NEW YORK (GenomeWeb News) – Using gene expression analyses, an international research team has identified molecular features that help breast cancers metastasize to the brain. The research appears online today in Nature.
The researchers used microarrays to compare the gene expression profiles of breast cancer cell lines that metastasized to the brain with those that didn't. They then checked the expression of these genes in clinically annotated breast tumors, identifying 17 genes that were differentially expressed in the tumors.
Several overlapped with genes shown to be differentially expressed in a lung metastasis signature, including the cyclooxygenase COX2 and an epidermal growth factor receptor ligand. Another gene, α2,6-sialtransferase ST6GALNAC5, which is involved in cell-cell communication, appeared to be exclusively over-expressed in brain metastases and appeared to help cells move from the blood into the brain.
"Our results draw attention to the role of the cell surface as a previously unrecognized participant in brain metastasis, and to the possibility of using drugs to disrupt its interactions," senior author Joan Massagué, a Howard Hughes Medical investigator and chair of Memorial Sloan-Kettering Cancer Center's cancer biology and genetics program, said in a statement. "Further study is necessary to explore the role of these genes in brain metastasis and their interest as therapeutic targets."
Metastasis from a primary site to other parts of the body is a factor in many cancer deaths. Whereas some metastases arise relatively shortly after cancer diagnosis, some take much longer. For example, breast cancer metastasis to the brain usually occurs several years after tumor removal.
Massagué and his colleagues hypothesized that this delay might reflect the fact that breast cancer cells need time to acquire the characteristics that provide them entry into the brain, which is protected by tight junctions between cells and a blood-brain-barrier.
In an effort to characterize the molecular features that eventually help breast cancer cells access the brain, the researchers first developed cells with a propensity for such metastasis.
They injected two different breast cancer cells — an ER- pleural malignant breast cancer cell line called CN34 and a metastatic ER- pleural breast cancer cell line called MDA231 — into mice and then isolated the populations that metastasized to the brain, using these to develop a brain metastatic line that they called BrM2.
The team first compared gene expression in BrM2 and parental cell lines with the Affymetrix HG-U133A gene expression chip, coming up with 243 genes that were either over- or under-expressed in the brain metastasis cell line.
When the researchers looked at the expression of these genes in 368 breast tumors that were linked to breast cancer relapse and brain metastasis using Affymetrix HG-U133A and HG-U133 plus 2.0 and Agilent microarrays, they found 17 genes differentially expressed in the brain relapse samples.
Of these, six overlapped with genes found in an 18-gene lung metastasis signature identified in 2007, including COX2, MMP1, ANGPTL4, LTBP1, FSCN1, and RARRES3. In addition, both signatures contained EGFR-binding molecules, though the identity differed between the brain and lung metastasis signatures.
Subsequent experiments in mice and human cell lines supported the notion that COX2 and the EGFR-pathway are involved in brain and lung metastases, possibly helping to promote cellular movement and invasiveness. Even so, the researchers speculated that breast cancer takes on additional features in order to specifically access the brain.
They found 26 candidate genes that were more highly expressed in breast cancer metastases to the brain than those to bone or lung. Among them: ST6GALNAC5, a gene that's usually expressed only in brain cells in mice and humans and codes for a α2,6-sialtransferase — an enzyme that modifies cell surface molecules and influences cell-cell interactions.
Consistent with its expression in brain metastatic cells, the researchers found that they could detect sialyl-related cell surface modifications in brain metastatic cell lines using staining techniques. And they found that nearly a quarter of brain metastases tested had ST6GALNAC5 messenger RNA levels resembling those in BrM2 cells.
Based on these results, the team concluded that ST6GALNAC5 expression specifically helps cancer cells enter the brain. Even so, they noted that additional factors are likely required for cancer expansion in the brain.
"These findings draw attention to the role of cell-surface sialylation as a previously unrecognized participant in brain metastasis, and to the possibility of therapeutically disrupting these interactions," Massagué and his co-authors wrote. "Our work also points to other candidate genes implicated in vascular permeability and leukocyte infiltration during brain inflammatory processes, and genes implicated in neurite extension and astrocyte cell processes."