Pervasive Somatic Mutations in NF1 Drive Cell Type and Spatially Specific Clonal Selection in Neurofibromatosis

Tumorigenesis is a multi-step process shaped by the accumulation of somatic driver mutations, but identifying the earliest steps and cell of origin of tumorigenesis remains challenging. Single cell genomics in >1.7 million single cells across donors with the cancer predisposition syndrome neurofibromatosis type 1 (NF-1), shows that somatic 'second-hit' mutations in NF1 are pervasive and show positive clonal selection in oligodendrocytes (OLs) and OL precursor cells (OPCs), but are exceedingly rare in neurons and other lineages. Combined spatial genotyping and transcriptomic analysis enabled dissection of cell autonomous and non-autonomous effects of second-hit mutations including clonal expansion of mutant OPCs with strong bias for cerebral gray matter versus white matter, suggesting that gray matter influences OPC proliferation and differentiation via non-cell autonomous interactions between OPCs and neurons. NF1 -null OL lineage cells upregulate CDKN2A -a tumor suppressor frequently lost in NF1 -mutant tumors- whereas brain cancers from these same individuals show CDKN2A loss across multiple cancer cell states, suggesting CDKN2A loss in OPCs is a critical step in tumor progression. Our findings reveal developmentally early, exquisitely lineage-specific NF1 somatic mutations in premalignant NF-1 brain, suggest cell autonomous and non-autonomous roles in clonal selection in brain, and suggest that clones harboring cancer driver mutations can persist over a lifetime in phenotypically normal tissues.