Tumor tissue mutation analysis

Formalin-fixed paraffin-embedded tumor tissue from the resected liver metastasis or primary tumor (where tumor tissue from liver metastasis was not available or inadequate for analysis) were analysed for somatic mutations in 15 genes recurrently mutated in CRC (SMAD4, TP53, AKT1, APC, BRAF, CTNNB1, ERBB3, FBXW7, HRAS, KRAS, NRAS, PIK3CA, PPP2R1A, RNF43, POLE). Tumor sections were macrodissected under a dissecting microscope to ensure a neoplastic cellularity of >30%. DNA was purified with a Qiagen FFPE Kit (Qiagen cat #56494). Primers were designed and sequencing results analyzed as previously described [16].