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

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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.