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. 2015 Jun 23;7(9):1104–1118. doi: 10.15252/emmm.201404914

Figure 2. Reactivation of MAPK pathway via amplification of a BRAF-encoding DNA fragment caused resistance to vemurafenib in three metastases.

Figure 2

  1. Hematoxylin–eosin (H&E) and p-ERK stainings on FFPE material of all metastases showed that all vemurafenib-resistant tumors had reactivation of the MAPK pathway. Scale bar represents 100 μm.
  2. CNA profiles were generated from the WES data with germ line DNA as a reference. Colors represent segmented log2 ratio values with red for gain and blue for loss. Further inspection of chromosome 7 where BRAF is located revealed amplification (7q34) of this region in three vemurafenib-resistant metastases, namely M032R1, M032R2, and M032R5.
  3. qPCR was performed on gDNA retrieved from each of the metastases, using primers for BRAF and CRAF and normalized on LINE levels. Bars represent the mean of three replicates, error bars indicate standard deviation. The results confirmed that BRAF was amplified in M032R1, M032R2, and M032R5.
  4. Staining for BRAFV600E with a mutant epitope-specific antibody confirmed the upregulation of BRAFV600E in R1, R2, and R5. Scale bar represents 100 μm.