5 December 1996 UROT96\THYMOSIN.MS-- 800 words
New protein causes step in metastasis;
thymosin beta-15 assay differentiates middle Gleason grades
With new assays for prostate cancer, you can see more than just changes in glandular architecture. You can actually see oncogenes turning on and off.
"Eventually we will have a panel of metastatic correlates for prostate cancer, maybe 6 or 8 molecules, whose expression is up- or downregulated as the tumors become capable of metastasis," said Bruce R. Zetter, Ph.D., Professor, Departments of Cell Biology and Surgery, Harvard Medical School.
"And by screening tissue against that panel, we may be able to get a good indication of the metastatic potential of any individual's prostate cancer," said Dr. Zetter.
Dr. Zetter just discovered one of those oncogenes, one that is responsible for the motility, and therefore metastasis, of prostate cancer cells. The gene produces a protein called thymosin beta-15.
Thymosin beta-15 could be a useful marker to distinguish aggressive tumors, particularly in the intermediate range of Gleason 5 to 7, according to Zetter. It could also be a therapeutic target.
Dr. Zetter compared metastatic and non-metastatic cell lines of Dunning rat prostate carcinoma, using standard molecular biology techniques to compare messenger RNA.
The difference turned out to be thymosin beta-15, which was expressed only in the metastatic lines. When he blocked the synthesis of thymosin beta-15 (by inserting anti-sense DNA), he could turn off the cell's mobility, and therefore its metastasis. He found the identical protein not only in rats but in humans, not only in cell culture but in pathology samples.
Thymosin beta-15 binds to actin, "but we don't understand that process too well," said Dr. Zetter.
"Thymosin beta-15 directly regulates cell motility in prostate carcinoma cells," he wrote in Nature Medicine (December 1996; 2(12): p. 1322-1328).
Then he found the identical protein in human prostate cancers--both in cell lines and pathology specimens.
One human prostate cancer cell line, PC-3ML, is highly metastatic, but a variant, PC-3, is not. Sure enough, the metastatic variant expressed thymosin beta-15, while the less aggressive line did not, when he tested them with reverse transcriptase-polymerase chain reaction (RT-PCR).
Then he examined pathology specimens with RNA probes and antibody stains.
Some glands with prostatic intraepithelial neoplasia expressed thymosin beta-15 mRNA, while others did not. The highly invasive tumors were consistently positive. Most of the negative cells were in noninvasive intraepithelial neoplasias. Only one negative specimen was as high as Gleason 8. "As a tumor progresses up the Gleason grading scale, this motility gene is more likely to be turned on," said Dr. Zetter. "By the time a cell is motile and metastatic, we should be able to identify it with this probe," unless there's another motility mechanism that can substitute for it, he said.
Specific tumor cell islands were strongly stained with RNA probes, while the stroma within and around the tumor cell masses, and the nonmalignant prostatic epithelium adjacent to the tumor, were not. "So we could watch the cells changing to a more motile phenotype," said Dr. Zetter.
Finally he created an antibody stain to thymosin beta-15, and examined 76 human prostate carcinoma specimens with a range of Gleason scores. Once again, the cytoplasm of the carcinoma cells in neoplastic prostates stained positive, but the histologically benign areas, and the stromal cells, did not. Benign prostatic hyperplasia specimens were negative.
There was a general correlation between thymosin beta-15 staining and Gleason score. Poorly differentiated adenocarcinomas with Gleason scores of 8-10 displayed the most extensive and intense thymosin beta-15 staining, followed by moderately differentiated prostate carcinomas with Gleason scores of 6-7. In poorly differentiated and invasive prostate carcinoma, single cells invading stroma displayed intense staining. "We've been able to see invasion before, but now we can see the genes that are turned on in these invading cells."
The great hope is that thymosin beta-15 will predict metastasis more precisely than Gleason grades alone. In the published study, Dr. Zetter knew the Gleason grade of the specimens, but not the clinical outcome. "We are right now doing a retrospective study," to see if thymosin beta-15 staining can predict clinical outcome, he said.
If one oncogene predicts outcome, two oncogenes may predict it better. "KAI-1 is the flip side of this story," said Dr. Zetter. "KAI-1 is a motility suppressor. It is present in the early stages of prostate cancer but drops out with progression. A cell that is upregulated in thymosin beta-15 and downregulated in the motility suppressor KAI-1 is going to be doubly motile," he said.
In preliminary studies, Zetter also found evidence of thymosin beta-15 in highly metastatic human breast cancer and melanoma cell lines.