Synergistic Suppressive Effect of Double Transfection of Tumor Necrosis Factor-α and Interleukin 12 Genes on Tumorigenicity of Meth-A Cells

Tumor necrosis factor-α(TNF-α) and interleukin 12 (IL-12), both potent antitumor cytokines, are known to be involved in the host's antitumor immune surveillance in tumor bearers, via different mechanisms. The former enhances the activities of dendritic cells, natural killer/lymphocyteactivated killer (NK/LAK) and cytotoxic T lymphocyte (CTL), while the latter induces Th1-type cellular immunity and enhances the activities of natural killer T (NKT), NK/LAK and CTL. In the present study, in the expectation of synergistic actions of these cytokines in stimulating the host's immune responses, we investigated the feasibility of a cancer vaccine involving double transfection with both genes in a murine model. The expression of major histocompatibility complex (MHC) class I, class II and B7.1 on the surface of the double transfectants was enhanced as revealed by FACS analysis. A significant decrease in tumorigenicity was observed in mice inoculated with the double transfectants. Cytotoxicity assay revealed that the activities of NK/LAK and CTL from spleens of mice bearing the double transfectants were enhanced. The induction of tumor-specific immunity was confirmed by rechallenge with parental Meth-A cells in mice that had rejected the double transfectants. Thus, double transfection of TNF-α and IL-12 genes was considered to bring about synergistic suppressive effects on the tumorigenicity of transfectants through the activation of killer cells by produced cytokines and the enhancement of expression of MHC class I, II and B7.1 molecules.     

Preparation of NK-enriched LAK cells--their potential cytotoxic and ADCC activities

We examined several culture methods to induce proliferation of natural killer (NK) cells from peripheral blood mononuclear cells (PBMC). In the presence of IL-2, a remarkable proliferation of NK cells was observed when PBMC were co-cultured with MMC-treated K562, which is known as a highly sensitive in vitro target cell for the NK assay. Addition of OK-432 or TNF-alpha and IL-1 beta also induced marked NK proliferation in a dose dependent manner. These NK-enriched lymphokine activated killer (LAK) cells showed highly cytotoxic activities against various MHC class I positive or negative tumor cells. They also showed potent ADCC activities against Herceptin-coated SK-BR-3, a HER2/neu positive breast cancer cell line. These results indicated that NK-enriched LAK cells are potent effector cells, and suggested novel therapeutic strategies for nonspecific immunotherapy as well as target immunotherapy in combination with anticancer antibodies, such as Herceptin.     

Natural killer cell-dependent suppression of systemic spread of human lung adenocarcinoma cells by monocyte chemoattractant protein-1 gene transfection in severe combined immunodeficient mice

Monocyte chemoattractant protein-1 (MCP-1) is a chemokine with various biological activities, including augmentation of cytotoxic activity of monocytes and natural killer (NK) cells. The present study was undertaken to determine whether transfection of the MCP-1 gene into lung cancer cells affected their tumorigenicity and metastatic potential by the NK cell-mediated mechanism. The human MCP-1 gene inserted into an expression vector (BCMGSNeo) was transfected into human lung adenocarcinoma (PC-14) cells. There was no difference in in vitro proliferation between MCP-1 gene-transfected PC-14 cells and the parent cells or mock-transfected cells. The tumorigenicity and in vivo tumor growth of MCP-1 gene-transfected PC-14 cells were similar to those of the parent cells or mock-transfected cells when tumor cells were injected into the s.c. space of NK cell-intact severe combined immunodeficient (SCID) mice. Although parent cells and mock-transfected cells inoculated i.v. formed lung metastatic colonies and pleural effusion, MCP-1 gene transfectants reduced the systemic spread in NK cell-intact SCID mice. Interestingly, these modulations in a systemic spread by MCP-1 gene transfection were not observed in NK cell-depleted SCID mice. Decreased survival of MCP-1 gene transfectants in the lung was observed in NK cell-intact SCID mice but not in NK cell-depleted SCID mice. Recombinant MCP-1 or the supernatant of MCP-1 gene transfectants enhanced the cytotoxicity of human CD56+ NK cells and spleen cells of SCID mice against PC-14 cells. These findings suggest that locally produced MCP-1 suppresses tumor progression by a NK cell-mediated mechanism, depending on organ microenvironment.     

