Rapid expansion and comprehensive profiling of lung tumor infiltrating lymphocytes using a highly scalable TCR sequencing method 2258388

Abstract

Abstract Introduction Tumor infiltrating lymphocytes (TILs) have the ability to migrate into the tumor microenvironment (TME), recognize malignant cells, and achieve their clearance. Recent trials leveraging adoptive cell therapy (ACT) with ex-vivo expanded TILs in solid cancers have been successful in achieving disease remission. Their success relies heavily on persistence of tumor-specific TILs in circulation in patients after ACT administration. Two main challenges that have been slowing down the large-scale production of these therapies are the absence of fast and contamination-free TIL expansion protocols and the need for methods to assess the functional state and clonotype make-up of the TIL product prior to infusion. Methods Here we showcase newly developed methods to overcome these challenges and demonstrate their utility by generating a lung tumor TILs multimodal single cell dataset. Results Specifically, we demonstrate that the CliniMACS Prodigy Platform is able to quickly and reliably expand tumor infiltrating T cells in less than 2 weeks and perform single cell immune profiling sequencing in a subset of these cells to characterize their transcriptional state and dissect their T cell receptor (TCR) diversity. Using the gene expression data, we identify all expected T cell subsets using canonical cell type markers. Furthermore, we show we can detect paired TCR CDR3α/β sequences > 92% of the cells profiled. Using this information, we estimate the frequency of each CDR3 clonotype and identify hyperexpanded clonotypes that might be tumor reactive and are good candidates for further investigation. Conclusion Overall, we developed methods for the rapid expansion and deep profiling of TILs. We performed a proof of concept experiment that demonstrated their utility and identified expanded clonotypes that can be used to inform the design of future immunotherapies. We hope our methods will enable immunologists to create safer, faster and patient-specific treatments that achieve long-term tumor remission. Funding Source n/a Topic Categories Technological Innovations in Immunology (TECH)