Cancer stem cells in solid tumors have been recently isolated and appear to be primarily responsible for the growth and spread of the disease. The presence of a stem cell population in a tumor has implications for the diagnosis and treatment of cancer, as it is these cancer stem cells that must be targeted to achieve a cure. Preliminary evidence demonstrates that there is a host of genes differentially expressed by the cancer stem cells and their non-tumorigenic progeny. Many of these genes are thought to play a role in essential cancer functions including proliferation, survival, self renewal and resistance to standard therapeutics. Uncovering the true functional stratification of the superficially uniform population of stem cells in leukemia, breast cancer and other solid cancers is a challenge which requires new kinds of measurements at the single-cell level. Based on recent discoveries by the Clarke and Weissman groups, we believe that gene expression in normal stem cells, as well as cancer stem cells is significantly regulated at the epigenetic level. Epigenetic regulation of gene expression is in part controlled by the posttranslational modification of histone proteins, as well as methylation of DMA, both of which result in the alteration of chromatin structure. These modifications are examined using Chromatin immunoprecipitation (ChIP) assays. Epigenetic analysis is a necessary complement to gene expression analysis in order to understand the control of normal and cancer stem cell self-renewal and discover new therapeutic targets. The microfluidic tools developed by the Quake lab for single cell gene expression (aim1), chip (aim 2) and high throughput in vitro cell culture (aim 3) will allow the Clarke and Weissman laboratories to perform gene expression and epigenetic analyses on rare, purified cells from model mice and primary human cancer or cancer xenografts. These assays will all use microfluidic platforms which have already been developed and validated in the Quake laboratory or are commercially available. New systems will also be designed as needed in the course of this project. Microfluidic epigenetic and genetic assays will allow the study of highly purified, homogeneous, rare cell populations that were previously inaccessible with the standard techniques. Microfluidic cell culture platforms will allow finding appropriate conditions to culture cancer stem cells in vitro and test new therapeutic targets
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