6. The Mechanism of GCP Activation

GCP has been found to induced significant apoptosis in cultured tumor cells as well as in normal cells in vitro. However, we found in animal experiments as well as in human monitor test, that GCP had no cytotoxic effects on normal tissues even through the blood concentration of genistein reached high levels in vivo. As described earlier, we found GCP exists mostly as glucuronide and sulfate conjugates. From this occurrence, we speculate there might be an activation process in the body that can transfer the non-active-genistein (conjugation style) into active-genistein (aglycone style).

It has been reported that tumor tissues express high amount of -glucuronidase (heparanase), an extracellular matrix degraded enzyme. This enzyme can degrade heparan sulfate and heparan sulfate proteoglycans, which are chief components of the extracellular matrix and vascular basement membrane. Therefore, heparanase is thought to be an important molecule for acceleration of cancer invasion and metastasis. The high expression of heparanase is strongly correlated with the metastasis of cancer.

To reveal our speculation that genistein conjugation can be activated in tumor tissue but not in normal tissues, we designed the following series of experiments to detect expression of -glucuronidase in local tumor tissues and in normal organ tissues, and checked whether the -glucuronidase in the tumor tissues could transfer the genistein conjugation to aglycone style.

A) Tumor cell transfer the genistein into aglycone in vitro.

We designed an in vitro system to detect the ability of cancer cells to change genistein conjugation into aglycone as compared to normal cells. The macrophage cells were obtained from the peritoneal as shown in following figure:

This result showed that when a 20% genistein conjugated serum was cultured with cancer cells, they transformed into an aglycone form before the in vitro -glucuronidase treatment, at a transformation rate of 60.69%. Macrophage cells did not make such a change (1.68% transformation).

B) Tumor tissue expresses high levels of  -glucuronidase

Studies revealed that  -glucuronidase (heparanase) was expressed significantly more in tumor tissues than in normal tissues.

C) Different existence style of genistein in tumor tissue or in normal tissue

We further measured the transformation rates of genistein-aglycone to total genistein (aglycone and conjugated forms) in tumor tissues and in normal tissues (liver and colon). The results suggest that tumor tissues might be able to transform the genistein conjugate into an aglycone form.

Summary: The results above reveal tumor tissues produce high level of heparanase which transfer genistein conjugates into genistein aglycones. The active genistein aglycone inhibits tumor growth. Genistein exerts its activity only when the conjugated glucuronide is cleaved by -glucuronidase (heparanase). This cleaving activity is mainly expressed by tumor tissue and not normal tissue. Therefore, the genistein attacks heparanase-rich tumor tissues and not normal tissues. Genistein-glucuronide exists in blood which might be the reason that GCP has no side effects.