Antiestrogen Therapy: Uncertainties and Risk Assessment

The incidence of breast cancer shows annual increases in the United Kingdom[1] and its frequency in the United States is much higher than, for example, Thailand or Japan.[2] The antiestrogenic drug, tamoxifen(Drug information on tamoxifen), ((Z)-2-[4-(1,2-diphenyl-1-butenyl)-phenoxy]-N,N-dimethylethanamine) (Figure 1) is a drug of choice used in adjuvant therapy for breast cancer. Women with negative lymph nodes and estrogen receptor-positive tumors, show a survival advantage that lasts for at least 5 years during tamoxifen therapy.[3] In the United States and United Kingdom, the usual human dose is 20 mg daily, although in the rest of Europe, dosages of 20, 30, or 40 mg have been used. This variation in the dosage of tamoxifen can make comparisons between the success of treatments and adverse effects between European countries and the United States more difficult.

A meta-analysis of trials of tamoxifen in breast cancer patients involving approximately 75,000 individuals clearly shows improved disease-free survival times and an approximate 39% reduction in the incidence of contralateral primary breast cancer.[4] This suggested that tamoxifen could have a role in the prevention of breast cancer. As adjuvant therapy, it has few serious undesirable side effects [5]. Epidemiological evidence indicates, however, that long-term administration of tamoxifen to breast cancer patients results in an increase in the incidence of endometrial tumors [reviewed in ref 6]. After an initial report in 1985 of an association between the treatment of breast cancer patients with tamoxifen, and an increase in endometrial cancer,[7] there have been a number of case and cohort studies to support the view that tamoxifen treatment and endometrial cancer are causally related. In 1994, the National Surgical Adjuvant Breast and Bowel Project (NSABP B-14) in Canada and the United States[8] reported on the rates of endometrial and other cancers in 2,843 breast cancer patients randomly assigned tamoxifen (20 mg/day) or a placebo. The average time of study was 5 to 8 years. Over this period, 23 endometrial tumors were found in the tamoxifen treatment groups and two in the placebo group. It was concluded in this study that the average hazard rate through followup was 1.6 per thousand in the tamoxifen group versus 0.2 per thousand in the placebo controls. A year later these findings were confirmed by the results of the Stockholm breast cancer study group reported by Rutqvist et al.[9] In this trial, 2,729 breast cancer patients under the age of 71 years were given tamoxifen (40 mg daily) or a placebo. The median follow up time was 9 years and showed tamoxifen treatment resulted in nearly a six-fold increase in endometrial cancer.

The effects of tamoxifen on the endometrium will have marked consequences on the risk-benefit analysis for the putative chemopreventive treatment of healthy women. In order to substantiate these data, analysis of tumor registries in the US involving a much larger cohort of 87,323 women with breast cancer has been investigated by the Surveillance Epidemiology and End Results (SEER) program.[10] Of these, 14,358 women received hormones (tamoxifen) for their first course of therapy. Epidemiological results indicated an increase in uterine cancers in this group. As with the earlier studies, the follow-up time for the treated patients is generally less than 10 years, limiting the evaluation of long-term effects of this drug.

In view of the long time period often needed for the development of tumors in humans, it is perhaps surprising that in the case of endometrial tumors, the onset is rapid, often within 5 years from the time from start of tamoxifen. There is a strong association between estrogen replacement therapy and endometrial cancer, which is also seen in the first 5 years of initiation of treatment,[11] suggesting an estrogen promoting-like action of tamoxifen. In the Stockholm trial, however, there was a marked time dependent increase in the cumulative incidence of endometrial tumors in treated women over a longer 15-year period.[9]

In the Stockholm study, there was about a three-fold increase in gastrointestinal cancers in the tamoxifen treated patients.[9] To our knowledge, this is the only study that has shown an increase in GI tract tumors in a tamoxifen treated population. This was not detected in either the SEER or the NSABP trials and none of these studies found any significant increase in liver cancers amongst the tamoxifen treated groups.

