PTEN gene expression and mutations in the PIK3CA gene as predictors of clinical benefit to anti-epidermal growth factor receptor antibody therapy in patients with KRAS wild-type metastatic colorectal cancer

Abstract

Purpose: To identify novel genetic markers predictive of clinical benefit from epidermal growth factor receptor-directed antibody therapy in patients with metastatic colorectal cancer.

Patients and methods: Seventy-six consecutive patients who received cetuximab or panitumumab, either alone or in combination with chemotherapy and with available tumor tissue were included. Tumor tissue was tested by pyrosequencing for mutations at known hot spots in the KRAS, BRAF, PIK3CA, PIK3R1, AKT1, and PTEN genes. PTEN promoter methylation status was analyzed by methylation-specific polymerase chain reaction, and expression was determined by immunohistochemistry (IHC). Forty-four patients had 4 weeks of therapy and were considered for clinical correlates.

Results: Consistent with previous studies, KRAS gene mutations were associated with a shorter progression-free survival (PFS) and overall survival (OS). Among the patients with wild-type KRAS, preservation of PTEN expression and PIK3CA wild-type status was associated with improved OS (median OS, 80.4 vs. 32.5 weeks; hazard ratio, 0.33; P = .0008) and a trend toward improved PFS (median PFS, 24.8 vs. 15.2 weeks; hazard ratio, 0.51; P = .06), compared with PTEN-negative or PIK3CA-mutant tumors. PTEN methylation was more common in the metastatic samples than in the primary samples (P = .02). The simultaneous presence of methylation and mutation in the PTEN gene was associated with IHC negativity (P = .026).

Conclusion: In addition to KRAS mutation, loss of PTEN expression (by IHC) and PIK3CA mutation is likely to be predictive of a lack of benefit to anti-EGFR therapy in metastatic colorectal cancer. PTEN promoter methylation and mutation status was predictive of PTEN expression and may be used as an alternative means of predicting response to EGFR-targeted therapy.

Figure I

Effect of KRAS, PIK3CA, and PTEN gene expression status in patients Panels A and B Kaplan Meier curves for progression free survival (PFS) and overall survival (OS) for patients based on the mutation status in the KRAS gene. There was a statistically significant difference in both PFS and OS in favor of the patients with a WT KRAS gene. Panels C and D Kaplan Meier curves for progression free survival (PFS), and overall survival (OS) based on the PTEN expression by IHC and PIK3CA mutation status among the KRAS WT patients. There was a statistically significant difference in OS, and a strong trend towards a difference in PFS, in favor of the patients with a preserved IHC PTEN/PIK3CA WT status.

Figure II

Panels A–D PTEN staining by IHC showing the scoring patterns; A: score of 0, B: score of 1, C: score of 2, and D: score of 3

Figure III

Agarose gel showing examples of methylation specific PCR (MSP) using primer set 3 (Supplementary table III). Abbreviations; 100 BP: DNA ladder with position marked for the 100 base pair location; Pos Con: positive control; M: PCR product using primers to amplify methylated DNA; U: PCR product using primers to amplify unmethylated DNA; 1,2 and 4: tumor samples with methylated DNA showing both methylated and unmethylated product; 3: tumor sample with unmethylated DNA with only unmethylated product