"Personalized cancer vaccines are therapeutic vaccines custom tailored to target tumor-specific mutations unique to a given patient. Such a treatment offers great hope for cancer patients as it does not harm healthy cells and has the potential to provide life-long remission. The emerging view of cancer as a dynamic evolutionary process that produces startling genetic heterogeneity and escapes treatment poses challenges for cancer vaccines. Yet progress in elucidating the nature of tumor heterogeneity has been impeded by experimental challenges, and the impact on vaccine design has not been investigated. The comprehensive research agenda outlined in this proposal aims to investigate tumor heterogeneity at the single cell level using novel experimental and theoretical methods with the end goal of maximizing the efficacy of personalized cancer vaccines. To accomplish these goals, I will first develop cost-effective high-throughput methods to measure the “mutanome” of hundreds of single cancer cells, integrating next-generation sequencing with microfluidics. Next, novel lineage treeing algorithms will be developed to reconstruct the evolutionary history of the cancer. I will then systematically apply this method to murine and human tumors and thus shed new light on cancer heterogeneity and evolution, reveal driver mutations with unprecedented resolution, enable the design of highly potent cancer vaccines and pave the way for new diagnostic tools. This proposal fits well within the scope of research conducted by the host institute, which aims to develop fundamentally new individualized cancer immunotherapies, with the first personalized cancer vaccines entering clinical trials in the near future. The host institute therefore has the experience and the resources to support this proposal and maximize its translational impact."