) having a common γ-chain of receptors on the maturation and differentiation of CD3⁺CD45RA⁺CD4⁺/CD8⁺ lymphocytes in homeostatic cultivation model in vitro was analyzed. It was found that the maximum IL-2 concentration in the helper CD45RA⁺-T-cell population mediates an increase in the number of CD45RA⁺CD4⁺ T lymphocytes with the phenotype of mature and immature terminally differentiated TEMRA T cells. IL-15 leads to the production of lymphocytes with CD27^–CD62L⁺ phenotype (presumably, TEMRA, in which the CD62L expression persists). In the CD45RA⁺CD8⁺ T lymphocyte populations, the studied cytokines (IL-2, IL-7, and IL-15) initiate the production of mature TEMRA (E) T lymphocytes and memory T cells with the CD45RA^?CD27⁺CD62L⁺ central phenotype (TCM).     

Naïve rat umbilical cord matrix stem cells significantly attenuate mammary tumor growth through modulation of endogenous immune responses

Background aimsUn-engineered human and rat umbilical cord matrix stem cells (UCMSCs) attenuate growth of several types of tumors in mice and rats. However, the mechanism by which UCMSCs attenuate tumor growth has not been studied rigorously.MethodsThe possible mechanisms of tumor growth attenuation by rat UCMSCs were studied using orthotopic Mat B III rat mammary tumor grafts in female F344 rats. Tumor-infiltrating leukocytes were identified and quantified by immunohistochemistry analysis. Potential cytokines involved in lymphocyte infiltration in the tumors were determined by microarray and Western blot analysis. The Boyden chamber migration assay was performed for the functional analysis of identified cytokines.ResultsRat UCMSCs markedly attenuated tumor growth; this attenuation was accompanied by considerable lymphocyte infiltration. Immunohistochemistry analysis revealed that most infiltrating lymphocytes in the rat UCMSC-treated tumors were CD3⁺ T cells. In addition, treatment with rat UCMSCs significantly increased infiltration of CD8⁺ and CD4⁺ T cells and natural killer (NK) cells throughout tumor tissue. CD68⁺ monocytes/macrophages and Foxp3⁺ regulatory T cells were scarcely observed, only in the tumors of the phosphate-buffered saline control group. Microarray analysis of rat UCMSCs demonstrated that monocyte chemotactic protein-1 is involved in rat UCMSC-induced lymphocyte infiltration in the tumor tissues.ConclusionsThese results suggest that naïve rat UCMSCs attenuated mammary tumor growth at least in part by enhancing host anti-tumor immune responses. Naïve UCMSCs can be used as powerful therapeutic cells for breast cancer treatment, and monocyte chemotactic protein-1 may be a key molecule to enhance the effect of UCMSCs at the tumor site.     

Systemic treatment with interleukin-4 induces regression of pulmonary metastases in a murine renal cell carcinoma model

Advanced metastatic renal cell carcinoma has been shown to be responsive to immunotherapy but the response rate is still limited. We have investigated the therapeutic potential of systemic interleukin-4 (IL-4) administration for the treatment of pulmonary metastases in the murine Renca renal adenocarcinoma model. Renca cells were injected iv in Balb/c mice to induce multiple pulmonary tumor nodules. From Day 5, Renca-bearing mice were treated with two daily injections of recombinant murine IL-4 for 5 consecutive days. IL-4 treatment induced a significant reduction in the number of lung metastases in a dose-dependent manner and significantly augmented the survival of treated animals. Immunohistochemistry studies, performed on lung sections, showed macrophage and CD8⁺ T cell infiltration in the tumor nodules 1 day after the end of IL-4 treatment. The CD8 infiltration increased by Day 7 after IL-4 treatment. Granulocyte infiltration was not detectable. To clarify further the role of the immune system in IL-4 anti-tumor effect, mice were depleted of lymphocyte subpopulations by in vivo injections of specific antibodies prior to treatment with IL-4. Depletion of CD8⁺ T cells or AsGM1⁺ cells abrogated the effect of IL-4 on lung metastases, whereas depletion of CD4⁺ T cells had no impact. These data indicate that CD8⁺ T cells and AsGM1⁺ cells are involved in IL-4-induced regression of established renal cell carcinoma.     

