By Charles Bankhead, Staff Writer, MedPage Today Published: June 11, 2011
Reviewed by Robert Jasmer, MD; Associate Clinical Professor of Medicine, University of California, San Francisco and Dorothy Caputo, MA, RN, BC-ADM, CDE, Nurse Planner
Explain that a single mutation (BRAF V600E) occurred in every patient in a cohort with hairy cell leukemia, suggesting that targeting the mutation could have major implications for treating the disease.Note that the method used was genome-wide massively parallel sequencing of tumor and normal cells from the same patient to identify recurrent somatic mutations.
A single mutation occurred in every patient in a cohort with hairy cell leukemia, suggesting that targeting the mutation could have major implications for treating the disease, Italian investigators reported.
The same BRAF V600E mutation appeared in 47 consecutive patients with hairy cell leukemia versus none of 195 patients with other types of peripheral B-cell lymphomas and leukemias.
In vitro studies showed that incubation of BRAF-mutated primary hairy cell leukemia cells with a specific inhibitor of BRAF led to a marked decrease in levels of two known downstream targets of BRAF kinase, according to an article published online in the New England Journal of Medicine.
"The BRAF V600E mutant is a potential therapeutic target in patients with hairy cell leukemia who do not have a response (or have a suboptimal response) to initial therapy with purine analogs, as well as in patients with repeated relapses or unacceptable toxic effects," Brunangelo Falini, MD, of the University of Perugia, and co-authors wrote in conclusion.
"Notably, BRAF V600E inhibitors have shown remarkable activity in patients with BRAF-mutated metastatic melanoma. These results, along with our in vitro finding that a specific active BRAF inhibitor causes MEK and ERK dephosphorylation in primary hairy cell leukemia cells, warrant the clinical testing of active BRAF inhibitors."
The findings were reported simultaneously at the European Hematology Association meeting.
Despite advances in the diagnosis and treatment of hairy cell leukemia over the past 50 years, little progress has occurred toward elucidation of the underlying genetic alterations associated with disease.
Gene-expression profiling has failed to pinpoint any recurrent genetic alterations, nor has genome-wide single-nucleotide polymorphism genotyping, the authors wrote in the introduction to their findings.
Falini and colleagues used a more powerful approach to examining the genetic basis of cancer: genome-wide massively parallel sequencing of tumor and normal cells from the same patient. They sought to identify recurrent somatic mutations in protein-coding genes, with the goal of gaining more insight into the origin of the disease and identifying new options for diagnosis and treatment.
Investigators obtained leukemic and nonleukemic genomic DNA from a single patient with hairy cell leukemia. By use of massively parallel sequencing, candidate somatic mutations were identified. Researchers used polymerase-chain-reaction (PCR) amplification and direct DNA sequencing to validate the findings from the index patient.
Falini and colleagues performed whole-exome sequencing of genomic DNA from the index patient and identified five unique variants specific to tumor DNA, the best known of which was BRAF.
The amino acid substitution at position 600 of the BRAF protein (V600E) is associated with several solid tumors. At the recent American Society of Clinical Oncology meeting, treatment with a BRAF inhibitor led to unprecedented activity in patients with metastatic melanoma.
With the finding from the index patient, Falini and colleagues screened DNA samples for 47 additional patients with hairy cell leukemia and the 195 patients with other leukemias and lymphomas. They identified the V600E mutation in samples from all of the patients with hairy cell leukemia but none of the patients with other hematologic malignancies.
Working from the understanding that V600E constitutively activates BRAF kinase activity, Falini and colleagues evaluated the phosphorylation status of MEK and ERK. The former is the kinase target immediately downstream from BRAF and the latter is the kinase phosphorylated by MEK.
Using antibodies specific to phosphorylated MEK and ERK, the investigators confirmed the presence of the two kinases in DNA specimens from five patients with BRAF-mutated hairy cell leukemia. Introduction of the BRAF inhibitor PLX-4720 led to marked decrease in phosphorylated MEK and ERK, whereas vehicle-treated samples retained MEK and ERK phosphorylation.
Falini had no relevant disclosures. Co-author Robin Foa disclosed relationships with Roche, Bristol-Myers Squibb, GlaxoSmithKline, and Celgene.
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