Novel Anti-CCR8 VHH for the Treatment of NSCLC

摘要 免疫检查点抑制剂(ICI)疗法的出现极大地改变了对 非小细胞肺癌(NSCLC)。然而，大多数接受ICIS治疗的患者要么没有反应，要么 发展为进行性疾病(PD)，尽管最初对治疗有反应。这背后的机制之一是 抑制性免疫细胞群在肿瘤微环境(TME)中的作用 T细胞(Tregs)，它起到保护肿瘤免受患者免疫反应的作用。我们计划瞄准并耗尽 这些Tregs通过C-C趋化因子G蛋白偶联受体8(CCR8)特异性地位于TME内，CCR8 在激活的肿瘤驻留Tregs中选择性上调，而在必要的效应T淋巴细胞中缺失。 选择性消除CCR8+Tregs有望促进对非小细胞肺癌更有效的免疫反应 肿瘤，同时避免与非选择性耗竭相关的危险的自身免疫副作用。 针对CCR8和其他G蛋白偶联受体(GPCRs)的抗体疗法历来是 这是一个挑战。这在很大程度上是由于它们的固有特性使其抗原较差，其中包括它们的低 丰度低，免疫原性差，构象异质性。为了解决这个问题，Abilita Bio开发了 启用膜蛋白(emp™)定向进化技术，产生增强的gpr变异体 在保持其结构和生物学特性的同时，对生物物理特性进行了革命性的改进 关联性。使用电磁脉冲™方法，我们进化出了构象稳定的CCR8版本，这是 作为蛋白抗原用于骆驼免疫和VHH单域抗体家族的发现 独特地靶向野生型人类受体的跨膜核心。我们的目标是利用这些 抗体具有构建一种具有最佳潜力的分子的优势特性。在建议的阶段 本人对SBIR的研究，我们将在体外完成对抗体HITS的彻底表征和优化，筛选出一种 铅分子，然后在一种新的CCR8人源化非小细胞肺癌小鼠模型中进行体内疗效研究 在与竞争对手的抗体进行面对面的比较时，以显示优势。我们将实现这一目标 通过执行以下科学目标。目标1：我们将分析我们多样化的VHH热门收藏 体外结合特性和生物物理稳定性，进而选择最佳分子进行进一步研究 人物刻画。目的2：我们将在作用机制研究中描述抗体的药理和效力。 选择一种用于体内研究的铅抗体。在目标3中：我们将在CCR8中测试我们的铅抗体的体内效果- 人源化非小细胞肺癌小鼠模型。据我们所知，这是CCR8的第一个活体疾病模型研究 治疗计划，其中人CCR8将在功能性TME的背景下作为靶点。成功 这项第一阶段研究的完成将为这些新药的治疗潜力提供关键的验证 抗体和证据支持我们最好的假说。这些研究的结果将支持继续 项目进展和未来申请SBIR第二阶段赠款，以支持完成临床前 发展。

Summary The advent of immune checkpoint inhibitor (ICI) therapies has dramatically changed the treatment landscape for non-small cell lung cancer (NSCLC). However, most patients treated with ICIs are either non-responders or develop progressive disease (PD), despite an initial response to therapy. One of the mechanisms behind this, is the action of suppressive immune cell populations within the tumor microenvironment (TME) such as regulatory T cells (Tregs), which act to shield tumors from the patient's immune response. We plan to target and deplete these Tregs specifically within the TME through the C-C Chemokine G Protein-coupled Receptor 8 (CCR8), which is selectively upregulated in activated tumor-resident Tregs and is absent from essential effector T lymphocytes. Selective elimination of CCR8+ Tregs is anticipated to promote a more effective immune response against NSCLC tumors, while avoiding the dangerous autoimmune side effects associated with non-selective depletion. Targeting CCR8 and other G protein-coupled receptors (GPCRs) with antibody therapeutics has historically been a challenge. This is largely due to the intrinsic properties that make them poor antigens, which include their low abundance, poor immunogenicity, and conformational heterogeneity. To address this, Abilita Bio developed the Enabled Membrane Protein (EMP™) directed evolution technology, which generates enhanced GPCR variants with transformative improvements in biophysical properties, while preserving their structure and biological relevance. Using the EMP™ methodology, we evolved a conformationally stabilized version of CCR8, which was used as a protein antigen for llama immunization and the discovery of VHH single-domain antibody families that uniquely target the transmembrane core of the wild-type human receptor. Our goal is to leverage these antibodies' advantageous properties to construct a molecule with best-in-class potential. In the proposed Phase I SBIR research, we will complete a thorough in vitro characterization and optimization of antibody hits, select a lead molecule, and then perform in vivo efficacy studies in a novel CCR8-humanized mouse model of NSCLC in a head-to-head comparison with a competitor’s antibody to show superiority. We will accomplish this goal through the execution of the following scientific aims. Aim 1: we will analyze our diverse VHH hit collection for binding characteristics and biophysical stability in vitro, and then select the best molecules for further characterization. Aim 2: we will characterize antibody pharmacology and potency in mechanism of action studies to select a lead antibody for in vivo studies. In Aim 3: we will test in vivo efficacy of our lead antibody in a CCR8- humanized mouse model of NSCLC. This is, to our knowledge, the first in vivo disease model study for a CCR8 therapeutic program, where human CCR8 will be targeted in the context of a functional TME. Successful completion of this Phase I research will provide critical validation of the therapeutic potential for these novel antibodies and evidence to support our best-in-class hypothesis. Results of these studies will support continued program advancement and a future application for an SBIR Phase II grant to support the completion of preclinical development.