Inhibition of matrix metalloproteinase-2 expression and bladder carcinoma metastasis by halofuginone

Matrix metalloproteinase-2 (MMP-2) plays a critical role in tumor cell inva sion and metastasis, Inhibitors of this enzyme effectively suppress tumor m etastasis in experimental animals and are currently being tested in clinica l trials. MMP-2 transcriptional regulation is a part of a delicate balance between the expression of various extracellular matrix (ECM) constituents a nd ECM degrading enzymes, Halofuginone, a low-molecular-weight quinazolinon e alkaloid, is a potent inhibitor of collagen type alpha 1 (I) gene express ion and ECM deposition. We now report that expression of the MMP-2 gene by murine (MBT2-t50) and human (5637) bladder carcinoma cells is highly suscep tible to inhibition by halofuginone, Fifty percent inhibition was obtained in the presence of as little as 50 ng/ml halofuginone, This inhibition is d ue to an effect of halofuginone on the activity of the MMP-2 promoter, as i ndicated by a pronounced suppression of chloramphenicol acetyltransferase a ctivity driven by the MMP-2 promoter in transfected MBT2 cells. There was n o effect on chloramphenicol acetyltransferase activity driven by SV40 promo ter in these cells, Halofuginone-treated cells failed to invade through rec onstituted basement-membrane (Matrigel) coated filters, in accordance with the inhibition of MMP-2 gene expression, A marked reduction (80-90%) in the lung colonization of MBT2 bladder carcinoma cells was obtained after the i .v. inoculation of halofuginone-treated cells as compared with the high met astatic activity exhibited by control untreated cells. Under the same condi tions, there was almost no effect of halofuginone on the rate of MBT2 cell proliferation. These results indicate that the potent antimetastatic activi ty of halofuginone is due primarily to a transcriptional suppression of the MMP-2 gene, which results in a decreased enigmatic activity, matrix degrad ation, and tumor cell extravasation. This is the first description, to our knowledge, of a drug that inhibits experimental metastasis through the inhi bition of MMP-2 at the transcriptional level. Combined with its known inhib itory effect on collagen synthesis and ECM deposition, halofuginone is expe cted to exert a profound anticancerous effect by inhibiting both the primar y tumor stromal support and metastatic spread.