Persistent Metabolic Effects of Tamoxifen: Considerations for an Experimental Tool and Clinical Breast Cancer Treatment.

The selective estrogen receptor (ER) modulator tamoxifen is frequently used in preclinical studies to induce Cre recombinase and generate conditional transgenic mice. In addition, it is often prescribed to treat ER-positive breast cancer, which is diagnosed in approximately 150 000 people each year. In mice, protocols to activate Cre-ER transgenes require tamoxifen administration by several methods, including oral gavage, IP injection, or intragastric injection, spanning a wide range of doses to achieve transgene induction. As a result, the reported metabolic effects of tamoxifen treatment are not always consistent with anecdotal reports from breast cancer patients, or with expected outcomes based on the overall metabolically protective role of estrogen. A greater awareness of tamoxifen's adverse metabolic effects is critical to designing studies with appropriate controls, especially those investigations focused on metabolic outcomes.

In the article “Tamoxifen Treatment in the Neonatal Period Affects Glucose Homeostasis in Adult Mice in a Sex-dependent Manner,” the authors evaluated metabolic effects of a single dose of tamoxifen given to postnatal day 1 mouse pups (1). Males and females received an intragastric injection of tamoxifen in oil vehicle or oil alone. At 6 weeks of age, mice were placed on a high-fat (36.1%)/high-sucrose diet and monitored for 2 months. Remarkably, this single tamoxifen exposure in the neonatal period led to persistent adverse metabolic effects in high-fat/high-sucrose–fed females but not males. By 10 weeks of age, neonatal tamoxifen-treated (NTT) males had significantly lower body mass due to lower lean mass as compared to vehicle-treated controls. Glucose tolerance and fasting glucose in NTT males were not different from controls, and insulin tolerance was relatively improved. Energy expenditure was slightly, but significantly, lower in NTT vs control males, consistent with a lower body mass. Physical activity at 10 weeks of age and cumulative food intake for the duration of the study did not differ between groups. Lower lean mass in NTT males was attributed in part to lighter tibia weight compared to controls. In contrast to the males, female NTT mice at 10 weeks of age had greater body mass, elevated fat mass, and lower lean mass compared to vehicle-treated controls. Glucose and insulin tolerance were impaired, and both fasting glucose and insulin were significantly elevated in female NTT vs controls. Despite the difference in body mass, physical activity and energy expenditure were lower in NTT compared to control females. The mechanisms responsible for these whole-body effects warrant further investigation. ER isoforms are expressed in cells throughout the body, including the muscle, adipose, liver, and brain. Each of these organs continues to grow after birth and participates in complex regulation of whole-body metabolism throughout life. Thus, a single exposure to tamoxifen could drive sex-dependent metabolic dysfunction through multiple coordinated or independent effects.

The understanding of how tamoxifen impacts the different metabolic tissues requires thorough analysis of the roles of estrogens and ER in energy balance and metabolic homeostasis. In mice, genomic deletion of ERα is linked to increased adiposity and impaired metabolic function in both males and females (2). Supplementing ovariectomized female mice with estradiol reduces adiposity and improves glucose and insulin tolerance almost certainly through ERα-mediated mechanisms. ERα in both liver and muscle plays a critical role in modulating systemic glucose homeostasis, insulin production, and ectopic lipid deposition in male and female mice (3). In adipose tissue, ERα ablation from mature white or brown adipocytes associated with greater fat mass in part through alteration in mitochondrial function (4); however, some adipose-specific Cre models are fraught with off-target gene expression that elevates estrogen production and confounds analysis of metabolism (5). While there are scores of published studies using various approaches to reduce ER isoforms in different metabolic tissues, it is still incompletely understood how canonical estrogen signaling, and therefore tamoxifen-mediated disruption of this signaling, influences energy balance and metabolism at the cellular level.

High doses of tamoxifen (25-300 mg/kg body weight/day) are frequently used to alter transgene expression, but the true metabolic effects of tamoxifen are difficult to interpret. Many studies have been done only in male mice and report effects such as adipose tissue atrophy and browning, as well as hepatic steatosis following acute exposure to supraphysiological tamoxifen levels (6,7). More recent studies, including that by Estrada-Meza et al (1), have attempted to address this gap. For example, administration of low-dose tamoxifen to female mice decreased physical activity through effects on ERα in the brain and resulted in thicker femurs compared to vehicle-treated controls (8). Importantly, outcomes of studies using tamoxifen doses that are relevant to humans may more closely resemble effects reported in breast cancer patients, such as weight gain, hepatic steatosis, and a tendency toward type 2 diabetes, suggesting the importance of revisiting prior work with a critical look at experimental design.

There are still many unanswered questions about how tamoxifen affects metabolism in both the short and long terms. For example, it is not clear whether, under physiologically relevant conditions, tamoxifen acts as an antagonist or a partial agonist of ER in metabolic cells and tissues. This requires consideration for the dose and route of tamoxifen exposure, individual age and sex, and whether or not opposing estrogens are present. Furthermore, the duration of tamoxifen effects both in experimental models and in breast cancer patients is unknown. This is important not only for interpreting studies that use tamoxifen-driven conditional gene deletion but also because many people live for several decades after discontinuing breast cancer treatment and endure adverse changes in metabolism. With continued improvements of our preclinical models, we will be better equipped to target chronic diseases that result from disrupted estrogen signaling.