"The transition from a normal to a cancer cell takes many steps," Michael Green told BioWorld Today. But for chronic myelogenous leukemia, one major step is the generation of the tyrosine kinase Bcr-Abl, a constantly active kinase.

In the Dec. 29, 2005, issue of Cell, Green and his colleagues from the Howard Hughes Medical Institute and University of Massachusetts Medical School in Worcester, reported on one molecular mechanism by which Bcr-Abl contributes to leukemia. The experiments are part of broader work teasing apart the complex interplay between iron, gene expression, cell death and a secreted protein known as 24p3 and its receptor.

24p3 is a known pro-apoptotic protein, and the scientists wanted to investigate the relationship between 24p3 and Bcr-Abl. "The simplest model that we imagined was that Bcr-Abl was preventing 24p3 from being expressed," said Green, professor of molecular medicine and senior author of the paper. "That turned out to be 180 degrees wrong."

Instead, Bcr-Abl up-regulates 24p3, which should have the opposite effect of the extended life span that leukemia cells have. "These cells should be dead," Green said.

The reason they are not is that Bcr-Abl, by an unknown mechanism, also prevents the expression of the 24p3 receptor. That renders the cancerous cells resistant to their own 24p3 and changes the balance of power between leukemia cells and their normal neighbors.

"Any normal cell in the vicinity gets killed because it has the 24p3 receptor," Green said.

Novartis AG's chronic myeloid leukemia drug, Gleevec (imatinib), blocks Bcr-Abl activity. As a result, several biotechnology companies, including New York-based Innovive Pharmaceuticals Inc., as well as Structural GenomiX Inc. and Ambit Biosciences Inc., both of San Diego, are working on treatments for Gleevec-resistant CML. There also is Breakthrough Therapeutics, of Greenwich, Conn. (See BioWorld Today, Dec. 9, 2004, and Jan. 6, 2006.)

In the experiments presented in Cell, Gleevec increased the expression of the 24p3 receptor in cells expressing the Bcr-Abl kinase; as a consequence, the cells no longer were protected from 24p3's apoptotic effects. Green said that "in patients with leukemia, if you were able to intervene to inhibit 24p3," - either through an antibody against circulating 24p3 or via a small molecule at the transcriptional level - "it could have a beneficial effect, particularly in instances such as acquired Gleevec resistance."

Green and his colleagues discovered 24p3 as a pro-apoptotic gene while studying how growth factor deprivation drives cells to suicide. In those studies, which were published in Science in 2001, they demonstrated that interleukin-3 deprivation leads to the up-regulation of 24p3, and that 24p3 induced apoptosis. The details of the experiments reported in Science led Green and his colleagues to believe that there must be a cellular receptor for 24p3, and the work described in Cell began with the isolation and characterization of that receptor.

At the membrane, the receptor can bind 24p3 whether 24p3 itself is bound to iron or not. In both cases, 24p3 is taken up into cells, but with opposite consequences. Once in the cell, iron-bound 24p3 will ditch its load (which actually consists of a complex of iron with a so-called siderophore) and return to the membrane empty. An empty 24p3 molecule, in contrast, will acquire an intracellular iron-siderophore complex and extrude it from the cell when it returns to the surface.

Intracellular iron levels, in turn, affect a cell's penchant for suicide, or apoptosis, in an inverse relationship; that is, the lower the intracellular iron, the higher the likelihood of apoptosis. Green said that under physiological conditions, "the weight of the evidence" favors the idea that most 24p3s are empty and thus bind intracellular iron and remove it from the cell, lowering iron levels.

Green and his colleagues investigated the exact mechanism by which iron deprivation drives cells to suicide. They were guided by the knowledge that interleukin-3 deprivation induces the pro-apoptotic protein Bim, a member of the well-known Bcl-2 family, which is already in the sights of biotechnology firms such as Berkeley Heights, N.J.-based Genta Inc. and Montreal's Gemin X Biotechnologies Inc. (See BioWorld Today, Dec 13, 2005, and Dec 30, 2005.)

Studies confirmed that "empty" 24p3 increased Bim levels; interfering with Bim levels via RNA interference reversed the effects of 24p3, as did the addition of either free iron or iron-bound 24p3, and prevented apoptosis in response to interleukin-3.

Green's lab is working on determining whether other oncogenic tyrosine kinases, like Bcr-Abl, alter the expression of 24p3 and its receptor. While he described his own space as a basic research lab, his group is "certainly interested" in the clinical applications of the discovery.

"We'd be happy to work with anyone who has a clinical interest in this," he said.