Recruitment of CCR6+ Foxp3+ regulatory gastric infiltrating lymphocytes in Helicobacter pylori gastritis

Helicobacter pylori (H. pylori) infection is associated with an inflammatory response in the gastric mucosa, leading to chronic gastritis, peptic ulcers, and gastric cancer. Increased T‐cell infiltration is found at sites of H. pylori infection. The CCR6⁺ subset of CD4⁺ regulatory T cells (Tregs), a newly characterized subset of Tregs, has been reported to contribute to local immune inhibition. However, whether CCR6⁺ Tregs are present in H. pylori gastritis, and what their relationship is to disease prognosis, remains to be elucidated. In this study, gastric infiltrating lymphocytes were isolated from endoscopic biopsy specimens of H. pylori gastritis patients and analyzed. We found that in gastric infiltrating lymphocytes, CCR6⁺ CD4⁺ CD25^high Tregs, which express high levels of CD45RO, are positively associated with more severe inflammation in gastric mucosa during H. pylori infection. Furthermore, the frequency of CCR6⁺ Tregs in gastric infiltrating lymphocytes, but not CCR6^? Tregs, is significantly increased in inflamed gastric tissues, which is inversely correlated with significantly lower expression of IFN‐γ⁺ CD8⁺ T cells. We also found that the frequency of CCR6⁺ Tregs is positively correlated with the frequency of CD4⁺ IFN‐γ⁺ T cells. In addition, the frequency of CCR6⁺ Tregs, but not that of CCR6^? Tregs, is significantly correlated with increased inflammation in H. pylori gastritis. This study demonstrates that immunosuppression in H. pylori gastritis might be related to the activity of CCR6⁺ Tregs, which could influence disease prognosis.     

MHC-class I-restricted CD4 T cells: a nanomolar affinity TCR has improved anti-tumor efficacy in vivo compared to the micromolar wild-type TCR

Clinical studies with immunotherapies for cancer, including adoptive cell transfers of T cells, have shown promising results. It is now widely believed that recruitment of CD4⁺ helper T cells to the tumor would be favorable, as CD4⁺ cells play a pivotal role in cytokine secretion as well as promoting the survival, proliferation, and effector functions of tumor-specific CD8⁺ cytotoxic T lymphocytes. Genetically engineered high-affinity T-cell receptors (TCRs) can be introduced into CD4⁺ helper T cells to redirect them to recognize MHC-class I-restricted antigens, but it is not clear what affinity of the TCR will be optimal in this approach. Here, we show that CD4⁺ T cells expressing a high-affinity TCR (nanomolar K _d value) against a class I tumor antigen mediated more effective tumor treatment than the wild-type affinity TCR (micromolar K _d value). High-affinity TCRs in CD4⁺ cells resulted in enhanced survival and long-term persistence of effector memory T cells in a melanoma tumor model. The results suggest that TCRs with nanomolar affinity could be advantageous for tumor targeting when expressed in CD4⁺ T cells.     

