Molecular mechanisms of T cell responses to a clonal neoantigen resulting from a mutated driver oncogene.

PROJECT SUMMARY/ABSTRACT Immunotherapy induces durable remissions in a subset of patients with highly mutated cancers. However, most cancers are modestly mutated and fail to respond to current immunotherapy treatments. This is especially true for malignancies caused by activating mutations in phosphatidylinositol 3-kinase catalytic subunit alpha (PIK3CA), the most commonly mutated driver oncogene in humans. Mutant PIK3CA cancers exhibit resistance to conventional treatments, including chemotherapy, hormonal therapy, and antibodies. Innovative new approaches that bring the curative potential of immunotherapy to PIK3CA mutated cancers are therefore urgently needed. We and others previously performed detailed immune monitoring studies of exceptional patient responders to resolve the mechanisms of successful immunotherapy. These analyses revealed that T cells from responders often recognize neoantigens (NeoAgs) - peptides derived from the protein products of somatic mutations presented by a patient’s unique complement of human leukocyte antigen (HLA) molecules. In >99% of cases, NeoAgs are exclusive to an individual patient because they result from passenger mutations that do not contribute to cancer cell fitness and therefore are subject to clonal heterogeneity. NeoAg clonal heterogeneity has emerged as a major cause of immunotherapy resistance. We hypothesize that clonally expressed NeoAgs derived from hotspot mutations in mutant PIK3CA can be immunogenic and are amenable to therapeutic targeting using T cell receptors (TCRs). In support of our hypothesis, we discovered through a mass spectrometry (MS) screen that a shared NeoAg derived from mutant PIK3CA is naturally processed and presented in the context of a prevalent HLA allele. We have termed this unique subset of antigens “public” NeoAgs because they are cancer-specific yet expressed by groups of patients, enabling the use of off-the-shelf reagents. Using a novel TCR discovery platform, we successfully generated multiple T cell clones specific for this PIK3CA public NeoAg, retrieved their unique TCR gene sequences, and exogenously transferred public NeoAg reactivity to non-specific T cells. These results confirm the immunogenicity of MS-identified public NeoAgs and enable the development of TCR-based gene therapies. Building on these preliminary data, we propose in Aim 1 to develop a novel therapeutic approach for cancers expressing a PIK3CA public NeoAg using TCR gene transfer and adoptive immunotherapy. In Aim 2, we will establish in cancer patients the frequency, immune-compartmentalization, and potential pathways of resistance to T cells specific for a PIK3CA public NeoAg. In Aim 3, we will resolve the physical basis for PIK3CA public NeoAg immunogenicity by studying the physical and structural properties of public NeoAg/HLA molecules, their wild type counterparts, and the complexes they form with TCRs. Together, this work will elucidate the fundamental principles governing NeoAg immunogenicity in humans, enabling innovative new precision immune-genomic approaches for common epithelial cancers currently lacking curative treatments.