Signaling Assay Identifies Juvenile Myelomonocytic Leukemia

A signaling assay that builds on the cell-sorting technique, called flow cytometry, could lead to a fast diagnostic test for the rare disease juvenile myelomonocytic leukemia (JMML).

In flow cytometry, fluorescently labeled antibodies are used to classify and sort cells based on proteins displayed on their outer surface. A new approach creates small holes in the cell membrane prior to sorting. These holes allow other antibodies to enter the cell and bind to signaling molecules involved in the cell's internal monologue.

Scientists tested the technique's clinical value by applying it to the diagnosis of JMML. Children with this rare disorder typically have fevers, grow poorly, suffer from infections, and generally look like they could have any one of a number of different diseases. A prompt diagnosis of JMML is particularly important because the only cure is a bone marrow transplant.

JMML cells tend to proliferate in response to very low levels of a growth-stimulating factor called granulocyte macrophage-colony stimulating-factor (GM-CSF); normal cells respond only at higher levels. However, it can take two to three weeks to grow enough cells in the laboratory to get a definitive answer to this test.

GM-CFS activates a particular cellular signaling cascade called the Janus kinases-signal transducers and activators of transcription (JAK-STAT) pathway. Although that pathway had not previously been directly implicated in JMML, Nikesh Kotecha, Ph.D., a graduate student in the laboratory of Prof. Garry Nolan, Ph.D., at Stanford University School of Medicine (Stanford, CA, USA), used an antibody that binds only to activated STAT5 to determine whether the cells of 12 patients with JMML displayed abnormally high levels of the protein in response to low doses of GM-CSF. Eleven of the 12 did so--confirming the involvement of the STAT pathway in the disorder.

The new technique also offers a way to monitor disease progress. With further refinement, the scientists hope that the technique can be used to screen the effectiveness of potential drugs for treatment of JMML and other disorders.

"I was surprised how much more we can learn about the inner nature of these cells by ‘interrogating' them with different conditions,” said Prof. Nolan, who is also a member of the Donald E. and Delia B. Baxter Laboratory in Genetic Pharmacology at Stanford. "Time and again we are finding this to be a powerful amplifier of the fate of a diseased cell and a good way to understand why it responds to certain treatments and not others.”

The study was published in the October 7, 2008 issue of Cancer Cell.

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Stanford University School of Medicine