抗人膀胱癌抗独特型抗体佐剂疫苗的动物实验研究

目的 探讨抗独特型抗体佐剂疫苗对膀胱癌的免疫治疗作用。方法 用抗人膀胱癌抗独特型抗体佐剂疫苗(Ab2 SAF)免疫BALB／C小鼠，即实验组；对照组用生理盐水，方法同实验组。免疫3次后，于末次免疫后1w，将新鲜人膀胱移行细胞癌组织移植于小鼠肾包度下，分别于移植后第2、4、6、8和10天处死小鼠，取血清进行抗抗独特型抗体(Ab3)分析。移植瘤行组织学检查，观察宿主淋巴细胞浸润情况及瘤细胞可见率。结果 实验组小鼠肾脏包膜下人膀胱癌细胞系很快受到排斥，在移植后第6天，淋巴细胞浸润即达高峰，而对照组在第10天才达高峰；癌细胞可见率，在移植后第6天即明显降低，而对照组呈逐渐下降。实验组Ab3为阳性，而对照组阴性。结论 Ab2作为抗原免疫小鼠后，通过诱发或激活对人膀胱癌特异性体液和细胞免疫反应，排斥癌细胞并抑制其生长。     

抗人膀胱癌/抗血管内皮生长因子双功能基因抗体的动物实验研究

目的 探讨抗人膀胱癌／抗人血管内皮生长因子(VEGF)双功能基因抗体对膀胱癌生长的抑翻作用。方法 应用双功能基因抗体作用于荷瘤动物模型观察抑瘤作用，应用免疫组化技术检测体内生长肿瘤中截血管密度。结果 双功能基因抗体对体内生长的肿瘤有明显的抑制作用，经双功能基因抗体治疗的瘤体中截血管密度明显低于对照组。结论 双功能基因抗体能够抑制肿瘤血管生成，从而为膀胱癌的治疗提供一种新的辅助方法。     

抗人膀胱癌/抗VEGF双功能基因抗体对血管内皮细胞增殖的影响

目的 研究抗人膀胱癌／抗VEGF双功能基因抗体体外抗血管内皮细胞增殖的活性。方法 MTT法检测抗人膀胱癌／抗VEGF双功能基因抗体对血管内皮细胞增殖的活性。结果 抗人膀胱癌／抗VEGF双功能基因抗体可以明显抑制血管内皮细胞增殖。结论 抗人膀胱癌／抗VEGF双功能基因抗体能通过抗肿瘤新生血管形成而发挥抗肿瘤作用，从而为膀胱癌的治疗提供了一种新的思路。     

抗人膀胱癌抗独特型抗体的制备和鉴定

目的：制备抗人膀胱癌抗独特型抗体（Ab2)并进行体外实验和鉴定，探讨通过免疫效应治疗或预防膀胱癌术后复发的可能性。方法：提取膀胱癌抗原（Ag)，应用膀胱癌抗原通过杂交瘤技术制备抗人膀胱癌单克隆抗体（Ab1)，Ab1经胃蛋白酶水解制备Ab1的F（ab‘)2片优，以F（ab‘）2片段免疫新西兰白兔，其血清经凝胶纯化后获抗人膀胱癌抗独特型抗体（Ab2），采用ELISA等方法进行鉴别。结果：Ab2与Ab1呈阳性反应，与BALB／C小鼠γ球蛋白呈阴极反应；Ab2与Ag竞争结合Ab1；Ab2与Ag的结合。结论：Ab1是针对膀胱移行细胞癌的单克隆抗体，Ab2是具有膀胱癌抗原的抗独特型抗体。     

抗人膀胱癌/抗CD3双功能基因抗体对LAK细胞增殖和细胞毒作用的体外研究

目的　探讨抗人膀胱癌 /抗 CD3双功能基因抗体对 LAK细胞增殖和增强细胞毒的作用。方法　采用 1 2 5I- Ud R释放试验等检测方法。结果　双功能抗体显著提高 LAK细胞与肿瘤细胞结合率 ,促进淋巴细胞增殖 ,加入双功能抗体后 ,L AK细胞对肿瘤细胞细胞毒性明显增加。结论　抗人膀胱癌 /抗 CD3双功能基因抗体可以大大促进 LAK细胞增殖和增加细胞毒性作用 ,为膀胱癌的治疗提供新的方法。     