顺铂联合VEGF抗体对食管癌移植瘤小鼠免疫调节以及癌细胞增殖和肺转移的影响

摘要 目的：研究顺铂联合血管内皮生长因子（vascular endothelial growth factor,VEGF）治疗对食管癌移植瘤小鼠免疫功能、癌细胞增殖以及肺转移的影响。方法：30只BALB/c小鼠通过皮下注射食管癌移植瘤模型。一周后,30只食管癌移植瘤模型小鼠被随机均分为3组,即模型组、顺铂组和联合组。模型组不进行治疗,顺铂组腹腔注顺铂治疗,联合组腹腔注射顺铂联合尾静脉注射VEGF抗体进行治疗,共治疗7周。比较各组小鼠体重,食管癌移植瘤体积和重量,卵巢癌细胞肺组织转移结节数、癌细胞转移面积和转移病灶总数,以及食管癌移植瘤外周血CD4⁺、CD8⁺以及CD4⁺/CD8⁺ T淋巴细胞比例。结果：（1）顺博组和联合组小鼠体重均显著高于对照组,而联合组小鼠体重显著高于顺铂组（P<0.05）;（2）顺铂组和联合组小鼠CD4+和CD8+细胞比例均显著低于对照组（P<0.05）,而CD4⁺/CD8⁺却显著高于对照组（P<0.05）;（3）联合组小鼠CD4⁺和CD8⁺细胞比例均显著高于对照组（P<0.05）,而CD4⁺/CD8⁺却显著低于顺铂组（P<0.05）;（4）顺铂组和联合组小鼠食管癌肿瘤组织体积和重量,肺转移结节数、转移面积和转移病灶数均显著低于对照组（P<0.05）,而联合组小鼠显著低于顺铂组（P<0.05）。结论：VEGF抗体可以显著增强顺铂在体内对食管癌的抗癌特性,并有助于增强食管癌移植瘤小鼠免疫功能、抑制癌细胞体内增殖和肺部转移。     

Tumor-infiltrating lymphocytes from nonrenal urological malignancies

Summary Tumor-infiltrating lymphocytes (TIL) were isolated from 15 of 20 surgical specimens of transitional cell carcinoma of the urinary bladder, prostate cancer, testicular cancer, Wilms tumor and adrenal cancer. Expansion was carried out in four different culture conditions, each containing 1000 U/ml interleukin-2: RPMI medium with or without 20% (by volume) of lymphokine-activated killer cell (LAK) supernatant and AIM V medium with or without 20% LAK supernatant. The resultant cell populations were then assayed for cytotoxicity against a variety of autologous and allogeneic tumor targets and phenotypic analysis was performed with fluorescein-labeled monoclonal antibodies. TIL growth was unrelated to the initial percentage of lymphocytes or tumor cells present in the enzymatically dispersed specimens or whether fresh or cryopreserved tissue was utilized. Better growth was seen in AIM V than in RPMI medium (P = 0.013); the beneficial effect of the addition of LAK supernatant to RPMI was indicated (P = 0.065), and the addition of LAK supernatant to AIM V did not improve the ability to culture TIL (P = 0.5) from these cancers. TIL in long-term culture were predominantly CD3⁺. The ratio of CD4⁺/CD8⁺ cells varied with time in culture and culture medium, but most cultures eventually became CD4⁺. Cells bearing B cell, natural killer cell, and macrophage markers disappeared early in culture. Overall 14/15 TIL samples were lytic against one of the autologous and allogeneic targets tested, but specific lysis against the autologous tumor from which it was derived was seen in only one TIL culture originating from a bladder cancer. Our results suggest that TIL can be expanded to therapeutic levels from a variety of urological malignancies and that their potential role in future therapy should be further explored.     

Modification of tumor cells by a low dose of Newcastle disease virus

Summary Augmented tumor-specific T cell responses were observed against the high metastatic murine lymphoma variant ESb when using as immunogen ESb tumor cells that had been modified by infection with a low dose of Newcastle disease virus (NDV). Such virus-modified inactivated tumor cells (ESb-NDV) were potent tumor vaccines when applied postoperatively for active specific immunotherapy of ESb metastases. We demonstrate here that immune spleen cells from mice immunized with ESb-NDV contain enhanced immune capacity in both the CD4⁺, CD8^– and the CD4^–, CD8⁺ T cell compartments to mount a secondary-tumor-specific cytotoxic T cell response in comparison with immune cells from mice immunized with ESb. ESb-NDV immune CD4⁺, CD8^– helper T cells also produced more interleukin 2 after antigen stimulation than the corresponding ESb immune cells. There was no participation of either CD4⁺ or CD8⁺ virus-specific cells in the augmented response. The specificity of the T cells for the tumor-associated antigen remaind unchanged. Thus, there is the paradox that the virus-mediated augmentation of the tumor-specific T cell response in this system involves increased T helper activity but does not involve the recognition of viral epitopes as potential new helper determinants.Abbreviations CTL cytolytic T lymphocytes - IL-2 interleukin 2 - rIL-2 recombinant IL-2 - mAb monoclonal antibody - NDV Newcastle disease virus - SSC syngeneic spleen cell     

