Yeast Display for the Identification of Peptide-MHC Ligands of Immune Receptors.

T cells detect peptide antigens presented by major histocompatibility complex (MHC) proteins via their T cell receptor (TCR). The sequence diversity of possible antigens, with trillions of potential peptide-MHC targets, makes it challenging to study, characterize, and manipulate the peptide repertoire of a given TCR. Yeast display has been utilized to study the interactions between peptide-MHCs and T cell receptors to facilitate high-throughput screening of peptide-MHC libraries. Here we present insights on designing and validating a peptide-MHC yeast display construct, designing and constructing peptide libraries, conducting selections, and preparing, processing, and analyzing peptide library sequencing data. Applications for this approach are broad, including characterizing peptide-MHC recognition profiles for a TCR, screening for high-affinity mimotopes of known TCR-binding peptides, and identifying natural ligands of TCRs from expanded T cells.

Identification of Highly Cross-Reactive Mimotopes for a Public T Cell Response in Murine Melanoma.

While immune checkpoint blockade results in durable responses for some patients, many others have not experienced such benefits. These treatments rely upon reinvigorating specific T cell-antigen interactions. However, it is often unknown what antigens are being recognized by T cells or how to potently induce antigen-specific responses in a broadly applicable manner. Here, we characterized the CD8+ T cell response to a murine model of melanoma following combination immunotherapy to determine the basis of tumor recognition. Sequencing of tumor-infiltrating T cells revealed a repertoire of highly homologous TCR sequences that were particularly expanded in treated mice and which recognized an antigen from an endogenous retrovirus. While vaccination against this peptide failed to raise a protective T cell response in vivo, engineered antigen mimotopes induced a significant expansion of CD8+ T cells cross-reactive to the original antigen. Vaccination with mimotopes resulted in killing of antigen-loaded cells in vivo yet showed modest survival benefit in a prophylactic vaccine paradigm. Together, this work demonstrates the identification of a dominant tumor-associated antigen and generation of mimotopes which can induce robust functional T cell responses that are cross-reactive to the endogenous antigen across multiple individuals.

Strategy management in collaborative clinical research partnerships.

PURPOSE: Today's clinical trial partnerships frequently join multi-disciplinary investigators and stakeholders, from different countries and cultures, to conduct research with a broad array of goals. This diversity, while a strength, can also foster divergent views about priorities and what constitutes success, thereby posing challenges for management, operations, and evaluation. As a sponsor and partner in such collaborations, we seek to assist and support their development and implementation of sound research strategies, to optimize their efficiency, sustainability, and public health impact. This report describes our efforts using an adaptation of the well-established Kaplan-Norton strategy management paradigm, in our clinical trials setting. We share findings from our first test of the utility and acceptance of this approach for evaluating and managing research strategies in a collaborative clinical research partnership. RESULTS: Findings from pilot studies and our first implementation in an ongoing clinical research partnership in Liberia, provide initial support for our hypothesis that an adapted version of the Kaplan-Norton strategy management model can have use in this setting. With leadership from within the partnership, analysis artifacts were gathered, and assessments made using standardized tools. Practical feasibility, resonance of the findings with partners, and convergence with other empirical assessments lend initial support for the view that this approach holds promise for obtaining meaningful, useable results for assessing and improving clinical research management. CONCLUSIONS AND IMPLICATIONS: Engaged leadership, thoughtful timing to align with partnership planning cycles, support for the process, and an eye towards the collaboration's long-term goals appear important for developing model understanding and practice. Skepticism about evaluations, and unease at exposing weaknesses, may hinder the effort. Acceptance of findings and associated opportunities for improvement by group leadership, support a growing sense of validity. Next steps aim to test the approach in other partnerships, streamline the methodology for greater ease of use, and seek possible correlations of strategy management assessments with performance evaluation. There is hardly a better example than the COVID-19 pandemic, to spotlight the need for efficient and effective clinical research partnerships to address global health challenges. While heartened by the collaborative spirit driving the effort so far, we cannot let our enthusiasm lull us into thinking that nobility of purpose or an abundance of good will is sufficient. Careful monitoring and adjustment of clinical research strategy in response to changes (e.g., demographics, pathogen evolution, research acceptance, political and cultural environments) are vital to making the needed adjustments that can guide these programs toward successful outcomes. We hope that our work can raise awareness about the importance, relevance, and feasibility of sound strategy management in clinical research partnerships, especially during this time when there is so much at stake.