抗人膀胱癌/抗VEGF双功能基因抗体的免疫电镜定位研究

目的 对抗人膀胱癌／抗VEGF双功能基因抗体进行免疫定位研究。方法 采用免疫组化和免疫电镜前染色法进行观察。结果 膀胱癌组90例，抗原检测阳性率86．7％(78／90)；对照组30例，抗原检测阳性率为6．67％(2／30)。经免疫组化染色后，电镜观察膀胱癌抗原阳性表达部位为细胞膜或细胞浆中的颗粒，细胞模性结构、线粒体、粗面内质网可见到阳性反应沉积物，电子密度增浓，线粒体肿胀、变性、微管系统变化明显。膀胱癌组抗原阳性率明显高于对照组，电镜观察细胞内改变明显。结论 抗人膀胱癌／抗VEGF双功能抗体对于膀胱癌高危人群的筛选以及膀胱癌的早期诊断具有重要临床意义，值得深入研究。     

抗人膀胱癌/抗VEGF双功能基因抗体的动物实验研究

目的探讨抗人膀胱癌/抗VEGF双功能基因抗体对人膀胱癌生长及淋巴转移的影响.方法通过构建人膀胱癌裸鼠皮下移植瘤模型并注射双功能抗体,观察肿瘤生长及盆腔淋巴结转移情况,同时采用免疫组化法检测肿瘤微血管密度值及凋亡的肿瘤细胞指数.结果肿瘤大小:实验组为(19.50±4.51),对照组为(57.62±8.31),两组比较差异有极显著性意义(P<0.01);肿瘤微血管密度:实验组为(2 351±207),对照组为(4 356±548),两组相比差异具有显著性意义(P<0.05);凋亡指数:实验组为19.25,对照组为9.31,两者比较,差异有统计学意义(P<0.05);盆腔转移:实验组无一只发生,而对照组为75.0%,两组比较,差异有极显著性意义(P＜0.01).结论抗人膀胱癌/抗VEGF双功能基因抗体对人膀胱癌具有良好的靶向性,能够通过抑制肿瘤微血管形成和加速肿瘤细胞凋亡,遏制实验性人膀胱癌的生长转移,为该抗体用于临床膀胱癌的治疗提供了一定的实验依据.     

钙纳米颗粒作为血吸虫病抗独特型抗体疫苗佐剂的研究

目的 探索钙（Ca）纳米颗粒作为日本血吸虫病抗独特型抗体NP30疫苗佐剂的可行性。方法 将钙纳米颗粒与NP30制备成Ca-NP30结合物（Ca-NP30)，主动免疫BALB/c小鼠，观察其对小鼠的保护性作用并探讨其免疫保护机制。结果 Ca纳米颗粒可增强NP30对宿主的保护性作用，减虫率明显提高，从单用NP30的30.4%提高到57.8%；血清特异性抗体IgG水平较对照组显著升高；足垫试验可引发迟发型变态反应。结论 Ca纳米颗粒可作为血吸虫病抗独特型抗体NP30疫苗的佐剂，其作用机制与同时引起宿主体液免疫和细胞免疫应答增强有关。     

中药用于血吸虫病抗独特型抗体疫苗佐剂的研究

目的　从中药中筛选血吸虫病抗独特型抗体疫苗的佐剂 ,并初步探讨其作用机制。方法　日本血吸虫抗独特型抗体 NP30主动免疫 BAL B/ c小鼠 ,分别联用虫草多糖、猪苓多糖、黄芪、甘草酸和人体免疫调理剂 (HIM) ,观察其对小鼠的保护作用 ,并检测小鼠血清特异性 Ig G、Ig G1和 Ig G2 a抗体水平。结果　黄芪和 HIM能明显提高 NP30对小鼠的保护性免疫作用 ,减虫率显著提高 ,从单用 NP30的 37.8%分别提高到 5 0 .6 %和 5 3.6 % ;但两者对血清特异性 Ig G、Ig G1和 Ig G2 a抗体均无明显升高作用。结论　黄芪和 HIM可明显提高血吸虫病抗独特型抗体疫苗 NP30的保护性免疫力 ,可作为 NP30的佐剂 ,其佐剂作用与所检测的抗体水平似不相关 ,推测可能与细胞免疫应答有关。     

抗人膀胱癌/抗VEGF双功能基因抗体对原位种植人膀胱癌的影响

目的 研究抗人膀胱癌/抗VEGF双功能基因抗体对裸鼠原位种植人膀胱癌生长和转移的抑制作用.方法 建立人膀胱癌裸鼠皮下移植瘤模型,观察双功能基因抗体对肿瘤生长和转移的影响,并计数肿瘤内微血管密度.结果 用药组与对照组比较,肿瘤生长和转移显著减慢和延缓,肿瘤内部微血管密度亦明显低.结论 抗人膀胱癌/抗VEGF双功能基因抗体能够通过抑制肿瘤微血管的形成来遏制人膀胱癌的生长和转移,从而为该抗体进一步体内试验和临床应用提供了实验基础.     

