Structure-based engineering of immune cytokine signaling

摘要： 免疫调节细胞因子与跨膜信号受体结合，以介导广泛的 功能包括通过JAK/STAT激活的白细胞的增殖、分化和扩增。多数 免疫调节性细胞因子具有对正常免疫至关重要的多余和独特的活性。 动态平衡，但这种功能多效性对这些细胞因子的有效利用提出了一个主要问题 免疫治疗癌症药物。细胞因子的多效性是不同的细胞因子受体 在多种不同的平衡细胞类型上表达，用于中和抗肿瘤作用和铅 对全身的毒性。在这个奖项的前一期，我们获得了对细胞外 不同免疫细胞因子复合体的结构结构，包括IL-2，IL-10， IL-12、IL-21、IL-22、IL-23、IL-27、干扰素、干扰素。在此续订应用程序中，这些结构模板位于 另一方面，在目标1中，我们的研究重点是细胞因子IL-2、干扰素和IL-12。这些免疫主控调节器 参与不同但重叠的免疫系统分支，分享多效性和毒性问题 如果不结合，可能会产生强大的癌症免疫治疗剂。我们建议对信号进行“调整” 通过基于结构的细胞因子工程，试图创造具有解偶联多效性、细胞子集的变体 偏好，增强抗肿瘤效果，降低毒性--无论是单独使用还是联合使用。在目标2中，我们 希望了解调节的细胞因子如何以不同的方式激活细胞内的信号的机制基础 细胞。基于我们20年来解决全长JAK结构的最新突破，我们继续 寻求细胞因子与其受体结合如何激活Janus Kinase(JAK)的结构信息 分子，通过重组和成像激活的JAK同源和异源二聚体与细胞因子和 细胞内JAK和STATS。以这种方式，通过将结构生物学、蛋白质工程、细胞 信号传递和体内肿瘤研究，我们建议获得细胞因子受体的完整分子快照 通过激活细胞内信号级联激活配体的初始参与的信号和 将这些信息用于癌症免疫疗法的工程。

ABSTRACT: Immunoregulatory cytokines engage transmembrane signaling receptors in order to mediate a wide range of functions including leukocyte proliferation, differentiation, and expansion through JAK/STAT activation. Most immunoregulatory cytokines possess both redundant and distinct activities that are critical to normal immune homeostasis, but this functional pleiotropy presents a major problem for the effective use of these cytokines as immunotherapeutic cancer drugs. Cytokine pleiotropy is a consequence of different cytokine receptors being expressed on multiple different counterbalancing cell types that serves to neutralize anti-tumor actions and lead to systemic toxicity. During the prior term of this award, we gained an appreciation for the extracellular structural architectures of a spectrum of different immune cytokine complexes, including those of IL-2, IL-10, IL-12, IL-21, IL-22, IL-23, IL-27, IFN, and IFN. In this renewal application, with these structural templates in hand, in Aim 1 we focus our studies on the cytokines IL-2, IFN, and IL-12. These immune master regulators engage different but overlapping branches of the immune system, and share the issues of pleiotropy and toxicity that, if uncoupled, could lead to powerful cancer immunotherapeutic agents. We propose to “tune” signaling through structure-based cytokine engineering to attempt to create variants with decoupled pleiotropy, cell subset preferences, enhanced anti-tumor efficacies, and reduced toxicity – both alone and in combination. In Aim 2, we wish to understand the mechanistic basis for how tuned cytokines can differentially activate signaling inside the cells. Based on a recent breakthrough in our 20-year quest to solve the full-length JAK structure, we continue to pursue structural information on how cytokine binding to their receptors activates Janus Kinase (JAK) molecules, by reconstituting and imaging activated JAK homo- and heterodimers bound to both cytokine and intracellular JAKs and STATs. In this fashion, by combining structural biology, protein engineering, cell signaling, and in vivo tumor studies, we propose to obtain a complete molecular snapshot of cytokine receptor signaling from the initial engagement of ligand through the activation of intracellular signaling cascades and leverage this information for the engineering of cancer immunotherapeutics.