Clonal hematopoiesis as a targetable mechanism of resistance to therapy in thyroid cancer.

About This Grant

PROJECT SUMMARY Anaplastic thyroid cancer (ATC) primarily affects elderly individuals and has a dismal prognosis compared to other thyroid cancer subtypes. One of the hallmarks of ATCs is their admixture with myeloid cells, primarily macrophages. The recent development of combinatorial treatment with BRAF and MEK inhibitors led to improved outcomes in Class 1 BRAF-mutant ATC patients. Although their initial response to MAPK inhibition is substantive, responses are not durable, leading to a median overall survival (OS) of ~15 months7. With aging, somatic mutations in hematopoietic stem and progenitor cells (HSPCs) promote clonal expansion over non- mutant HSPCs. When this is present in the absence of malignant transformation it is termed clonal hematopoiesis (CH). CH mutations arise most frequently in epigenetic modifier genes, such as DNMT3A and TET2. CH is associated with an increased risk of atherosclerotic cardiovascular disease and other diseases associated with aging. We showed in a pan-cancer analysis that CH, in particular CH with putative driver mutations (CH-PD), is associated with adverse outcomes in solid tumor patients, including those with ATC. The specific interactions between CH leukocytes and tumor cells in the TME and their impact on therapeutic response remain uncharted. We find that TET2-mutant CH is enriched in the TME of patients with ATC and other solid tumors. We developed syngeneic immunocompetent mouse models of concurrent Tet2-mutant CH and orthotopically implanted BrafV600E-driven ATC to explore the mechanisms involved. Using single cell-CITE-RNASeq we found that Tet2- mutant macrophages selectively infiltrate mouse BrafV600E-mutant ATC and cause resistance to BRAF-MEK inhibition through overexpression of Tgfβ-family ligands. Importantly, inhibition of the effects of Tgfβ at three distinct nodes restores sensitivity to MAPK pathway inhibition, opening a path for synergistic strategies to improve outcomes of patients with ATCs and concurrent CH. The mechanisms by which Tgfβ activation render ATCs insensitive to MAPK inhibition remain to be defined. We will investigate whether macrophage Tgfβ ligand overproduction induces resistance to MAPK inhibitors through cancer cell autonomous mechanisms and/or by its immune suppressive effects and use genetic approaches to nominate the key Tgfβ ligands responsible for treatment resistance. The mechanisms delineating how DNMT3A CH leads to worse outcomes in solid cancers in general, and ATC in particular, have eluded explanation. We will determine whether Dnmt3a-CH affects ATC biology and response to therapy in mice and test the hypothesis that this is driven by the infiltrating mutant myeloid population. Finally, we found that CH-PD is associated with worse OS in patients with ATC, but the effect of individual CH genes has not been established. We will determine whether specific CH genotypes impact OS in ATC and if this manifests at low CH variant allelic fractions (VAF) using a high sensitivity assay. We will also investigate whether advanced thyroid cancers of any type with a high tumor-to-blood CH VAFratio (i.e. CH mutant cell enrichment in the tumor as compared to blood) have worse clinical outcomes.