Progression of RAS-mutant leukemia during RAF inhibitor treatment

Abstract

Vemurafenib, a selective RAF inhibitor, extends survival among patients with BRAF V600E-mutant melanoma. Vemurafenib inhibits ERK signaling in BRAF V600E-mutant cells but activates ERK signaling in BRAF wild-type cells. This paradoxical activation of ERK signaling is the mechanistic basis for the development of RAS-mutant squamous-cell skin cancers in patients treated with RAF inhibitors. We report the accelerated growth of a previously unsuspected RAS-mutant leukemia in a patient with melanoma who was receiving vemurafenib. Exposure to vemurafenib induced hyperactivation of ERK signaling and proliferation of the leukemic cell population, an effect that was reversed on drug withdrawal.

Figures

Figure 1. The Clinical Course of a Patient with BRAF V600K–Mutant Melanoma and NRAS G12R–Mutant Chronic Myelomonocytic Leukemia on Treatment with Vemurafenib

The white-cell count and absolute monocyte count (Panel A) were monitored over the course of treatment. Solid triangles indicate doses of ipilimumab and the open triangle represents the time at which the bone marrow–biopsy specimen and the peripheral-blood smear were obtained. Shaded areas represent the periods of treatment with vemurafenib (Vem), administered at doses of either 960 mg or 720 mg twice daily. Myeloid dysplasia is seen in the peripheral-blood smear and the bone marrow aspirate (Panel B, arrows). The insets in Panel B show the biopsy specimen and aspirate at higher magnification. Computed tomographic (CT) scans show the response of the metastatic melanoma after treatment with vemurafenib at 8 and 16 weeks (Panel C). The red circles denote the location of melanoma metastases in the subcarinal lymph nodes (top row) and in the liver (bottom row).

Figure 2. RAS-Mutant Leukemia Cells, Hyperactivated in the Presence of Selective RAF Inhibitors

Peripheral-blood mononuclear cells (PBMCs) were collected from the patient when he was (+) and was not (−) receiving treatment with vemurafenib (720 mg twice daily). ERK signaling was assessed by measuring levels of phosphorylated ERK (pERK) with flow cytometry. The ratio of the median fluorescence intensity (MFI) of staining for pERK was compared with the MFI of staining for total ERK (tERK) in the aberrant monocyte population (CD14+, CD56+, HLA-DRhigh) and the lymphocyte population (CD3+) in the presence or absence of vemurafenib (Panel A). Total bone marrow mononuclear cells were plated in methylcellulose in the presence or absence of PLX4720 (2 μM or 20 μM), PD325901 (200 nM), or granulocyte–macrophage colony-stimulating factor (GM-CSF) (10 ng per milliliter). After 7 days, the total number of colony-forming units (CFUs) was enumerated (Panel B). PBMCs were treated for 4 hours with PLX4720 (2 μM or 20 μM) or PD325901 (100 nM) in the presence of GM-CSF (10 ng per milliliter). The ratio of the MFI of pERK to the MFI of tERK in the monocyte population was determined with the use of flow cytometry (Panel C). The graphs depict the mean of the experimental triplicates (Panel A), duplicates (Panel B), or quadruplicates (Panel C), and the T bars represent the standard error of the mean. In Panels A and C, the P values were calculated with the use of a paired t-test.