Molecular engineering and systematic evaluation of bispecific aptamers to develop potent and efficacious therapies for the immunomodulation of Non-Small Cell Lung Cancer

项目摘要 癌症将绕过心血管疾病，成为美国的头号死因， 是世界范围内的主要死因非小细胞肺癌（NSCLC）是一个主要的促成因素， 统计数据。化疗药物输送的最新进展和小分子抑制剂的开发， 例如酪氨酸激酶抑制剂在降低疾病流行率和负担方面是不可或缺的。 此外，最近FDA批准的免疫调节疗法，如免疫检查点抑制剂， 嵌合抗原受体（CAR）T细胞、嵌合抗原受体（ICI）T细胞和双特异性抗体的研究强调了以下重要性： 免疫系统逃避在疾病进展和复发中起作用。不幸的是，管理这些 靶向和免疫调节疗法经常遇到肿瘤获得性抗性（例如，二次 突变; T细胞耗竭）并引发非特异性或靶向/非靶向肿瘤副作用（例如，免疫相关 不良事件）。这表明需要替代或辅助NSCLC疗法，其不仅有效， 有效，但行使广泛的治疗指数。我们建议利用适体技术作为一种潜力 解决这一需求的方式。适体是单链寡核苷酸，其以高亲和力结合其靶标。 特异性和亲和力以及它们作为外来物质的免疫原性的相对缺乏，与抗体相比， 使它们成为调节免疫系统的理想试剂。此外，它们的易操作性使得 设计和优化这些试剂的分子工程相对简单。我们的目标是开发 双重结合免疫细胞CD 3 ε和NSCLC肿瘤的新型免疫调节双特异性适体（bsApts） 相关抗原（TAA）以诱导有效免疫突触的形成。我们建议使用分子 合理设计bsApt和系统评估特定bsApt属性的工程技术， 如（i）化合价，（ii）亲和力，和（iii）接头长度/类型，它们诱导人工免疫细胞活化的能力， 体外和体内抗肿瘤反应。我们还建议采用转录组学方法来更好地了解 设计/靶标如何影响肿瘤异质性、肿瘤浸润淋巴细胞表型和脱靶免疫 细胞激活次要目标是改善限制bsApt临床应用的药代动力学特性 长期目标是将我们对这些属性的发现推广到当前和未来， 靶向广泛的实体癌和血液癌的双特异性疗法。

PROJECT SUMMARY Cancer is set to bypass cardiovascular disease as the number one cause of death in United States and it is a leading cause of death worldwide. Non-Small Cell Lung Cancer (NSCLC) is a major contributing factor to this statistic. Recent advancements in chemotherapeutic delivery and the development small molecule inhibitors, such as tyrosine kinase inhibitors, have been indispensable in decreasing disease prevalence and burden. Additionally, the recent FDA approvals of immunomodulating therapies, such as immune checkpoint inhibitors (ICIs), chimeric antigen receptor (CAR) T cells, and bispecific antibodies, emphasizes the importance that immune system evasion plays in disease progression and relapse. Unfortunately, administration of these targeted and immunomodulating therapies is often met with tumor acquired resistance (e.g., secondary mutations; T cell exhaustion) and incites non-specific or on-target/off tumor side effects (e.g., immune related adverse events). This suggests a need for alternative or adjuvant NSCLC therapies that are not only potent and efficacious but exercise a wide therapeutic index. We propose to exploit aptamer technology as one potential way to address this need. Aptamers are single strand oligonucleotides that bind to their targets with high specificity and affinity and their relative lack of immunogenicity as a foreign substance, compared to antibodies, make them ideal reagents to modulate the immune system. Furthermore, their ease of manipulation makes molecular engineering to design and optimize such reagents relatively straightforward. Our goal is to develop novel immunomodulating bispecific aptamers (bsApts) that dually bind to immune cell CD3ε and NSCLC tumor associated antigens (TAAs) to induce formation of effective immune synapses. We propose to use molecular engineering techniques to rationally design bsApts and systematically evaluate specific bsApt properties, such as (i) valency, (ii) affinity, and (iii) linker length/type in their ability to induce artificial immune cell activation in vitro and anti-tumor responses in vivo. We also propose to take a transcriptomics approach to better understand how designs/targets affect tumor heterogeneity, tumor infiltrating lymphocyte phenotypes, and off-target immune cell activation. Secondary goals look at improving pharmacokinetic properties that limit bsApt clinical translatability while long-term goals look to generalize our findings on these properties to current and future bispecific therapies that target a wide range of solid and hematological cancers.