Enhancing the immunogenicity of idiotypes for therapy of Non-Hodgkin's lymphoma

描述（由申请人提供）：非霍奇金淋巴瘤（NHL）的特点是临床病程缓慢，中位生存期为6-10年。在美国，NHL的发病率每年增加约2.5%。显然，全国需要改进这种恶性肿瘤的治疗方法。基于独特型的卵泡性非霍奇金淋巴瘤微小残留病的临床试验显示，无病生存率和分子缓解都有所增加。目前，在随机III期临床试验中，最好的基于独特型的疫苗使用KLH作为蛋白质载体偶联到B细胞受体（BCR）独特型（Id）和GM-CSF作为佐剂。重要的是，那些对Id-KLH疫苗有抗独特型反应（50-70%）的患者有更好的无病生存。选择KLH作为载体蛋白为T细胞提供抗id应答的帮助在很大程度上是历史的，可能有更好的蛋白载体可以提高临床疗效。1可能是细菌蛋白李斯特菌溶素0 （LLO）。我们已经证明，当肿瘤相关抗原与截断的非溶血性LLO （LLOdetox）作为融合蛋白递送到免疫系统时，肿瘤相关抗原的免疫原性增强。我们在这个项目中的目标是探索LLOdetox的效用，以开发更有效的抗淋巴瘤免疫疗法。我们将测试将BCR独特型与LLOdetox融合是否会增强抗独特型免疫，以治疗小鼠B细胞淋巴瘤模型，即来自C3H/HeN小鼠的38C13 B细胞肿瘤。该模型已被研究人员广泛用于测试使用38C13独特型作为免疫原的疫苗策略。在初步研究中，我们从38C13的B细胞独特型中产生了一个重组可变区单链片段（scFV），并设计了一种用于监测同基因小鼠38C13肿瘤生长和残留疾病的FACS技术。在Specific Aim 1中，我们将使用大肠杆菌表达系统制备重组LLOdetox，并将38Cl3scFv化学偶联到rLLOdetox或KLH上。在特异性目标2中，我们将确定38C13scFv与rLLOdetox结合是否比传统疫苗结合38C13scFv- klh具有更好的抗肿瘤功效和免疫力。在I期完成后，我们希望能够证明38Cl3scFv-LLOdetox是一种优于38C13scFv-KLH的抗肿瘤免疫治疗药物。我们还将测试3种可能的佐剂，以增强其递送。这将为专注于准备进入临床试验的II期提案奠定基础。

DESCRIPTION (provided by applicant): Non-Hodgkin's Lymphoma (NHL) is characterized by an indolent clinical course with a median survival of 6-10 years. The incidence of NHL in the US increases by about 2.5% per year. Clearly, there is a national need for improved therapies for this malignancy. Idiotype-based clinical trials for minimal residual disease of follicular NHL have shown increased disease free survival as well as molecular remission. Currently, the best idiotype-based vaccine, which is in randomized phase III clinical trials, uses KLH as a protein carrier coupled to the B cell receptor (BCR) idiotype (Id) and GM-CSF as an adjuvant. Importantly, those patients who responded to the Id-KLH vaccine with an anti-idiotype response (50-70%) had a better disease free survival. The choice of KLH as the carrier protein to provide T cell help for the anti-Id response is largely historical and there may be better protein carriers that could improve clinical efficacy. 1 could be the bacterial protein listeriolysin 0 (LLO). We have shown enhanced immunogenicity to tumor associated antigens when they are delivered to the immune system as fusion proteins with a truncated non-hemolytic form of LLO (LLOdetox). Our objective in this project is to explore the utility of LLOdetox to develop more effective anti-lymphoma immunotherapeutics. We will test whether fusing the BCR idiotype to LLOdetox will enhance anti-idiotypic immunity in the treatment of a mouse model of B cell lymphoma, the 38C13 B cell tumor from the C3H/HeN mouse. This model has been used extensively by investigators to test vaccine strategies using the 38C13 idiotype as an immunogen. In preliminary studies, we have produced a recombinant single chain fragment of the variable regions (scFV) from the B cell idiotype of 38C13 and devised a FACS technique for monitoring 38C13 tumor growth and residual disease in syngeneic mice. In Specific Aim 1, we will prepare recombinant LLOdetox using an E. coli expression system and chemically conjugate 38Cl3scFv to either rLLOdetox or to KLH. In Specific Aim 2, we will determine whether conjugating 38C13scFv to rLLOdetox provides better anti- tumor efficacy and immunity than the conventional vaccine conjugate 38C13scFv-KLH. At the completion of Phase I, we hope to have demonstrated that 38Cl3scFv-LLOdetox is a superior anti-tumor immunotherapeutic than 38C13scFv-KLH. We will also have tested 3 possible adjuvants to enhance its delivery. This will set the stage for a Phase II proposal focused on preparations for entering clinical trials.