IN-VIVO TOXICITY, PHARMACOKINETICS, AND ANTICANCER ACTIVITY OF GENISTEIN LINKED TO RECOMBINANT HUMAN EPIDERMAL GROWTH-FACTOR

Epidermal growth factor receptor (EGFR)-associated protein tyrosine ki nase (PTK) complexes have vital anti-apoptotic functions in human brea st cancer cells. We have shown previously that targeting the naturally occurring PTK inhibitor genistein to the EGFR-associated PTK complexe s using the EGF-Genistein (Gen) conjugate triggers rapid apoptotic cel l death in human breast cancer cells and abrogates their in vitro clon ogenic growth. In the present study, we examined the in vivo toxicity profile, pharmacokinetics, and anticancer activity of EGF-Gen, No toxi cities were observed in mice treated with EGF-Gen at dose levels as hi gh as 40 mg/kg administered i.p. as a single dose or 140 mg/kg adminis tered i.p. over 28 consecutive days. EGF-Gen significantly improved tu mor-free survival in a severe combined immune deficiency (SCID) mouse xenograft model of human breast cancer, when it was administered 24 h after inoculation of tumor cells. At 100 mu g/kg/day x 10 days (1 mg/k g total dose), which is >100-fold less than the highest tested and non toxic cumulative dose (i.e., 140 mg/kg) in mice, EGF-Gen was more effe ctive than cyclophosphamide (50 mg/kg/day x 2 days), Adriamycin (2.5 m g/kg x 1 day), or methotrexate (0.5 mg/kg x 1 day), the most widely us ed standard chemotherapeutic drugs for breast cancer, and resulted in 60% long-term tumor-free survival. Furthermore, treating SCID mice wit h established s.c. human breast cancer xenografts of 0.5-cm diameter w ith EGF-Gen at this dose level resulted in disappearance of the tumors in two of five mice and >50% shrinkage in three of five mice within 1 0 days, whereas all of the control tumors in five PBS-treated mice as well as five mice treated with unconjugated Gen (1 mg/kg/day x 10 days ) showed >200% increase in diameter during the same observation period . EGF-Gen treatment reduced the growth rate of breast cancer xenograft s of 1.0-cm diameter, but unlike with tumors of 0.5-cm diameter, it fa iled to cause shrinkage or disappearance of these larger tumors. The l evel of EGF-Gen systemic exposure that was effective in SCID mice was achieved in cynomolgus monkeys without any significant side effects de tectable by clinical observation, laboratory studies, or histopatholog ical examination of multiple organs. EGF-Gen might be useful in the tr eatment of breast cancer as well as other EGFR-positive malignancies.