Susceptibility of rat colon carcinoma cells to lymphokine activated killer-mediated cytotoxicity is decreased by alpha1,2-fucosylation

The presence of alpha1,2-fucosylated glycans at the surface of rat colon carcinoma cells has been associated with an increased tumorigenicity and resistance to natural killer/lymphokine activated killer (NK/LAK) cytotoxicity. We now report that transfection of rat alpha1,2-fucosyltransferases cDNA (FTA and FTB) into REG cells, which are spontaneously devoid of this enzymatic activity, allows expression of histo-blood group H antigen and increases their resistance to LAK, but not NK cell lysis. Conversely, transfection of PRO cells, which spontaneously express alpha1, 2-fucosyltransferase activity, with the FTA cDNA in the antisense orientation decreases expression of the H antigen together with their resistance to LAK cell lysis, but again, not to NK cell lysis. Furthermore, REG cells that are rejected by immunocompetent syngeneic rats are similarly rejected by rats depleted of NK cells by antibody 3.2.3, directed against the NKR-P1 molecule. Thus, the rejection of REG cells by immunocompetent rats and their earlier reported increased tumorigenicity after transfection with an alpha1, 2-fucosyltransferase cDNA cannot be ascribed to NK cell sensitivity or resistance, respectively. The increased resistance to LAK cell lysis, however, may be relevant to tumor progression.     

AK细胞及环磷酰胺治疗肝癌的实验研究

为研究１ 种新的过继免疫化疗治疗肝癌的方法,采用肝癌细胞株Ｈ２２ 接种于近交系Ｂａｌｂ／ｃ 小鼠皮下,制成肿瘤模型。使用ＩＬ２ 、肿瘤活化的杀伤细胞（ＡＫ细胞）及环磷酰胺（Ｃｙ） 进行治疗,检测小鼠脾淋巴细胞ＮＫ、ＬＡＫ及ＣＴＬ 活性,用流式细胞仪检测Ｌ３Ｔ４ 亚群及Ｌｙｔ２ 亚群含量。结果表明在过继免疫化疗组小鼠ＬＡＫ 及ＣＴＬ活性明显高于其它治疗组（Ｐ＜ ０．０１）,荷瘤小鼠肿瘤结节生长较ＩＬ２ 组及Ｃｙ 组明显缓慢（Ｐ＜ ０．０１）,生存期也明显高于其它各治疗组（Ｐ＜０ ．０１）。该研究表明过继免疫化疗有较强的抗肿瘤作用。     

ANTITUMOR EFFECTS OF HUMAN IL-15 GENE MODIFIED LUNG CANCER CELL LINE

ＡＮＴＩＴＵＭＯＲＥＦＦＥＣＴＳＯＦＨＵＭＡＮＩＬ－１５ＧＥＮＥＭＯＤＩＦＩＥＤＬＵＮＧＣＡＮＣＥＲＣＥＬＬＬＩＮＥＳｈｅｎＹｏｎｇｑｕａｎ１沈永泉ＣｕｉＬｉａｎｘｉａｎ２崔莲仙ＨｅＷｅｉ１何维ＸｕｅＬｉ１薛莉ＢａＤｅｎｉａｎ１巴德年１Ｉｎｓｔ...     

