An astrocytoma CNS WHO grade 4 of a patient carrying a homozygous MSH6 germline frameshift variant, i.e. NM_000179.3(MSH6):c.691del p.(V231Yfs*15), had a TMB of 42.2 mut/Mb and served as positive control (Fig. 3a). Here, MSH6 is linked to astrocytoma (excluding glioblastoma).