Abstract P5-12-09: A novel commercial LC-MS/MS assay for tamoxifen (TAM) and its major metabolites
Authors: Clarke NJ , Weber DM , Goldman SM , Morton JW , Lagier RJ , Birse CE , Bender RA , Waldman FM 
Published: May 2016

Abstract
The standard of care for women presenting with early stage- ER-positive breast cancer (BC) following "curative" surgery has been 5 years of TAM. Adjuvant treatment with TAM has changed the natural history of BC, producing a significant reduction in 5- and 10-year recurrence rates; however, because of its adverse effects, many women (approx. 40%) do not complete the recommended 5 years of treatment. Furthermore, since TAM is a pro-drug that needs to be converted to endoxifen to be effective, inter-individual variability in endogenous enzymatic activity (i.e., CYP2D6) can affect endoxifen exposure. Certain drugs (e.g., SSRIs) can also reduce endoxifen exposure by inhibiting CYP2D6.
It is thought that a reduction in endoxifen exposure reduces the efficacy of TAM treatment and increases recurrence risk. However, several recent studies contradict this hypothesis and suggest that a) there is more than one pathway to get to endoxifen, even in the presence of variant CYP2D6; and b) TAM may act through its other metabolites as well, not just endoxifen.
Thus, a CYP2D6 genetic test may overly simplify our understanding of TAM metabolism and prompt clinicians to draw the wrong conclusions. As such, it would seem useful to develop an assay to directly measure each patient's unique serum metabolite levels. The ability to quantitate all the major TAM metabolites would also allow researchers to assess which metabolite level(s) most closely correlates with both recurrence and toxicity, allowing individualized patient dosing. In this regard, results from the BIG1-98 study suggest that some metabolites may be more closely associated with adverse effects than others. This finding could be clinically useful when combined with outcomes data, as poor adherence to TAM may be an unrecognized reason for a number of recurrences that could potentially be avoided by therapeutic drug monitoring (TDM) using a sensitive and specific assay.
With this in mind we developed an HPLC-MS/MS method that quantitatively measures TAM and 6 of its major metabolites in a single assay. This high-throughput assay has been validated to CLIA '88 standards and is run in a high-volume commercial CLIA certified laboratory. Although some of the metabolites had been measured previously by HPLC or LC-MS/MS, this is the first assay that measures all of the major metabolites, including the newly identified norendoxifen.
Results
Serum from 100 women taking TAM at 20 mg/d for &gt; 6 months was tested using this assay, and observed ranges were calculated for this patient cohort. The observed ranges from the analysis are shown in Table 1, along with the lower limit of quantitation (LLOQ) for each analyte.
Table 1. Observed ranges and LLOQs for Tamoxifen and MetaboliteAnalyteLLOQ (ng/mL)Observed Range (ng/mL)Endoxifen0.40.93-43.19Tamoxifen1.512.5-233.1N-Desmethyl Tamoxifen1.53.0-374.04-Hydroxy Tamoxifen0.20.24-5.05N-Desmethyl 4'- Tamoxifen0.41.17-19.954'-Hydroxy Tamoxifen0.40.4-6.33Norendoxifen1.2&lt;7.3
Conclusions
A novel commercial assay has been developed for TAM and its metabolites, which for the first time allows physicians to use a TDM approach for their TAM-treated patients.
Citation Format: Clarke NJ, Weber DM, Goldman SM, Morton JW, Lagier RJ, Birse CE, Bender RA, Waldman FM. A novel commercial LC-MS/MS assay for tamoxifen (TAM) and its major metabolites. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-12-09.