Cytokine-induced killer cells: NK-like T cells with cytotolytic specificity against leukemia

Cytokine-induced killer (CIK) cells are a unique population of cytotoxic T lymphocytes (CTL) with the characteristic CD3+CD56+ phenotype. These cells have demonstrated higher proliferative and cytolytic activities in comparison to the reported CD3-CD56+ lymphokine activated killer (LAK) cells that are essentially activated natural killer (NK) cells. CIK cells are non-MHC-restricted in target cell recognition and killing. We have shown the feasibility of generating CIK cells from a series of marrow samples of patients with acute myeloid leukemia (AML) collected at diagnosis. At maturity, the CIK cells exhibit potent cytotoxicity against autologous AML targets as well as allogeneic myeloid leukemia cells, regardless of the HLA types of these targets. This observed cytotoxicity is not entirely due to NK cells as prior pre-absorption of the NK cells cytolytic activities does not abolish the subsequent cytotolytic activities against leukemic targets. It has also been reported by others that CIK cells are cytolytic against chronic myeloid leukemia (CML) cells, both in vitro and in the SCID mouse tumor model. In a mouse transplant model across MHC barrier, the CIK cells generated from the donor do not induce graft vs. host disease as observed for unfractionated donor splenocytes. In comparison to untreated control mice, the infusion of CIK cells results in the prolonged survival of murine leukemia-bearing mice. CIK cells also express CD94, part of the NK receptor comprising of CD94-NKG2 heterodimer. However, only low level of the killer immunoglobulin-like receptors are expressed by the CIK cells. In addition, as reported for the classical CTL, CIK cells could interact with dendritic cells (DC) to result in the enhancement of cytotolytic activities against tumor cells. The characteristic biological properties of the CIK cells would, therefore, enable them to be exploited for anti-leukemic therapy.     

Retaliation against tumor cells showing aberrant HLA expression using lymphokine activated killer-derived T cells.

Lymphokine activated killer (LAK) cells represent mixtures of natural killer (NK) and non-MHC-restricted CTLs that have the capacity to lyse a variety of tumor cells and MHC class I-negative target cells. Although it is clear that NK cells are negatively regulated by interactions with MHC class Ia or class Ib molecules, the regulation of LAK-derived T cells has not been clarified to date. In the studies presented here, we demonstrate that IFN-gamma treatment of tumor cells can induce their resistance to LAK-derived T cells in a manner similar to that seen for NK cells. The IFN-gamma-mediated suppression of LAK activity correlates with increased MHC class I expression by the tumor cells, and the inhibition of LAK-mediated cytotoxicity can be reversed in the presence of class I-specific antibody. Furthermore, the expression of MHC class Ia or class Ib molecules in class I-negative cell lines can reduce their susceptibility to LAK-mediated cytotoxicity. This principle of negative regulation by MHC class I molecules applies to LAK-derived T cells generated from peripheral blood lymphocytes of renal cell carcinoma patients and healthy, control donors. Although LAK-derived T cells can be inhibited in their lytic activity through interactions with MHC class Ia and class Ib molecules, they do not express the known inhibitory receptors specific for these ligands that are found on NK cells. Apparently, LAK-derived T cells are negatively regulated by as yet undefined inhibitory receptors.     

Overexpression of interleukin-15 in vivo enhances antitumor activity against MHC class I-negative and -positive malignant melanoma through augmented NK activity and cytotoxic T-cell response

Interleukin (IL)-15, a pleiotropic cytokine, is involved in the development and maintenance of NK cells and memory CD8+ T cells. We examined the effects of in vivo overexpression of IL-15 on protection against 2 types of murine B16 melanoma lines, MHC class I-negative B16.44 and MHC class I-positive B16F10 cells, using IL-15 transgenic (Tg) mice that we have recently constructed. The tumor growth was severely retarded in IL-15 Tg mice after subcutaneous (s.c.) inoculation with B16.44 or B16F10 cells. IL-15 Tg mice showed an augmented NK cell activity against B16.44 cells, and in vivo depletion of NK cells by anti-asialoGM1 Ab treatment abrogated the antitumor activity in IL-15 Tg mice. On the other hand, IL-15 Tg mice inoculated with B16F10 cells developed a significant level of CTL response against B16F10 cells, and in vivo depletion of CD8+ T cells by anti-CD8 MAb treatment abrogated the antitumor activity. Thus, overexpression of IL-15 augmented antitumor activity against different tumors via augmentation of different antitumor mechanisms. These results suggest a possible therapeutic application of IL-15 for human neoplasms expressing a wide range of MHC class molecules.     

