Glioblastoma is one of the most heterogeneous and most aggressive human tumors. It remains an unmet clinical need, as limited benefits were observed both with targeted treatment and immunotherapy approaches. We hypothesize that significant variation in tumor microenvironments, generating differences in oxygen and nutrient supply between these areas, may contribute to survival of specific cancer cell subpopulations. By developing new in situ applications, suited for formalin-fixed archival specimen, we will retain the relative localization of cells with distinct genomic features within the tissue sections. Spatial analysis could infer preferences for particular neighborhoods or niches and cellular interactions. Our proposal will overcome several limitations of currently available methods, to allow identification of non-hotspot mutations at the single cell level in situ and simultaneously profile the tumor microenvironment.
In Aim 1, we will use two different approaches to develop an assay that will mark cells with mutations relevant to therapy response in standard intact biopsy sample. Our new methods will yield a ?yes-no? binary result for presence of any, including non-hotspot, mutations within a genomic region of interest, at single cell level in situ. Thus, they could facilitate screening for actionable mutations in pathology units, with minimal required equipment and streamlined analysis pipeline. By analyzing genetically heterogeneous cellular localizations we may identify direct interactions between diverse populations of cancer cells. To complement this approach, in Aim 2 we will adapt our previously published method, to not only account for genetically defined populations of cancer cells, but to include markers of normal cells, such as leukocytes, glial cells and neurons, that all contribute to development of a complex tumor ecosystem. The proposed projects would complement the parental grant by expanding our methodological capabilities to better characterize the relationship between heterogeneity and glioblastoma progression.
Effective eradication of cancer is often impeded by the presence of genetically diverse subpopulations of cells with varyingability to survive anti-cancer treatment. The STAR-FISH assay I developed allows for the monitoring of intratumor genetic diversity during treatment at the single cell level and in situ. Dissecting mechanisms underlying intratumor heterogeneity will lead to improved treatment design and patient outcomes.
