A Potential Synergistic Anticancer Effect of Paclitaxel and Amifostine on Endometrial Cancer Donghai Dai, 1 Anna M. Holmes, 1 Tan Nguyen, 1 Suzy Davies, 1 Daniel P. Theele, 2 Claire Verschraegen, 3 and Kimberly K. Leslie 1 1 Reproductive Molecular Biology Laboratory, Division of Maternal-Fetal Medicine, Department of Obstetrics and Gynecology; 2 Animal Research Facility, Department of Neuroscience; and 3 Cancer Research and Treatment Center, University of New Mexico Health Sciences Center, Albuquerque, New Mexico Abstract Although paclitaxel is one of the most effective chemothera- peutic agents, its usefulness is still limited in advanced and recurrent endometrial cancer. Amifostine protection of normal tissues against the side effects of chemotherapeutic agents has been clinically proven in cancer patients; however, its application in endometrial cancer has not been fully evaluated. We have investigated the use of paclitaxel and amifostine in controlling the growth of poorly differentiated endometrial cancer cells, Hec50co, in vitro and in vivo . Our studies show that amifostine had direct anticancer effects on endometrial cancer cells in vitro by arresting the cell cycle at the G 1 phase and inducing apoptosis. Amifostine also inhibited s.c. tumor growth in athymic mice. Paclitaxel IC 50 value was reduced from 14 to 2 nmol/L with pretreatment of a single dose of 178 Mmol/L of amifostine for 72 hours. Amifostine also synergized with paclitaxel in the arrest of the cell cycle at the G 2 -M phase and in the induction of apoptosis. This two-drug regimen inhibited s.c. tumor growth as well as improved mouse survival significantly more than paclitaxel alone. Amifostine also significantly improved paclitaxel-induced cytotoxic effects on peripheral blood profiles. Our studies show that amifostine has direct anticancer effects on endometrial cancer. Our data have also shown a potential anticancer synergy between amifos- tine and paclitaxel in vitro and in vivo, whereas amifostine maintained a protective role in peripheral blood profiles. The dual specificity of amifostine action should be further investigated. (Cancer Res 2005; 65(20): 9517-24) Introduction Endometrial cancer is the fourth most common malignancy in women (1). The American Cancer Society estimated that there were 40,320 new cases and 7,090 deaths from endometrial cancer in 2004 (1). Currently, there is no effective therapy for advanced or recurrent endometrial cancer. Paclitaxel is an antimicrotubule agent which is among the most potent single chemotherapeutic agents. Paclitaxel binds to the h-tubulin subunit and stabilizes the microtubules, resulting in disruption of normal microtubule dynamics during cell division (2). Failure of microtubule separation during the G 2 -M phase blocks cell mitosis and results in apoptosis. Application of this agent in endometrial cancer treatment is under investigation. A Gyneco- logic Oncology Group trial reports an overall response rate of 35.7% in advanced or recurrent endometrial cancer (3). The response rate is about 27.3% when paclitaxel is used in patients who have failed prior chemotherapy (4). The overall survival, objective response, and progression-free survival were improved by adding paclitaxel to the doxorubicin and cisplatin regimen (5). However, the median survival of patients treated with paclitaxel-based regimens is around 12 months. Unfortunately, the benefits of paclitaxel are also associated with serious adverse effects. These commonly include myelosuppression, gastrointestinal toxicity, and peripheral neurop- athy, among others (5). It was reported that 62% of patients treated with paclitaxel had life-threatening leucopenia (3). In another study, where paclitaxel was used as a second-line chemotherapy (4), serious neutropenia occurred in more than 58% of patients with one treatment-related death. Improvement of outcome with the addition of paclitaxel to the doxorubicin and cisplatin regimen was accompanied by a much higher treatment-related death rate (5). Thus, it is imperative to search for more effective and safer chemotherapeutic regimens. Amifostine (WR2721) was developed in 1979 as an antiradiation agent through the screening of 4400 chemicals by the Walter Reed Army Institute of Research (6). Since then, its protective effects against chemotherapy and radiotherapy in cancer patients have led to amifostine being FDA-approved as a cytoprotective agent against the adverse effects of chemotherapy and radiotherapy in normal tissues. Amifostine is a prodrug, which must first be converted to its active form, WR1065, by the enzyme alkaline phosphatase, which is located on the cell membrane. WR1065 is further metabolized through intracellular oxidation to the sym- metrical disulfide WR33278, cysteamine, and mixed disulfides (7). The underlying mechanisms of amifostine protective effects are still not fully understood, but they are attributed to its ability to scavenge free radicals (8) and to its antimutagenic effects (9). This protection is based on the conversion of amifostine to the active form, WR1065, which is rapidly taken up into normal tissues (10). Amifostine dephosphorylation by membrane alkaline phosphatase occurs effectively in normal tissue, which has a suitable pH of 6.6 to 8.2 for alkaline phosphatase to work. The uptake of WR1065 is minimal in cancer cells because tumor tissue usually contains less membrane-bound alkaline phosphatase (11–14) and the environ- ment of the tumors is more acidic, making the enzymatic activity less than optimal (15). Additionally, normal tissues contain the WR1065 transporter that allows active absorption against a concentration gradient (16). Taken together, these differences lead to a concentration of WR1065 that is about 50 to 100 times higher in normal tissue than in malignant tissue (16), thereby providing normal tissues with a remarkable protection against cytotoxic therapies. Requests for reprints: Donghai Dai, Department of Obstetrics and Gynecology, University of New Mexico Health Sciences Center, MSC 10 5580, Albuquerque, NM 87131. Phone: 505-272-0095; Fax: 505-272-3921; E-mail: DDai@salud.unm.edu. I2005 American Association for Cancer Research. doi:10.1158/0008-5472.CAN-05-1613 www.aacrjournals.org 9517 Cancer Res 2005; 65: (20). October 15, 2005 Research Article Research. on November 5, 2015. © 2005 American Association for Cancer cancerres.aacrjournals.org Downloaded from