Combined immunogene therapy of IL-6 and IL-15 enhances anti-tumor activity through augmented NK cytotoxicity

Many tumors evade host immunity by lowering expression of major histocompatibility complex (MHC) molecules. Theoretically, low MHC expression should activate natural killer (NK) cells and in some cases suppress tumor growth; nevertheless, some tumors also produce high concentrations of immunosuppressive cytokines, such as transforming growth factor (TGF)-beta, to inhibit the activity of NK cells. Using a canine transmissible venereal tumor (CTVT) model, we have previously demonstrated that IL-6 is a strong antagonist for TGF-beta. Herein, we found that IL-6 alone was unable to significantly promote TGF-beta-inhibited NK activities. Conversely, IL-15 alone strongly promoted NK activities; however, NK activities were inhibited to baseline levels following the addition of TGF-beta. Therefore, a new strategy using combined immunogene therapy of both IL-6 and IL-15 mediated by electroporation was used in this study. This combined IL-6 and IL-15 treatment effectively relieved the inhibitory effect of TGF-beta and activated NK cell cytotoxicity of lymphokine-activated killer (LAK) cells. Similarly, in isolated DX5+ NK cells, only IL-6 and IL-15 in combination significantly overcame the inhibitory effect of TGF-beta and promoted NK cytotoxicity. The group of BALB/c mice injected with plasmids with IL-6 and IL-15 genes (pIL-6/pIL-15) had the highest percentages of DX5+ NK cells as compared with either the pIL-6 or pIL-15 groups. Further, in SCID mice inoculated with CTVT, electroporation-mediated delivery of pIL-6/pIL-15 was significantly more efficient in suppressing both tumor establishment and tumor growth as compared with pIL-6 or pIL-15 inoculation alone. In addition, the anti-asialo GM-1 antibody abolished NK activities in SCID mice and resulted in outgrowth of the tumors. Together, these results suggest that the TGF-beta-associated inhibition of NK cytotoxicity cannot be adequately restored by simply antagonizing TGF-beta with IL-6: the co-existence of NK activating factors such as IL-15 is also important in restoring TGF-beta-inhibited cytotoxicity. This study highlights the therapeutic potential of the pIL-6/pIL-15 combination by inhibiting TGF-beta activity and enhancing NK cytotoxicity.     

The level of MHC class I expression on murine adenocarcinoma can change the antitumor effector mechanism of immunocytokine therapy

The huKS1/4-IL2 fusion protein, directed against the human epithelial cell adhesion molecule (huEpCAM) has been shown to induce a strong CD8+ T-cell-dependent, natural killer (NK) cell-independent, antitumor response in mice bearing the huEp-CAM-transfected CT26 colon cancer CT26-EpCAM. Here we investigate the effectiveness of huKS1/4-IL2 against CT26-Ep21.6, a subclone of CT26-EpCAM, expressing low levels of MHC class I. In vitro antibody-dependent cellular cytotoxicity (ADCC) assays in the presence of huKS1/4-IL2 demonstrate that murine NK cells from spleen and blood can kill CT26-Ep21.6 significantly better than they kill CT26-EpCAM. NK-mediated ADCC of CT26-EpCAM can be enhanced by blocking the murine NK cell-inhibitory receptor, Ly-49C. A potent in vivo antitumor effect was observed when BALB/c mice bearing experimental metastases of CT26-Ep21.6 were treated with huKS1/4-IL2. The depletion of NK cells during huKS1/4-IL2 treatment significantly reduced the antitumor effect against CT26-Ep21.6. Together our in vitro and in vivo data in the huEp-CAM-transfected CT26 models indicate that the amount of MHC class I expressed on the tumor target cell plays a critical role in the in vivo antitumor mechanism of huKS1/4-IL2 immunotherapy. A low MHC class I level favors NK cells as effectors, whereas a high level of MHC class I favors T cells as effectors. Given the heterogeneity of MHC class I expression seen in human tumors and the prevailing T-cell suppression in many cancer patients, the observation that huKS1/4-IL2 has the potential to effectively activate an NK cell-based antitumor response may be of potential clinical relevance.     

