Radiotherapy--the use of x-rays and other ionizing radiation to treat cancer--involves a deadly compromise. Bombarding a tumor with cell-killing beams inevitably affects healthy tissue in the tumor's vicinity.
Mary Helen Barcellos-Hoff, a biologist in LBL's Life Sciences Division, has taken an important step toward reducing complications in cancer patients after radiotherapy. She has shown that ionizing radiation, such as that used in cancer treatment, activates transforming growth factor-beta (TGFb), a cell-signal molecule known for its role in scarring.
Her study provides the strongest evidence yet that the molecule is the cause of radiogenic fibrosis, a hardening of tissue that is common following radiotherapy. "We think TGFb sows the seeds that determine whether or not irradiated tissue undergoes permanent dysfunction," Barcellos-Hoff said.
More importantly, the research suggests a new way of improving radiotherapy. Blocking TGFb activation could decrease side effects, and provide the basis for more aggressive radiotherapy and better tumor control.
Typically one-fifth of cancer patients receiving radiotherapy suffer some degree of fibrosis in the tissue surrounding a tumor. Fibrosis is the result of an overproduction of extracellular matrix (ECM), the fibrous network of molecules between cells that determines structure and regulates function of tissues. In fibrosis, tissues gradually lose their elasticity as ECM fills up the space between cells.
The dangers of fibrosis depend on where it occurs. In the breast, it can cause loss of mobility; in lungs, it can limit breathing and the ability to cough up infectious material; in the kidney it can restrict fluid flow and lead to infection.
Scientists believe that TGFb's link to fibrosis has to do with its regulating the production and remodeling of the ECM. One of TGFb's best understood roles in the body is coordinating the production of ECM to heal wounds. Fibrosis in some ways is like wound healing gone haywire, with TGFb converting healthy tissue to tough tissue similar to that seen in scars.
However, finding how TGFb might trigger fibrosis has been a challenge. The stumbling block has been the two-faced nature of the molecule. Most growth factors and other cell signals, such as hormones, affect surrounding tissue immediately after release. In contrast, TGFb must be activated--it enters the surrounding tissue in a latent form, bound to a larger carrier molecule known as latency-associated peptide.
Scientists have known for years that in order to have a biological effect on cells, TGFb somehow has to be released from the carrier peptide. Unfortunately, they have had no way to distinguish whether TGFb in tissue is latent or active, and thereby investigate what triggers activation. "All the biochemical methods to measure it actually caused activation," Barcellos-Hoff said.
To look at the effect of radiation on TGFb activation, Barcellos-Hoff had to create a better test. First, she obtained genetically engineered cells, provided by collaborators at UCSF, that secreted either the latent or active form of the molecule. She was able to grow the cells as tumor tissue, then use the tissue to select for antibodies that would bind to one or the other form of TGFb.
By adding the antibodies to tissue, then using fluorescence to highlight the antibodies, Barcellos-Hoff could look at the distribution of the two types of TGFb. She used the test on mouse mammary tissue before and after therapy-level doses of radiation.
The tests showed that radiation had a dramatic effect on TGFb. Levels of the latent form, which were high in the tissue to begin with, dropped sharply within an hour of the treatment. Levels of the active form, which were nonexistent at first, increased. Active TGFb remained high for at least a week following radiation exposure.
Activation also varied with the amount of radiation--the higher the dose, the more TGFb was activated. Barcellos-Hoff found restructuring of the ECM following the treatments, suggesting that activated TGFb was already beginning to influence cells in the irradiated tissue.
Barcellos-Hoff is now examining how molecules such as antibodies could block the effects of TGFb prior to radiation treatment. She is also looking at ways by which radiation might trigger activation.
"I think a problem with TGFb research has been that people have had to lump apples with oranges--latent TGFb with active TGFb," Barcellos-Hoff said. "Being able to discriminate between the two types should clarify the role of TGFb."
In the long run, being able to pinpoint active versus latent TGFb should help scientists understand the sometimes contradictory results they have obtained in TGFb studies. Cancer research, for instance, indicates that TGFb can be both a positive and negative indicator of the disease.