ANTIIDIOTYPE VACCINE THERAPY OF HUMAN COLORECTAL CANCER

我们已经产生了一种小鼠抗独特型抗体，3HL，即癌胚抗原(CEA)的内图像。我们已经证明，晚期转移性结直肠癌(CRC)患者和高危结直肠癌患者在手术后辅助性环境中，在足够数量的Chl抗ID吸附到氢氧化铝上进行免疫后，可以产生针对CEA的主动免疫反应。我们证明了一种以免疫球蛋白G为主的多克隆体液免疫反应，与纯化的CEA和CEA阳性细胞特异性结合，介导抗体依赖的细胞毒性。接种疫苗的患者在抗ID3HL、CEA或3H1和CEA合成肽的存在下，也表现出体外T细胞增殖反应。其中一名患者对3HL有客观的临床反应，几名患者继续接受疫苗治疗，病情稳定12-36个月。毒性仅限于注射部位的局部反应，并伴有轻微发烧。在这个项目中，我们将进行临床试验，将佐剂5FU和亚叶酸钙和抗ID3HL联合不同的免疫佐剂，如GS-21，GM-CSF或小剂量IL-2治疗Dukes B和C期结直肠癌患者。将观察患者的免疫反应和临床结果。C57BL/6小鼠移植人CEA转基因小鼠结肠癌细胞株MC38的动物模型将被用来建立更有效的利用抗ID3HL衍生物的免疫方法，并对其抗肿瘤免疫机制进行剖析。我们将研究3HL的单链抗体分子(ScFv)作为一种免疫原，与多种佐剂和细胞因子混合。我们将用相关的抗独特型试剂冲击树突状细胞(DC)，探讨其在肿瘤免疫中的作用。此外，将使用腺病毒载体(ADV)将3H1-scFv或CEA导入DC，继而将其用于疫苗潜力测试。我们还将从结直肠癌患者的白细胞分离产品中分离出具有潜在治疗性的DC(其中一些可能经过3H1疫苗处理)，并用抗ID 3HL、CEA或相关多肽冲击它们，或将表达这些基因的ADV转导它们，并在体外检测它们的抗原提呈和T细胞刺激功能或T细胞杀伤功能。这些研究将是使用基于DC的自体抗ID疫苗和单链抗体对结直肠癌患者进行临床试验的前奏。

We have generated a murine monoclonal anti-idiotype ( antibody, 3Hl, that is the internal image of the carcinoembryonic antigen (CEA). We have demonstrated that patients with advanced metastatic colorectal cancer (CRC) and patients with high risk CRC in the post-surgical adjuvant setting generate an active immune response against CEA following an adequate number of immunizations with the Chl anti-Id adsorbed to aluminum hydroxide. We demonstrated a predominantly IgG polyclonal humoral immune response with specific binding to purified CEA and CEA positive cells that mediated antibody-dependent cellular cytotoxicity. The vaccinated patients also demonstrated in vitro T cell proliferative responses in the presence of anti-Id 3Hl, CEA or synthetic peptides derived from 3H1 and CEA. While one of the patients had objective clinical response to 3Hl, several continue on vaccine therapy with stable disease from 12-36 months. Toxicity was limited to local reactions at the site of the injection with mild fevers. In this project, we will conduct clinical trials in which patients with Dukes B and C colorectal carcinoma will be treated with adjuvant 5FU and leucovorin and anti-Id 3Hl in combination with different immunologic adjuvants such as GS-21, GM-CSF or low-dose IL-2. The patients will be observed for immune responses and for clinical outcome. Animal models in C57BL/6 mice transplanted with the human CEA transfected murine colon cancer cell line MC38 will be used to develop more powerful methods of immunization utilizing the anti-Id 3Hl derivatives and to dissect the mechanisms of anti-tumor immunity. We will investigate a single chain Fv molecule (scFv) of 3Hl as an immunogen mixed with variety of adjuvants and cytokines. We will investigate the role of dendritic cell (DC) in developing tumor immunity after pulsing them with relevant anti-idiotypic reagents. In addition, adenoviral vectors (AdV) will be used to introduce 3H1-scFv or CEA into DC, which in turn will be tested for vaccine potential. We will also generate potentially therapeutic DC from leukapheresis products of CRC patients (some of them may be pretreated with 3H1 vaccine), and pulse them with the anti-Id 3Hl, CEA or relevant peptides or transduce them with Adv expressing these genes and examine their antigen presenting and T cell stimulatory function or T cell cytolytic function in vitro. These studies will be a prelude to clinical trials for CRC patients with autologous DC based anti-Id vaccine and scFv.