Breast cancer is still a leading cause of cancer death in women in the United States and Europe [
1]. Adjuvant chemotherapy has been shown to provide disease-free and overall survival benefits for patients with node-positive breast cancer in large meta-analyses conducted by the Early Breast Cancer Trialists' Collaborative Group [
2]. A study by Weiss
et al. [
3], however, showed that the overwhelming majority (80%) of node-positive patients relapse and die within 26 years of diagnosis despite the use of cyclophasphamide-methotrexate-5-fluorouracil (CMF)-based adjuvant chemotherapy. The development of drugs and strategies to improve relapse-free and overall survival therefore remains a high priority.
Paclitaxel (Pacl), belonging to the chemical class of taxanes, is capable of inducing
in vitro apoptosis, independently of p53 status, through its microtubule-stabilizing activity and, as recently published, by inducing the release of cathepsin B from lysosomes [
5]. From a clinical point of view, taxanes appear to be promising, although their real impact on the natural history of breast cancer has yet to be defined. A similar impact of Pacl on clinical response has been reported, however, for patients with wild-type or p53-mutated cancers [
6‐
9], the latter representing 15% of
in situ breast carcinomas and 50% of invasive disease [
10]. Moreover, the loss of normal p53 function sensitizes
in vitro cells to drug activity [
11‐
13], hence the interest to use the taxane combination with other drugs as clinical therapy. The association of a taxane with an anthracycline is based on evidence that the two drugs have different mechanisms of action and toxicities and are not cross-resistant. Moreover,
in vitro studies [
14‐
17] on human cell lines and primary breast cancer cultures have shown a schedule-dependent interaction of the two drugs. Phase I and II studies with the doxorubicin (Dox)-Pacl sequence defined by preclinical studies have reported objective responses ranging from 40% to more than 90% [
18,
19], and a recent prospective phase III study on advanced breast cancer reported an impressive response rate (up to 94%) for the sequence, which was shown to prolong overall survival compared to the fluorouracil, epirubicin, cyclophosphamide (FEC) combination [
20].
5-Fluorouracil (5-FU), one of the most important agents for the treatment of colorectal, head and neck, pancreatic and breast carcinomas, is a pro-drug that requires conversion to 5-fluoro-deoxyuridine-monophosphate (5FdUMP) and 5-fluoro-deoxyuridine-triphosphate (5FUTP) in cancer cells. Several
in vitro studies on human solid tumor cell lines have demonstrated the positive and schedule-dependent interaction of Pacl and 5-FU [
21‐
23]. A synergistic effect was obtained only when tumor cells were exposed to Pacl followed by antimetabolites. Conversely, simultaneous exposure to the two drugs or pretreatment with 5-FU reduced overall cell killing compared to Pacl alone. Specifically, it has been demonstrated that a short pretreatment with Dox increases the activity of Pacl and of the Pacl→gemcitabine sequence [
24]. In the light of these preclinical and clinical observations, the present study aimed to investigate the cytotoxic effect produced by the combination of Dox, Pacl and 5-FU in human breast cancer cell lines and to define, at the preclinical level, the most effective treatment scheme.