A number of other trials, results for which were published prior to the Stockholm Breast Cancer Study Group or NSABP findings and which used smaller numbers of breast cancer patients, failed to find any increase in endometrial cancers. Examples include the Nolvadex Adjuvant Trial Organization (NATO) involving 564 patients treated with tamoxifen[12] and the Scottish trial with 374 treated women (both 20 mg/day).[13] These studies had follow-up times of 5 or more years. While the NATO trial failed to find endometrial cancers, three uterine sarcomas were reported in the Scottish tamoxifen treated group. The numbers of patients taking part were not sufficiently large to give the analytical power of the US and Swedish investigations. It appears that it is women of less than 50 years of age who are more susceptible to tamoxifen-related endometrial tumors than women over the age of 50, who have a relative risk only slightly above that of untreated women. From the epidemiological evidence, it was concluded by the International Agency for Research on Cancer (IARC) that there was sufficient evidence for the association of tamoxifen therapy and endometrial tumors in breast cancer patients, leading to classification of this drug as a Class 1 human carcinogen.[6]

Effects of Toremifene(Drug information on toremifene) on the Human Endometrium

The chlorinated structural analogue of tamoxifen, toremifene (Figure 1) is under investigation in breast cancer patients in a number of Phase III trials using daily doses of 60 to 240 mg.[14] Toremifene was not yet registered in many countries when these trials were initiated, and the number of individuals being given this drug is small, relative to the SEER and NSABP studies. Although toremifene is not genotoxic or carcinogenic in the rat liver assay (see below), the numbers of patients in the Phase III studies are presently too small to make realistic assessments of the ability of these compounds to result in endometrial tumors in women. With larger numbers of participants, epidemiological results should give a fascinating insight into the probable mechanisms of endometrial tumors caused by tamoxifen. (Note: Toremifene is now registered in most countries.)

Cellular and Molecular Actions

At the cellular level, the actions of tamoxifen are not completely understood. In breast tissue, the antiproliferative effect of the drug is primarily mediated through inhibition of estrogen activities by binding to the estrogen receptor. Tamoxifen allows dimerization of the receptor to occur but classically blocks transcriptional activation. Activation of the estrogen receptor is dependent on the differential action of the N- and C-terminal transcription activation functions AF-1 and AF2, respectively. Tamoxifen inhibits AF2 activity and functions as an estrogen antagonist where AF2 is required. However, AF2 is not required for all promoters, allowing this drug to demonstrate partial estrogen agonist activity.[15] Recently, a second estrogen receptor gene ER-b has been reported [16]. ER-b binds to DNA and can dimerize with ER-a. This adds further to the degree of complexity to transcription activation/inhibition in response to tamoxifen. Apart from these actions, tamoxifen can stimulate the paracrine secretion of transforming growth factor b and insulin-like growth factor 1 from stromal cells as well as inhibiting protein kinase C and calmodulin-dependent cAMP phosphodiesterase.[17] In human endometrial tissues tamoxifen is thought to act as a estrogen agonist, inducing cell proliferation and thus promoting any endogenous DNA lesions.

Liver Carcinogenicity in Rats

In rats, several lifetime bioassays of tamoxifen have shown that this drug results in the formation of liver carcinomas but not endometrial tumors.[18,19] With respect to liver tumors, there appears to be little difference in the susceptibility between male or female animals to this drug. Mice given similar doses of tamoxifen do not get such tumors.[20] There was a need to establish the mechanism for the formation of the liver neoplasms in the rat and to establish whether these factors operate in human liver or reproductive tissues, in order to permit better risk-benefit analysis for the many thousands of healthy women who are taking this drug in the current chemopreventative trials.

Liver DNA Damage Following Exposure of Rats or Mice to Tamoxifen

Using the ³²P-postlabeling technique, it was shown that DNA adducts were formed in the livers of two rodent species following tamoxifen administration.[21,22] The level of such DNA damage in mouse liver was about one-quarter to one-third of that in rats. Following tamoxifen treatment in rats, a DNA adduct pattern consisting of approximately 12 individual spots was consistently detected in livers of three different strains.[23] Using the postlabeling assay, no adducts could be detected in the rat liver at doses less than 5 mg/kg/day given for 7 days. Using the newly developed and highly sensitive technique of accelerator mass spectrometry, however, the binding of [¹⁴C]tamoxifen to rat liver DNA has been shown following a single dose of 0.3 mg/kg, comparable to the human therapeutic dose.[24] It should be noted, however, that because of the very much shorter half-life of tamoxifen in rodents, rats must be given ~20 mg/kg daily to achieve similar plasma levels (~250 ng/mL) as women taking this drug therapeutically.