AML-M6 evolution and vemurafenib treatment. (A) Upper image: during vemurafenib treatment (gray box) lactate dehydrogenase (LDH; purple) levels (maximum 4297 U/L, normal range <250 U/L) and transaminases (green) increased. LDH levels were above normal levels before vemurafenib was administered, but liver enzymes rose with vemurafenib. During vemurafenib treatment, fluorescence-activated cell sorting demonstrated 40% of erythroblasts (red: glycophorin A⁺, CD36⁺, CD71⁺, CD34⁻) in the peripheral blood. After stopping vemurafenib, the LDH levels and liver enzymes returned to normal and erythroblasts disappeared from the peripheral blood (upper and lower images), suggesting dependence on vemurafenib. At 1.8 months after the stop of vemurafenib treatment, erythroblasts with the above-mentioned immunophenotype were again found in the peripheral blood, accompanied by massively elevated liver enzymes and LDH levels exceeding 16 000 U/L. BRAF V600E allele frequency (steel blue, middle image) and hairy cells (CD25, CD103, CD11c; steel blue, lower image) were significantly reduced upon vemurafenib treatment, whereas AML with PI3KCA mutation (E545K) emerged in the AML clone (middle image). We performed deep sequencing (Ion Torrent, >16 000 reads) on a trephine biopsy sample taken 44 days before vemurafenib was administered. The PI3KCA mutation E545K could not be confirmed within the sensitivity of the assay. (B) Immunohistochemistry revealed the strong positivity of p-ERK in glycophorin A⁺ intrasinusoidal erythroblasts (bone marrow biopsy) as a sign of ERK activation. Original magnification ×40. Orange box indicates AML induction treatment. GPT, glutamate pyruvate transaminase.