Abstract: Objective:To explore genes associated with sensitivity to anti-EGFR therapy.Methods:From March2012to August 2013, 31metastatic colorectal cancer patients in Peking University Cancer Hospital & Institute were treated with anti-EGFR mono-therapy. A total of 21genes associated with oncogenesis, metastasis, and EGFR signaling pathway were profiled in these31patients by using tar-geted next-generation sequencing technology. Results:A total of 31patients with Kras exon 2 wild type received anti-EGFR therapy as third-line treatment. Among these patients, the median progression-free survival (PFS) was 89days, overall survival was311 days, and objective response rate was 16. 1%. Five cases harbored Kras exons 3/4 or Nras exons 2/3 mutations. These five Ras mutation patients showed disease progression during the first evaluation with 31-day PFS. One PIK3CA mutation case exhibited disease progression dur-ing the first evaluation (PFS 35days), and one case showed mTOR mutation with91-day PFS. The PFS of two cases with SMAD 4 muta-tion were 58and 59days, whereas that of the case containing FBXW 7 mutation was 93days. Among the 26Ras wild- type patients, MLL3, TP 53, and APC were the three genes with the highest mutation frequencies of 92. 3% (24/26), 53. 8% (14/26), and 42. 3% (11/ 26), respectively. Conclusion: Extended Ras analysis (including Kras and Nras exons 2/3/4) is recommended for patients who are candi-dates for anti-EGFR therapy. Mutations in the downstream effectors of the EGFR signaling pathway, such as PI 3KCA and mTOR, may al-so have a predictive role in anti- EGFR therapy. Mutations beyond the EGFR pathway such as FBXW7 and SMAD4 may be associated with anti-EGFR efficacy and deserve further attention.