B7-H4 expression associates with cancer progression and predicts patient’s survival in human esophageal squamous cell carcinoma

A retrospective cohort study including 112 patients suffering from esophageal squamous cell carcinoma (ESCC) was performed to investigate the expression of B7-H4 in ESCC and determine its association with patient’s clinicopathological parameters and survival. Expression levels of B7-H4 on tumor cells and densities of tumor infiltrating lymphocytes (TILs) in the surgical specimens of ESCC tissues were characterized using immunohistochemical assays. Uni- and multivariate analyses were performed to evaluate the prognostic value of B7-H4 expression levels and densities of TILs in tumor sections. Positive B7-H4 immunostaining was observed in 107 of 112 (95.5%) of ESCC tissue sections. We further divided all patients into two major subgroups, a lower B7-H4 expression group with 46 patients and a higher B7-H4 expression group with 66 patients. We found that expression levels of B7-H4 on tumor cells were significantly correlated with patient’s gender (P = 0.0288), distant metastasis (P = 0.0500), and TNM stage (P = 0.0258). Moreover, tumor cell B7-H4 expression was inversely correlated with densities of CD3⁺ T cells in tumor nest (P = 0.0424) and CD8⁺ T cells in tumor stroma (P = 0.0229). The overall survival rate of the patients with higher B7-H4 expression was significantly worse than that of the patients with lower B7-H4 expression (P = 0.0105, Hazard Ratio: 1.854, 95%CI:1.152–2.902). Markers of cell-mediated immune responses such as CD3, CD8, and T-bet were associated with better patient survival. The present study demonstrated that B7-H4 expression in human ESCC is associated with cancer progression, reduced tumor immunosurveillance and worse patient outcomes. B7-H4 can serve as a novel prognostic predictor for human ESCC and a potential target for the immune therapy against this malignancy.     

T cell augments the antitumor activity of tumor-targeting <Emphasis Type="Italic">Salmonella</Emphasis>

Systemic administration of Salmonella to tumor-bearing mice leads to preferential accumulation within tumor sites and retardation of tumor growth. However, the detailed mechanism of Salmonella-induced antitumor immune response via host T cell remains uncertain. Herein, we used wild-type, CD4⁺ T-cell-deficient, and CD8⁺ T-cell-deficient mice to study the role of T cell in the antitumor immune responses induced by Salmonella enterica serovar Choleraesuis (Salmonella Choleraesuis). When systemically administered into mice bearing tumors, Salmonella Choleraesuis significantly inhibited tumor growth by 50%. In contrast, in T-cell-deficient mice, there was only 34–42% inhibition of tumor growth. We found that treatment with Salmonella Choleraesuis significantly upregulates interferon-γ in wild-type and CD8⁺ T-cell-deficient mice, but not in CD4⁺ T-cell-deficient mice. Furthermore, immunohistochemical staining of the tumors revealed more infiltration of macrophages and neutrophils in wild-type mice after Salmonella Choleraesuis treatment compared with those in T-cell-deficient mice. The antitumor therapeutic effect mediated by Salmonella Choleraesuis is associated with an inflammatory immune response at the tumor site and a tumor T helper 1-type immune response. In conclusion, these results suggest that tumor-targeted therapy using Salmonella Choleraesuis, which exerts tumoricidal effects and stimulates T cell activities, represents a potential strategy for the treatment of tumor.