腺病毒介导的人GM-CSF基因转染瘤苗增强机体免疫功能的实验研究

目的 研究腺病毒介导的人ＧＭ－ＣＳＦ基因转染瘤苗体内抗肿瘤免疫作用极其机理。方法 应用ＧＭ－ＣＳＦ基因转染、的瘤苗对小鼠肝癌模型进行免疫基因治疗,观察该疗法对荷瘤小鼠脾细胞ＮＫ、ＬＡＫ、ＣＴＬ活性的影响、脾淋巴细胞围化反应的影响以及荷瘤习的生存期。结果 经ＧＭ－ＣＳＦ基因转染瘤苗治疗后,插细胞ＣＴＬ活性显著升高,而ＮＫ、ＬＡＫ细胞活性未见明显增强、脾淋巴细胞转化反应显著增强、生存期显著延长。结论     

In vivo effects of recombinant human interleukin 2 on antitumor and antiviral natural immunity in induced or natural murine immunodeficiency states

We have examined the ability of in vivo treatment of mice with recombinant interleukin 2 (rIL-2) to affect natural immunity measured against tumor (YAC-1) or virally infected (herpes simplex type 1) target cells. rIL-2 treatment leads to significant increases in natural killer/lymphocyte-activated killer (NK/LAK) function and spleen cells recovered. This effect is dose dependent and strain related. The latter parameter correlated with the pretreatment NK activity level of the strain. The rIL-2 induced NK/LAK augmentation is also kinetically restricted as treatment must have occurred within 48-72 h of assay to be effective. The rIL-2 therapy effectively enhances both antitumor and antiviral NK/LAK activity and results in a noticeable increase in asialo-GM1-positive cells in the spleens of treated mice as well as a significant increase in IL-2 receptor expression as monitored by either cytometry or radioligand binding. In vivo treatment of mice with an antibody directed to the ASGM1 determinant effectively reduces the rIL-2 augmentation of both antitumor and antiviral activity even though this treatment does not affect the pretreatment level of antiviral activity. Various natural and induced immunodeficiency states (immunotherapy, irradiation, immunosuppressive drugs, cytoreductive drugs) have been examined for the ability of in vivo treatment with rIL-2 to enhance NK/LAK activity. In vivo rIL-2 administration is differentially effective in enhancing NK/LAK activity in these situations. Notably, in these induced immunodeficiency states, although NK/LAK activity is commonly enhanced, the number of spleen cells recovered often is only marginally affected. Thus, as expected, a limiting aspect in this use of a natural immunomodulator is the number of potentially responsive cells present in the immunodeficiency condition. In addition, correlations between rIL-2 effect, several of the immunodeficiency states, and vascular leak syndrome are briefly discussed.     

Recombinant gamma interferon provokes resistance of human breast cancer cells to spontaneous and IL-2 activated non-MHC restricted cytotoxicity

Natural and lymphokine activated killer cells (NK and LAK) are believed to play an important role in the control of tumour progression and metastasis. Their specific receptors on tumours cells are still unknown. Several studies suggest that these cells recognise and eliminate abnormal cells with deleted or reduced expression of MHC class I molecules. Previous reports suggest that interferons (IFN), by increasing MHC class I expression on target cells, induce resistance to killing by NK cells. We investigated the role of MHC molecule expression by two human breast cancer cell lines T47D and ZR75-1 in their susceptibility to NK and LAK cells. These two cell lines spontaneously express low levels of HLA class I antigens but no HLA class II molecules. After IFN-gamma treatment they both overexpressed MHC class I and de novo expressed class II molecules as detected by flow cytometry, quantified by a radioimmunoassay and analysed by two-dimensional gel electrophoresis. Opposed to untreated cells these IFN-gamma treated cells were resistant to NK and LAK lysis. Furthermore, preincubation of IFN-gamma treated breast cancer cells with F(ab')2 fragments of monoclonal antibodies to HLA class I and HLA class II molecules was unable to restore lysis. In contrast, several complete monoclonal antibodies including anti-HLA class I and HLA class II induced the lysis of target cells whether or not they had been treated by IFN-gamma. The therapeutic use of monoclonal antibodies directed against antigens expressed on tumour cells (ADCC) in conjunction with interferon therapy should be discussed in lymphokine-based strategies for treatment of cancer patients.     