Anti-idiotypic antibody and recombinant antigen vaccines in colorectal cancer patients.

The colorectal carcinoma (CRC)-associated GA733 antigen (also known as CO17-1A, KS1-4, KSA or EpCAM) has been the target of a phase II/III randomized trial of passive immunotherapy with monoclonal antibody CO17-1A and phase I active immunotherapy trials with polyclonal anti-idiotypic antibodies mimicking the CO17-1A or GA733 epitope on the antigen. The CO17-1A antigen was molecularly cloned and the extracellular domain expressed in baculovirus (BV) GA733-2E. Whereas, anti-idiotypic antibody mimics a single epitope on the antigen, BV GA733-2E expresses multiple potentially immunogenic epitopes. In animals, the immunogenicity of BV GA733-2E in aluminum hydroxide was superior to that of anti-idiotype in the same adjuvant. Here, we compared the immunogenicity of anti-idiotypic antibody and GA733-2E antigen in CRC patients. These studies indicate that the antigen is superior to the anti-idiotype antibody in inducing humoral and cellular immunity in CRC patients.     

Immune responses to transgene and retroviral vector in patients treated with ex vivo-engineered T cells

Adoptive transfer of immune effector cells that are gene modified by retroviral transduction to express tumor-specific receptors constitutes an attractive approach to treat cancer. In patients with metastatic renal cell carcinoma, we performed a study with autologous T cells genetically retargeted with a chimeric antibody receptor (CAR) directed toward carbonic anhydrase IX (CAIX), an antigen highly expressed in renal cell carcinoma. In the majority of patients, we observed distinct humoral and/or cellular anti-CAIX-CAR T-cell immune responses in combination with a limited peripheral persistence of transferred CAIX-CAR T cells in the majority of patients. Humoral immune responses were anti-idiotypic in nature and neutralized CAIX-CAR-mediated T-cell function. Cellular anti-CAIX-CAR immune responses were directed to the complementarity-determining and framework regions of the CAR variable domains. In addition, 2 patients developed immunity directed against presumed retroviral vector epitopes. Here, we document the novel feature that therapeutic cells, which were ex vivo engineered by means of transduction with a minimal γ-retroviral vector, do express immunogenic vector-encoded epitopes, which might compromise persistence of these cells. These observations may constitute a critical concern for clinical ex vivo γ-retroviral gene transduction in general and CAR-retargeted T-cell therapy in particular, and underscore the need to attenuate the immunogenicity of both transgene and vector.     

Sendai virus-specific T-cell clones: induction of cytolytic T cells by an anti-idiotypic antibody directed against a helper T-cell clone.

We used a monoclonal anti-idiotypic antibody that is directed against a Sendai virus-specific helper T-cell clone to induce an anti-viral immune response in vivo. Splenocytes primed with the anti-idiotypic antibody mediated an antigen-specific cytolytic response. Preimmunization of mice with the anti-idiotypic antibody resulted in protection against a subsequent lethal infection with Sendai virus.     

Modulation of anti-idiotypic immune response by immunization with the autologous M-component protein in multiple myeloma patients

Multiple myeloma is characterized by a proliferation of clonal B lymphocytes and plasma cells. The idiotypic structure of clonal immunoglobulin (Ig) expressed on the tumour B-cell surface can be regarded as a tumour-specific antigen and, as such, a potential target for anti-idiotypic T and B cells in an immune regulation of the tumour-cell clone. Active immunization using the autologous monoclonal Ig as a ‘vaccine’ was shown to induce tumour-specific immunity in murine B-cell tumours and in human B-cell lymphoma. With the aim to induce or amplify an anti-idiotypic response in multiple myeloma, five stage I–III patients were repeatedly immunized with the autologous monoclonal IgG. Induction of idiotype-specific cellular immunity was analysed in vitro by an enzyme-linked immunospot assay (interferon-γ and interleukin-4 secreting cells). B cells secreting anti-idiotypic IgM antibodies were also analysed. An anti-idiotypic T-cell response was amplified 1.9–5-fold in three of the five patients during immunization. The number of B cells secreting anti-idiotypic antibodies also increased in these three patients. In two of the patients induction of idiotype-specific immunity was associated with a gradual decrease of blood CD19⁺ B cells. The induced T-cell response was eliminated during repeated immunization. Further studies are warranted to optimize the immunization schedule in order to achieve a long-lasting T-cell immunity against idiotypic determinants on the tumour clone. A role for immunity in controlling the tumour clone remains to be established.     

