Exploring molecular encoding in the context of T cell-based immunotherapy

The immune system plays a pivotal role in maintaining general health and providing protection against the abundance of invading pathogens that an individual may come across in their lifetime. Regulation and clearance of cells undergoing malignant transformation is another critical function of the immune system. However, the heterogeneous nature of cancer has proven to be a major obstacle for obtaining effective therapeutic outcome in the majority of patients. The concept of immunotherapy, where the aim is to boost the patient’s natural immunity towards cancer, has revolutionized the way cancer treatment is approached. Nonetheless, despite massive advancements in the field, the multifactorial nature of the interactions between the immune system and cancer has resulted in mixed therapeutic outcomes. The factors which contribute to clinical response are not universal, which partly explains why the majority of patients still do not respond favorably to strategies such as immune checkpoint blockade therapy. As such, there has been a great effort in recent years to identify the key parameters, or immune predictors, which dictate the patient’s outcome to immune-based therapies. A comprehensive understanding of these immune predictors would enable a more effective design of treatment strategies, based on the profile of the given cancer type or individual patient.
The overall objective of this thesis is to evaluate tumor antigen recognition and novel tools for the detection of antigen-specificity, as well as to highlight nanoparticle-based strategies for delivery of therapeutic cargo, in the context of T cells and cancer therapy. These concepts are connected by one common element, the use of DNA barcodes as a form of molecular encoding.
In manuscript I, DNA barcode-labeled MHC multimers are utilized in combination with a multicolor T cell phenotype panel in a novel approach that allows the parallel analysis of T cell recognition with corresponding phenotypic characteristics. This was performed to assess the characteristics of neoantigen-reactive CD8+ T cells (NARTs), one of the primary drivers of immunotherapy-based tumor elimination, in the hopes of understanding whether T cell-intrinsic factors are involved in the response to therapy. This study was performed in a diverse cohort of cancer patients from a Phase I basket trial, in an effort to identify a profile of NARTs that could distinguish responding from non-responding patients. Although, due to the small sample size and heterogeneity of the cohort, no significant difference was observed between the patient groups, related to the breadth and magnitude of the detected NARTs. There also appeared to be no significant phenotypic profile associated with the responding patient NARTs. However, insights into a favorable antigen-reactive T cell profile were made evident. Tendencies for increased expression of markers such as Ki67, CD27 and TCF-1 were observed in some of the patients, post-therapy. Indicating that the presence of proliferating, early-activated NARTs with self-renewal properties could provide therapeutic benefit. Further studies in larger cohorts would be warranted to provide concise conclusions.
In the first additional results section, a novel strategy was applied to make use of DNA barcodes for encoding MHC tetramer reagents for the detection of viral-reactive CD8+ T cells. The second additional results section implemented a DNA barcode in the nanoparticle design, as a proof of concept for developing a screening platform to identify formulations successful in the in vivo delivery of messenger RNA cargo. These preliminary results indicate that the use of DNA barcodes could be widely applicable for encoding purposes, not only to identify relevant drug and T cell targets but also to facilitate the design of more effective nanoparticle delivery systems for enhancing current therapeutic strategies.