Induction of EBV-latent membrane protein 1-specific MHC class II-restricted T-cell responses against natural killer lymphoma cells

EBV-encoded latent membrane protein 1 (LMP1) has oncogenic potential and is expressed in many EBV-associated malignancies. Although LMP1 is regarded as a potential tumor-associated antigen for immunotherapy and several LMP1-specific MHC class I-restricted CTL epitopes have been reported, little is known regarding MHC class II-restricted CD4 helper T-lymphocyte (HTL) epitopes for LMP1. The goal of the present studies was to determine whether MHC class II-restricted CD4 T-cell responses could be induced against the LMP1 antigen and to evaluate the antitumor effect of these responses. We have combined the use of a predictive MHC class II binding peptide algorithm with in vitro vaccination of CD4 T cells using candidate peptides to identify naturally processed epitopes derived from LMP1 that elicit immune responses against EBV-expressing tumor cells. Peptide LMP1(159-175) was effective in inducing HTL responses that were restricted by HLA-DR9, HLA-DR53, or HLA-DR15, indicating that this peptide behaves as a promiscuous T-cell epitope. Moreover, LMP1(159-175)-reactive HTL clones directly recognized EBV lymphoblastoid B cells, EBV-infected natural killer (NK)/T-lymphoma cells and naturally processed antigen in the form of LMP1+ tumor cell lysates presented by autologous dendritic cells. Because the newly identified epitope LMP1(159-175) overlaps with an HLA-A2-restricted CTL epitope (LMP1(159-167)), this peptide might have the ability to induce simultaneous CTL and HTL responses against LMP1. Overall, our data should be relevant for the design and optimization of T-cell epitope-based immunotherapy against various EBV-associated malignancies, including NK/T cell lymphomas.     

Natural Killer (NK) and Lymphokine-Activated Killer (LAK) Cell Activity in Healthy Volunteers with Special Emphasis on the Familial Incidence of Cancer

Natural killer (NK) and lymphokine activated killer (LAK) cellactivities in 133 healthy volunteers were analyzed with regardto the volunteer's sex, age, smoking history and the familialincidence of cancer. None of these factors had any influence on NK and LAK cell activities.It was concluded that identifying individuals with increasedrisk of cancer development by examining NK and LAK activitieswould be difficult.     

Melanoma cells interfere with the interaction of dendritic cells with NK/LAK cells

Dendritic cells (DCs) and natural killer (NK) cells are key players at the interface between innate resistance and acquired immunity. NK cells can induce DC maturation, a differentiation process whereby DCs respond to a environmental stimulus and acquire the ability of eliciting adaptive immunity. Conversely, maturing DCs promote NK functions in vivo and in vitro. This interplay has important consequences on the immune response to pathogens and possibly to neoplastic cells. Here, we show that B16 melanoma cells actively modulate the interaction between DCs derived from bone marrow precursors and NK/LAK cells propagated from the spleen of C57BL/6 mice. DCs increased in a dose-dependent manner the ability of NK/LAK cells to kill melanoma cells and to produce cytokines. This activatory cross-talk entailed the production of IL-18 by DCs and of IFN-gamma by NK/LAK cells. Melanoma cells were not a passive target of NK activity; they regulated the outcome of the interaction between DCs and NK/LAK cells, inhibiting the in vitro production of cytokines as effectively as the genetic deletion of IL-18 or IFN-gamma. Interference with the NK/DC interaction possibly represents a mechanism used by growing tumors to evade the immune response.     