Modulation of anti-idiotypic immune response by immunization with the autologous M-component protein in multiple myeloma patients

Multiple myeloma is characterized by a proliferation of clonal B lymphocytes and plasma cells. The idiotypic structure of clonal immunoglobulin (Ig) expressed on the tumour B-cell surface can be regarded as a tumour-specific antigen and, as such, a potential target for anti-idiotypic T and B cells in an immune regulation of the tumour-cell clone. Active immunization using the autologous monoclonal Ig as a 'vaccine' was shown to induce tumour-specific immunity in murine B-cell tumours and in human B-cell lymphoma. With the aim to induce or amplify an anti-idiotypic response in multiple myeloma, five stage I–III patients were repeatedly immunized with the autologous monoclonal IgG. Induction of idiotype-specific cellular immunity was analysed in vitro by an enzyme-linked immunospot assay (interferon-γ and interleukin-4 secreting cells). B cells secreting anti-idiotypic IgM antibodies were also analysed. An anti-idiotypic T-cell response was amplified 1.9–5-fold in three of the five patients during immunization. The number of B cells secreting anti-idiotypic antibodies also increased in these three patients. In two of the patients induction of idiotype-specific immunity was associated with a gradual decrease of blood CD19⁺ B cells. The induced T-cell response was eliminated during repeated immunization. Further studies are warranted to optimize the immunization schedule in order to achieve a long-lasting T-cell immunity against idiotypic determinants on the tumour clone. A role for immunity in controlling the tumour clone remains to be established.     

Suppression of a "recurrent" idiotype results in profound alterations of the whole B-cell compartment.

The progeny of BALB/c female mice actively immunized with the trinitrophenyl-binding myeloma protein MOPC460 and producing anti-idiotypic antibodies during pregnancy were compared with mice born of normal mothers for several characteristics of B lymphocytes and their precursors. In all cases, maternal anti-idiotypic immunity resulted in the suppression of the expression of that idiotype by immunocompetent cells in the progeny, as shown by limiting-dilution analysis in single clones of mitogen-reactive IgM-secreting cells. At critical concentrations of circulating maternal antibodies, suppression of the antibody idiotype was found to be accompanied by a large increase in the total number of mature small B lymphocytes. This increase can be accounted for by the selective expansion of B cells bearing nonimmunoglobulin surface structures crossreactive with a MOPC460 idiotope recognized by a monoclonal antibody. In addition, the large majority of newly formed mature B lymphocytes, as well as a large fraction of immunoglobulin-negative cells in the bone marrow of suppressed mice, bear such nonimmunoglobulin MOPC460 crossreactive determinant(s). These results suggest that the suppression of a given "recurrent" idiotype has profound consequences for a large part of the immune system.     

Induction of opsonic antibodies to Pseudomonas aeruginosa mucoid exopolysaccharide by an anti-idiotypic monoclonal antibody.

Mucoid strains of Pseudomonas aeruginosa are the major pulmonary pathogens for cystic fibrosis patients. Opsonizing antibodies to the mucoid exopolysaccharide (MEP) antigen may protect animals and some cystic fibrosis patients from infection. However, MEP does not readily elicit opsonic antibodies either during chronic infection or after vaccination. To evaluate alternative means to induce opsonic antibodies, a murine monoclonal anti-idiotypic antibody directed to an opsonic monoclonal antibody specific to MEP was produced. The anti-idiotypic antibody bound to F(ab')2 fragments of the opsonic antibody, blocked binding to MEP, bound to cross-reactive idiotopes on human opsonic antibodies to MEP, and elicited MEP-specific antibodies in syngeneic and allogeneic mice. These anti-idiotype-induced, MEP-specific antibodies fixed complement to mucoid P. aeruginosa cells and opsonized them for phagocytic killing by human leukocytes. These studies demonstrate the potential utility of anti-idiotypic monoclonal antibody for generating protective immunity against bacterial polysaccharides.