局部注射小鼠白细胞介素12逆转录病毒包装细胞对大鼠肝癌的治疗

目的:研究低剂量环磷酸胺(Cy)联合MHC Ⅰ类限制性肿瘤抗原多肽Mutl致敏、白细胞介素2(IL-2)基因修饰的树突状细胞(DCs)对转移性肺癌小鼠的治疗作用及其免疫学机理.方法:制备小鼠骨髓来源的DCs,用转移性Lewis肺癌特异性多肽Mutl预激经IL-2基因修饰的DCs联合低剂量Cy治疗转移性肺癌小鼠.通过FACS分析其脾细胞内T淋巴细胞比例的变化,~51Cr释放法检测CTL和NK细胞杀伤活性.结果:肿瘤抗原多肽致敏、IL-2基因修饰的DCs与小剂量Cy联合后,能比单用DCs更有效地治疗转移性肺癌,小鼠脾细胞中CD8~ T细胞和NK1.1~ 细胞明显比例升高,联合治疗组诱导出的CTL杀伤活性最高.结论:以肿瘤抗原多肽冲击致敏的IL-2基因修饰的DCs联合小剂量Cy能更有效地促进荷瘤宿主免疫应答,具有显著地体内抑制肺癌转移的效果.     

低剂量环磷酰胺增强MHCⅠ类限制性肿瘤抗原多肽冲击致敏的树突状细胞的抗肿瘤转移效果

目的:研究低剂量环磷酸胺(Cy)联合MHC Ⅰ类限制性肿瘤抗原多肽Mutl致敏、白细胞介素2(IL-2)基因修饰的树突状细胞(DCs)对转移性肺癌小鼠的治疗作用及其免疫学机理.方法:制备小鼠骨髓来源的DCs,用转移性Lewis肺癌特异性多肽Mutl预激经IL-2基因修饰的DCs联合低剂量Cy治疗转移性肺癌小鼠.通过FACS分析其脾细胞内T淋巴细胞比例的变化,~51Cr释放法检测CTL和NK细胞杀伤活性.结果:肿瘤抗原多肽致敏、IL-2基因修饰的DCs与小剂量Cy联合后,能比单用DCs更有效地治疗转移性肺癌,小鼠脾细胞中CD8~+T细胞和NK1.1~+细胞明显比例升高,联合治疗组诱导出的CTL杀伤活性最高.结论:以肿瘤抗原多肽冲击致敏的IL-2基因修饰的DCs联合小剂量Cy能更有效地促进荷瘤宿主免疫应答,具有显著地体内抑制肺癌转移的效果.     

Augmented Systemic Immunity in Mice Implanted with Tumor Necrosis Factor-α Gene-transduced Meth-A Cells

Syngeneic BALB/c mice bearing methylcholanthrene-induced fibrosarcoma (Meth-A) cells transduced with a tumor necrosis factor (TNF) gene showed a longer life span and tumor regression as compared with mice carrying TNF-non-producing Meth-A cells. To elucidate the mechanism of the reduced tumorigenicity of TNF-producing Meth-A, we compared systemic immune responses between mice bearing high TNF producer (C5) and unmodified Meth-A cells (M0). The results indicated that the cytotoxic activity of lymphokine-activated killer cells (LAK) induced from spleen cells of mice bearing C5 was higher against both M0 and C5 than that of LAK from mice bearing M0. Also, C5 was more sensitive to LAK induced from spleen cells of C5- and M0- bearing mice than M0. We also found that cytotoxic T lymphocyte from spleen cells of mice transplanted with C5 was more cytotoxic to M0 than that from mice with M0. In addition, the population of Lyt2 (CD8)-positive T cells was higher in freshly isolated spleen cells of mice transplanted with C5 than from mice with M0. Finally, we observed a higher expression of MHC class 1 antigen on C5 than on M0. These observations suggest that the augmented host systemic immunity in mice carrying TNF gene-modified Meth-A cells is one of the mechanisms of the reduced tumorigenicity of C5 and that the increased systemic immunity can be ascribed to the increased immunogenicity of the tumor cells. Thus, the use of TNF gene-modified tumor cells as vaccines appears to be promising for therapeutic and/or prophylactic application.